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Sample records for detecting chromosomal alterations

  1. Detection of chromosomal alterations in the circulation of cancer patients with whole-genome sequencing.

    PubMed

    Leary, Rebecca J; Sausen, Mark; Kinde, Isaac; Papadopoulos, Nickolas; Carpten, John D; Craig, David; O'Shaughnessy, Joyce; Kinzler, Kenneth W; Parmigiani, Giovanni; Vogelstein, Bert; Diaz, Luis A; Velculescu, Victor E

    2012-11-28

    Clinical management of cancer patients could be improved through the development of noninvasive approaches for the detection of incipient, residual, and recurrent tumors. We describe an approach to directly identify tumor-derived chromosomal alterations through analysis of circulating cell-free DNA from cancer patients. Whole-genome analyses of DNA from the plasma of 10 colorectal and breast cancer patients and 10 healthy individuals with massively parallel sequencing identified, in all patients, structural alterations that were not present in plasma DNA from healthy subjects. Detected alterations comprised chromosomal copy number changes and rearrangements, including amplification of cancer driver genes such as ERBB2 and CDK6. The level of circulating tumor DNA in the cancer patients ranged from 1.4 to 47.9%. The sensitivity and specificity of this approach are dependent on the amount of sequence data obtained and are derived from the fact that most cancers harbor multiple chromosomal alterations, each of which is unlikely to be present in normal cells. Given that chromosomal abnormalities are present in nearly all human cancers, this approach represents a useful method for the noninvasive detection of human tumors that is not dependent on the availability of tumor biopsies.

  2. Detection of Chromosomal Structural Alterations in Single Cells by SNP Arrays: A Systematic Survey of Amplification Bias and Optimized Workflow

    PubMed Central

    Iwamoto, Kazuya; Bundo, Miki; Ueda, Junko; Nakano, Yoko; Ukai, Wataru; Hashimoto, Eri; Saito, Toshikazu; Kato, Tadafumi

    2007-01-01

    Background In single-cell human genome analysis using whole-genome amplified product, a strong amplification bias involving allele dropout and preferential amplification hampers the quality of results. Using an oligonucleotide single nucleotide polymorphism (SNP) array, we systematically examined the nature of this amplification bias, including frequency, degree, and preference for genomic location, and we assessed the effects of this amplification bias on subsequent genotype and chromosomal copy number analyses. Methodology/Principal Findings We found a large variability in amplification bias among the amplified products obtained by multiple displacement amplification (MDA), and this bias had a severe effect on the genotype and chromosomal copy number analyses. We established optimal experimental conditions for pre-screening for high-quality amplified products, processing array data, and analyzing chromosomal structural alterations. Using this optimized protocol, we successfully detected previously unidentified chromosomal structural alterations in single cells from a lymphoblastoid cell line. These alterations were subsequently confirmed by karyotype analysis. In addition, we successfully obtained reproducible chromosomal copy number profiles of single cells from the cell line with a complex karyotype, indicating the applicability and potential of our optimized workflow. Conclusions/Significance Our results suggest that the quality of amplification products should be critically assessed before using them for genomic analyses. The method of MDA-based whole-genome amplification followed by SNP array analysis described here will be useful for exploring chromosomal alterations in single cells. PMID:18074030

  3. Numeric alterations in chromosomes 7 and 8 detected by fluorescent in situ hybridization correlate with high-grade localized prostate cancer.

    PubMed

    Barranco, M A; Alcaraz, A; Corral, J M; Solé, M; Mallofré, C; Llopis, J; Rodríguez, A; Ribal, M J; Alvarez-Vijande, R; Carretero, P

    1998-01-01

    To compare the ability of flow cytometry (FCM) and fluorescent in situ hybridization (FISH), using a small set of 4 enumeration chromosome probes to detect aneuploidy in prostate tumors, and to correlate it with histological grade and pathological stage. Among 28 suitable cases, 21 could be analyzed by FISH and FCM techniques. DNA centromeric probes were used in FISH analysis to enumerate chromosomes 7, 8, 10 and 12. (a) Of the 21 cases studied by FISH, 5 were diploid, 14 aneuploid and 2 were tetraploid. When studied by FCM, these tumors were: 14 diploid, 6 aneuploid, and 1 tetraploid. FISH proved to have a higher ability for detecting DNA aneuploidy than FCM while been equally specific, since all tumors aneuploid by FCM were also found to be aneuploid by FISH. (b) Of the 14 aneuploid tumors, 12 were of high histological grade, while only 2 of the 7 nonaneuploid were of high grade. A statistically significant association was observed between high histological grade and FISH aneuploidy (p = 0.033). (c) All the aneuploid tumors showed chromosome 7 and/or 8 aneusomy. Trisomy 7 and monosomy 8 were the most frequent alterations present in 56 and 42% of the aneuploid tumors, respectively. FISH analysis of chromosome 7 and 8 alterations proved to be more sensitive than FCM in the detection of aneuploid prostate tumors. This aneuploidy was significantly associated with a poor pathological prognosis.

  4. Study of chromosomal alterations in bovine leukosis.

    PubMed

    Predescu, E; Athanasiu, P; Nastac, E; Hozoc, M

    1977-01-01

    The results of a cytogenetic study of the "CT 384" cell line obtained from bovine leukemic lymph nodes are presented. Multiple chromosomal alterations were found in the 265 metaphases examined: numeric anomalies (aneuploidy and polyploidy), morphologic aberrations (dicentric, annular, giant, filamentous chromosomes) and chromosomal lesions (arm breaks).

  5. Detection of numerical abnormalities of chromosome 9 and p16/CDKN2A gene alterations in ovarian cancer with fish analysis.

    PubMed

    Aravidis, Christos; Panani, Anna D; Kosmaidou, Zoi; Thomakos, Nikolaos; Rodolakis, Alexandros; Antsaklis, Aristeidis

    2012-12-01

    The molecular events leading to the development of ovarian cancer are not well-established. Defects of the retinoblastoma protein (pRb)/cyclin-D1/p16 pathway have been shown to play a critical role in the development of human malignancies. In particular, the p16/cyclin-dependent kinase inhibitor 2A (CDKN2A) gene located on chromosomal region 9p21 frequently is altered in several types of cancer. To investigate both the presence of numerical abnormalities of chromosome 9 and p16 gene alterations in ovarian cancer, we studied 28 cases by the fluorescence in situ hybridization (FISH) technique using a DNA p16 probe and an a-satellite probe specific for chromosome 9. Numerical abnormalities of chromosome 9 were found in all studied cases. Polysomy 9 was detected in 10 cases while monosomy 9 in seven cases. In 11 cases, there were two cell populations, one with polysomy 9 and the other with monosomy 9. In all cases, the p16 gene deletion was observed. Among them, 25 cases presented deletion of p16 gene in 21.43%-86.3% of the examined cells. Three cases carried deletion of the p16 gene in a lower proportion (12.04%-19.49%). In five cases with p16 gene deletion, homozygous deletion was detected. Numerical aberrations of chromosome 9 and p16 gene deletion are common findings in ovarian cancer. Data suggest that the p16 gene, located in the short arms of chromosome 9, may play a role in ovarian carcinogenesis. In addition, polysomy 9 could lead to activation of a number of oncogenes, thus participating in the neoplastic process in the ovaries.

  6. Altered chromosome 6 in immortal human fibroblasts.

    PubMed

    Hubbard-Smith, K; Patsalis, P; Pardinas, J R; Jha, K K; Henderson, A S; Ozer, H L

    1992-05-01

    Human diploid fibroblasts have a limited life span in vitro, and spontaneous immortalization is an extremely rare event. We have used transformation of human diploid fibroblasts by an origin-defective simian virus 40 genome to develop series of genetically matched immortal cell lines to analyze immortalization. Comparison of a preimmortal transformant (SVtsA/HF-A) with its uncloned and cloned immortalized derivatives (AR5 and HAL) has failed to reveal any major alteration involving the simian virus 40 genome. Karyotypic analysis, however, demonstrated that all of the immortal cell lines in this series have alterations of chromosome 6 involving loss of the portion distal to 6q21. The karyotypic analysis was corroborated by DNA analyses. Southern analysis demonstrated that only one copy of three proto-oncogene loci (ros1, c-myb, and mas1) on 6q was retained in immortal cells. Polymerase chain reaction analysis of the microsatellite polymorphism at 6q22 (D6S87) showed loss of heterozygosity. In addition, elevated expression of c-myb (6q22-23) was observed. We hypothesize that the region at and/or distal to 6q21 plays a role in immortalization, consistent with the presence of a growth suppressor gene.

  7. Altered chromosome 6 in immortal human fibroblasts.

    PubMed Central

    Hubbard-Smith, K; Patsalis, P; Pardinas, J R; Jha, K K; Henderson, A S; Ozer, H L

    1992-01-01

    Human diploid fibroblasts have a limited life span in vitro, and spontaneous immortalization is an extremely rare event. We have used transformation of human diploid fibroblasts by an origin-defective simian virus 40 genome to develop series of genetically matched immortal cell lines to analyze immortalization. Comparison of a preimmortal transformant (SVtsA/HF-A) with its uncloned and cloned immortalized derivatives (AR5 and HAL) has failed to reveal any major alteration involving the simian virus 40 genome. Karyotypic analysis, however, demonstrated that all of the immortal cell lines in this series have alterations of chromosome 6 involving loss of the portion distal to 6q21. The karyotypic analysis was corroborated by DNA analyses. Southern analysis demonstrated that only one copy of three proto-oncogene loci (ros1, c-myb, and mas1) on 6q was retained in immortal cells. Polymerase chain reaction analysis of the microsatellite polymorphism at 6q22 (D6S87) showed loss of heterozygosity. In addition, elevated expression of c-myb (6q22-23) was observed. We hypothesize that the region at and/or distal to 6q21 plays a role in immortalization, consistent with the presence of a growth suppressor gene. Images PMID:1373811

  8. Alterations at chromosome 17 loci in peripheral nerve sheath tumors

    SciTech Connect

    Lothe, R.A.; Slettan, A.; Saeter, G.

    1995-01-01

    Little is known about the molecular genetic changes in malignant peripheral nerve sheath tumors (MPNST). Inactivation of the TP53 gene in l7p has been reported in a few tumors. The MPNST is one of the manifestations of neurofibromatosis 1 (NF1), suggesting that the NF1 gene in 17q might be important. We present a study of 15 neurofibromas and MPNST from nine individuals. Seven patients had NF1 and six of these developed MPNST. Genetic alterations at nine polymorphic loci on chromosome 17 were examined. Allelic imbalance was detected only in the malignant tumors from NF1 patients (4/6). Complete loss of heterozygosity of 17q loci was found in three of these tumors, all including loci within the NF1 gene. Two of the malignant tumors also showed deletions on 17p. No mutations were detected within exon 5-8 of the TP53 in any of the MPNST, and none of them were TP53 protein-positive using immunostaining with mono- and polyclonal antibodies against TP53. The numbers of chromosome 17 present in each tumor were evaluated by use of fluorescence in situ hybridization (FISH) on interphase nuclei with a centromere-specific probe. A deviation from the disomic status of chromosome 17 was observed in two of the MPNST from NF1 patients. These results support the hypothesis of inactivation of both NF1 gene alleles during development of MPNST in patients with NF1. In contrast to other reports, we did not find evidence for a homozygous mutated condition of the TP53 gene in the same tumors. Finally, FISH analysis was in accordance with the DNA analysis in the deduction of the numbers of chromosome 17 in these tumors. 29 refs., 3 figs., 2 tabs.

  9. A highly specific coding system for structural chromosomal alterations.

    PubMed

    Martínez-Frías, M L; Martínez-Fernández, M L

    2013-04-01

    The Spanish Collaborative Study of Congenital Malformations (ECEMC, from the name in Spanish) has developed a very simple and highly specific coding system for structural chromosomal alterations. Such a coding system would be of value at present due to the dramatic increase in the diagnosis of submicroscopic chromosomal deletions and duplications through molecular techniques. In summary, our new coding system allows the characterization of: (a) the type of structural anomaly; (b) the chromosome affected; (c) if the alteration affects the short or/and the long arm, and (d) if it is a non-pure dicentric, a non-pure isochromosome, or if it affects several chromosomes. We show the distribution of 276 newborn patients with these types of chromosomal alterations using their corresponding codes according to our system. We consider that our approach may be useful not only for other registries, but also for laboratories performing these studies to store their results on case series. Therefore, the aim of this article is to describe this coding system and to offer the opportunity for this coding to be applied by others. Moreover, as this is a SYSTEM, rather than a fixed code, it can be implemented with the necessary modifications to include the specific objectives of each program. Copyright © 2013 Wiley Periodicals, Inc.

  10. Altered fingerprints: analysis and detection.

    PubMed

    Yoon, Soweon; Feng, Jianjiang; Jain, Anil K

    2012-03-01

    The widespread deployment of Automated Fingerprint Identification Systems (AFIS) in law enforcement and border control applications has heightened the need for ensuring that these systems are not compromised. While several issues related to fingerprint system security have been investigated, including the use of fake fingerprints for masquerading identity, the problem of fingerprint alteration or obfuscation has received very little attention. Fingerprint obfuscation refers to the deliberate alteration of the fingerprint pattern by an individual for the purpose of masking his identity. Several cases of fingerprint obfuscation have been reported in the press. Fingerprint image quality assessment software (e.g., NFIQ) cannot always detect altered fingerprints since the implicit image quality due to alteration may not change significantly. The main contributions of this paper are: 1) compiling case studies of incidents where individuals were found to have altered their fingerprints for circumventing AFIS, 2) investigating the impact of fingerprint alteration on the accuracy of a commercial fingerprint matcher, 3) classifying the alterations into three major categories and suggesting possible countermeasures, 4) developing a technique to automatically detect altered fingerprints based on analyzing orientation field and minutiae distribution, and 5) evaluating the proposed technique and the NFIQ algorithm on a large database of altered fingerprints provided by a law enforcement agency. Experimental results show the feasibility of the proposed approach in detecting altered fingerprints and highlight the need to further pursue this problem.

  11. Alteration of the metaphase checkpoint by B chromosomes.

    PubMed

    Herron, W B; Lluesma, S M; Arana, P; Wise, D

    2004-01-01

    The B chromosome polymorphism in Spanish populations of the grasshopper, Eyprepocnemis plorans (Charpentier) is ancient and widespread. Meiocytes containing B chromosomes were analyzed in our laboratory using the 3F3/2 monoclonal antibody, which binds to a kinetochore phosphoepitope whose degree of phosphorylation is sensitive to tension applied to the kinetochore. Further, the tension created by the spindle at metaphase controls a checkpoint (the metaphase checkpoint) that allows the cell to begin anaphase when all chromosomes are aligned at the metaphase plate. Fluorescence patterns of the 3F3/2 phosphoepitope in cells containing B chromosomes were determined using confocal laser scanning microscopy. The phosphorylation pattern of kinetochores in these cells was shown to be different from that of cells without Bs. This suggests that the metaphase checkpoint has been modified in some way. We propose that B chromosomes in these grasshopper populations may have survived during evolution due to an alteration of the metaphase checkpoint, making it more permissive to the presence of misaligned chromosomes. Copyright 2004 S. Karger AG, Basel

  12. Alterations in Chromosomal Synapses and DNA Repair in Apoptotic Spermatocytes of Mus m. Domesticus

    PubMed Central

    Ayarza, E.; González, M.; López, F.; Fernández-Donoso, R.; Page, J.; Berrios, S.

    2016-01-01

    We investigated whether apoptotic spermatocytes from the mouse Mus m. domesticus presented alterations in chromosomal synapses and DNA repair. To enrich for apoptotic spermatocytes, the scrotum’s temperature was raised by partially exposing animals for 15 min to a 42ºC water bath. Spermatocytes in initial apoptosis were identified in situ by detecting activated caspase-9. SYCP1 and SYCP3 were markers for evaluating synapses or the structure of synaptonemal complexes and Rad51 and γH2AX for detecting DNA repair and chromatin remodeling. Apoptotic spermatocytes were concentrated in spermatogenic cycle stages III-IV (50.3%), XI-XII (44.1%) and IX-X (4.2%). Among apoptotic spermatocytes, 48% were in middle pachytene, 44% in metaphase and 6% in diplotene. Moreover, apoptotic spermatocytes showed several structural anomalies in autosomal bivalents, including splitting of chromosomal axes and partial asynapses between homologous chromosomes. γH2AX and Rad51 were atypically distributed during pachytene and as late as diplotene and associated with asynaptic chromatin, single chromosome axes or discontinuous chromosome axes. Among apoptotic spermatocytes at pachytene, 70% showed changes in the structure of synapses, 67% showed changes in γH2AX and Rad51 distribution and 50% shared alterations in both synapses and DNA repair. Our results showed that apoptotic spermatocytes from Mus m. domesticus contain a high frequency of alterations in chromosomal synapses and in the recruitment and distribution of DNA repair proteins. Together, these observations suggest that these alterations may have been detected by meiotic checkpoints triggering apoptosis. PMID:27349323

  13. Alterations and Chromosomal Variants in the Ecuadorian Population

    PubMed Central

    Paz-y-Miño, César; Cumbal, Nadia; Araujo, Santiago; Sánchez, Ma. Eugenia

    2012-01-01

    Medical genetics is a field marked by fast progress. Even though it was at one point confined to a group of relatively rare diseases, today it has become a central component in the understanding of disorders and it is the subject of interest for all medical specialties. This paper, shares data on the chromosomal alterations and variations that have been diagnosed in Ecuadorian patients since 1998. A total of 2,636 individual cases have been analyzed by G-banding technique until February 2012. The present work shows this collection of data and the important findings that have appeared throughout these years in hopes that it can contribute to have a deeper understanding of the incidence of chromosomal aberrations and alterations in the Ecuadorian population. PMID:23091347

  14. Chromosome 8p alterations in sporadic and BRCA2 999del5 linked breast cancer

    PubMed Central

    Sigbjornsdottir, B. I.; Ragnarsson, G.; Agnarsson, B. A.; Huiping, C.; Barkardottir, R. B.; Egilsson, V.; Ingvarsson, S.

    2000-01-01

    Chromosomal losses involving the short arm of chromosome 8 are frequent in a variety of tumour types, including breast cancer, suggesting the presence of one or more tumour suppressor genes in this region. In this study, we have used 11 microsatellite markers to analyse loss of heterozygosity (LOH) at chromosome 8p in 151 sporadic breast tumours and 50 tumours from subjects carrying the BRCA2 999del5 mutation. Fifty percent of sporadic tumours compared to 78% of BRCA2 linked tumours exhibit LOH at one or more markers at 8p showing that chromosome 8p alterations in breast tumours from BRCA2 999del5 carriers are more pronounced than in sporadic breast tumours. The pattern of LOH is different in the two groups and a higher proportion of BRCA2 tumours have LOH in a large region of chromosome 8p. In the total patient material, LOH of 8p is associated with LOH at other chromosome regions, for example, 1p, 3p, 6q, 7q, 9p, 11p, 13q, 17p, and 20q, but no association is found between LOH at 8p and chromosome regions 11q, 16q, 17q, and 18q. Furthermore, an association is detected between LOH at 8p and positive node status, large tumour size, aneuploidy, and high S phase fraction. Breast cancer patients with LOH at chromosome 8p have a worse prognosis than patients without this defect. Multivariate analysis suggests that LOH at 8p is an independent prognostic factor. We conclude that chromosome 8p carries a tumour suppressor gene or genes, the loss of which results in growth advantage of breast tumour cells, especially in carriers of the BRCA2 999del5 mutation.


Keywords: chromosome 8; BRCA2; LOH; breast cancer PMID:10807692

  15. Dual Detection of Chromosomes and Microtubules by the Chromosomal Passenger Complex Drives Spindle Assembly

    PubMed Central

    Tseng, Boo Shan; Tan, Lei; Kapoor, Tarun M.; Funabiki, Hironori

    2010-01-01

    SUMMARY Chromosome-dependent spindle assembly requires the chromosomal recruitment and activation of Aurora B, the kinase subunit of the chromosomal passenger complex (CPC). It remains unclear how the chromosome-activated kinase spatially transmits signals to organize the micron scale spindle. Here we reveal that the CPC must detect two structures, chromosomes and microtubules, to support spindle assembly in Xenopus egg extracts. While Aurora B is enriched on chromosomes in metaphase, we establish that a fraction of Aurora B is targeted to the metaphase spindle and phosphorylates microtubule-bound substrates. We demonstrate that chromosomally activated Aurora B must be targeted to microtubules to drive spindle assembly. Moreover, although the CPC-microtubule interaction can activate Aurora B, which further promotes microtubule assembly, this positive feedback is not initiated without chromosomes. We propose that the dual detection of chromosomes and microtubules by the CPC is a critical step in assembling spindles around and only around chromosomes. PMID:20627073

  16. Dual detection of chromosomes and microtubules by the chromosomal passenger complex drives spindle assembly.

    PubMed

    Tseng, Boo Shan; Tan, Lei; Kapoor, Tarun M; Funabiki, Hironori

    2010-06-15

    Chromosome-dependent spindle assembly requires the chromosomal recruitment and activation of Aurora B, the kinase subunit of the chromosomal passenger complex (CPC). It remains unclear how the chromosome-activated kinase spatially transmits signals to organize the micron-scale spindle. Here we reveal that the CPC must detect two structures, chromosomes and microtubules, to support spindle assembly in Xenopus egg extracts. While Aurora B is enriched on chromosomes in metaphase, we establish that a fraction of Aurora B is targeted to the metaphase spindle and phosphorylates microtubule-bound substrates. We demonstrate that chromosomally activated Aurora B must be targeted to microtubules to drive spindle assembly. Moreover, although the CPC-microtubule interaction can activate Aurora B, which further promotes microtubule assembly, this positive feedback is not initiated without chromosomes. We propose that the dual detection of chromosomes and microtubules by the CPC is a critical step in assembling spindles around and only around chromosomes.

  17. Analysis of chromosomal alterations induced by asbestos and ceramic fibers.

    PubMed

    Dopp, E; Schiffmann, D

    1998-08-01

    increase of chromosomal breakage in the pericentric heterochromatin regions of chromosomes 1 and 9 in AFC after exposure to asbestos and ceramic fibers. The number of hyperdiploid cells was also significantly increased. These results show that asbestos as well as ceramic fibers are inducers of structural and numerical chromosomal alterations.

  18. Chromosome 8p alterations in sporadic and BRCA2 999del5 linked breast cancer.

    PubMed

    Sigbjörnsdottir, B I; Ragnarsson, G; Agnarsson, B A; Huiping, C; Barkardottir, R B; Egilsson, V; Ingvarsson, S

    2000-05-01

    Chromosomal losses involving the short arm of chromosome 8 are frequent in a variety of tumour types, including breast cancer, suggesting the presence of one or more tumour suppressor genes in this region. In this study, we have used 11 microsatellite markers to analyse loss of heterozygosity (LOH) at chromosome 8p in 151 sporadic breast tumours and 50 tumours from subjects carrying the BRCA2 999del5 mutation. Fifty percent of sporadic tumours compared to 78% of BRCA2 linked tumours exhibit LOH at one or more markers at 8p showing that chromosome 8p alterations in breast tumours from BRCA2 999del5 carriers are more pronounced than in sporadic breast tumours. The pattern of LOH is different in the two groups and a higher proportion of BRCA2 tumours have LOH in a large region of chromosome 8p. In the total patient material, LOH of 8p is associated with LOH at other chromosome regions, for example, 1p, 3p, 6q, 7q, 9p, 11p, 13q, 17p, and 20q, but no association is found between LOH at 8p and chromosome regions 11q, 16q, 17q, and 18q. Furthermore, an association is detected between LOH at 8p and positive node status, large tumour size, aneuploidy, and high S phase fraction. Breast cancer patients with LOH at chromosome 8p have a worse prognosis than patients without this defect. Multivariate analysis suggests that LOH at 8p is an independent prognostic factor. We conclude that chromosome 8p carries a tumour suppressor gene or genes, the loss of which results in growth advantage of breast tumour cells, especially in carriers of the BRCA2 999del5 mutation.

  19. [Spectral karyotyping of seven prenatally detected marker chromosomes and complex chromosome aberrations].

    PubMed

    Song, Hua-lei; Chen, Bao-jiang; Fang, Qun; Xie, Ying-jun; Lin, Shao-bin; Wu, Jian-zhu

    2012-08-01

    To perform spectral karyotyping (SKY), fluorescence in situ hybridization (FISH) and conventional karyotyping on prenatally detected marker chromosomes and complex chromosomal aberrations. Five marker chromosomes and 2 complex chromosome aberrations diagnosed by G banding were collected. SKY was performed to verify the composition of marker chromosomes. FISH was used to confirm the diagnosis when necessary. In certain cases, C or N banding technique was employed to verify the composition of chromosomes. Results of ultrasonography and pregnancy outcome were reviewed. Among the 5 marker chromosomes, 2 were large and 3 were medium in size, 4 were de novo and one was inherited from the father. By SKY analysis, 2 marker chromosomes have originated from non-acrocentric chromosomes (4 and 9), whilst the other two have originated from acrocentric chromosomes (21 and 22). The remainder was derived from X chromosome. The SKY results were confirmed by FISH in 3 cases. Four cases have chosen to terminate the pregnancy after genetic counseling. A fetus with inherited paternal marker chromosome was delivered at term, and showed normal development during the first year of life. As for the other 2 cases with complex chromosome aberrations, by SKY examination, one had duplication in chromosome 8 and the other had chromosome rearrangements derived from translocation between chromosomes 2 and 6. In the latter case the fetus was delivered at term but showed developmental retardation at 6 months. SKY in combination with FISH can facilitate identification of the origins of marker chromosomes as well as complex chromosomal aberrations. With combined information from ultrasonography, SKY and FISH, effective counseling may be offered to the patients.

  20. The TP53 tumour suppressor gene in colorectal carcinomas. I. Genetic alterations on chromosome 17.

    PubMed Central

    Meling, G. I.; Lothe, R. A.; Børresen, A. L.; Graue, C.; Hauge, S.; Clausen, O. P.; Rognum, T. O.

    1993-01-01

    In 231 colorectal carcinomas, allele variation at four restriction fragments length polymorphisms (RFLP) loci on chromosome 17 have been studied by Southern analysis. Heterozygous loss of the TP53 gene was found in 68% (129/189) of the carcinomas informative on both chromosome arms. In 41% (77/189) of the carcinomas the loss was found only on 17p. Two probes were used to detect alterations on 17p, pBHP53 and pYNZ22. When loss was demonstrated with pYNZ22, pBHP53 also always showed loss (n = 45), whereas when loss was demonstrated with pBHP53, only 45 of 54 (83%) showed loss with pYNZ22. Loss on 17q was found in 34% (64/189) of the carcinomas, and 6% (12/189) had loss on this chromosome arm, only. Loss on 17q was significantly associated with loss on 17p (P < 0.01). These data confirm that the TP53 gene is the target of loss on chromosome arm 17p in colorectal carcinomas, and demonstrate that loss of the TP53 gene is most frequently part of limited, subchromosomal loss. Furthermore, the results do not suggest any additional tumour suppressor gene(s) on chromosome 17 involved in colorectal carcinogenesis. Images Figure 2 PMID:8094008

  1. Chromosomal and genetic alterations in human hepatocellular adenomas associated with type Ia glycogen storage disease.

    PubMed

    Kishnani, Priya S; Chuang, Tzu-Po; Bali, Deeksha; Koeberl, Dwight; Austin, Stephanie; Weinstein, David A; Murphy, Elaine; Chen, Ying-Ting; Boyette, Keri; Liu, Chu-Hao; Chen, Yuan-Tsong; Li, Ling-Hui

    2009-12-15

    Hepatocellular adenoma (HCA) is a frequent long-term complication of glycogen storage disease type I (GSD I) and malignant transformation to hepatocellular carcinoma (HCC) is known to occur in some cases. However, the molecular pathogenesis of tumor development in GSD I is unclear. This study was conducted to systematically investigate chromosomal and genetic alterations in HCA associated with GSD I. Genome-wide SNP analysis and mutation detection of target genes was performed in ten GSD Ia-associated HCA and seven general population HCA cases for comparison. Chromosomal aberrations were detected in 60% of the GSD Ia HCA and 57% of general population HCA. Intriguingly, simultaneous gain of chromosome 6p and loss of 6q were only seen in GSD Ia HCA (three cases) with one additional GSD I patient showing submicroscopic 6q14.1 deletion. The sizes of GSD Ia adenomas with chromosome 6 aberrations were larger than the sizes of adenomas without the changes (P = 0.012). Expression of IGF2R and LATS1 candidate tumor suppressor genes at 6q was reduced in more than 50% of GSD Ia HCA that were examined (n = 7). None of the GSD Ia HCA had biallelic mutations in the HNF1A gene. These findings give the first insight into the distinct genomic and genetic characteristics of HCA associated with GSD Ia. These results strongly suggest that chromosome 6 alterations could be an early event in the liver tumorigenesis in GSD I, and may be in general population. These results also suggest an interesting relationship between GSD Ia HCA and steps to HCC transformation.

  2. Detection Of Amplified Or Deleted Chromosomal Regions

    SciTech Connect

    Stokke, Trond , Pinkel, Daniel , Gray, Joe W.

    1997-05-27

    The present invention relates to in situ hybridization methods for the identification of new chromosomal abnormalities associated with various diseases. In particular, it provides probes which are specific to a region of amplification in chromosome 20.

  3. Detection of amplified or deleted chromosomal regions

    SciTech Connect

    Stokke, Trond; Pinkel, Daniel; Gray, Joe W.

    1995-01-01

    The present invention relates to in situ hybridization methods for the identification of new chromosomal abnormalities associated with various diseases. In particular, it provides probes which are specific to a region of amplification in chromosome 20.

  4. Detection of amplified or deleted chromosomal regions

    DOEpatents

    Stokke, T.; Pinkel, D.; Gray, J.W.

    1995-12-05

    The present invention relates to in situ hybridization methods for the identification of new chromosomal abnormalities associated with various diseases. In particular, it provides probes which are specific to a region of amplification in chromosome 20. 3 figs.

  5. Defective histone supply causes condensin-dependent chromatin alterations, SAC activation and chromosome decatenation impairment

    PubMed Central

    Murillo-Pineda, Marina; Cabello-Lobato, María J.; Clemente-Ruiz, Marta; Monje-Casas, Fernando; Prado, Félix

    2014-01-01

    The structural organization of chromosomes is essential for their correct function and dynamics during the cell cycle. The assembly of DNA into chromatin provides the substrate for topoisomerases and condensins, which introduce the different levels of superhelical torsion required for DNA metabolism. In particular, Top2 and condensin are directly involved in both the resolution of precatenanes that form during replication and the formation of the intramolecular loop that detects tension at the centromeric chromatin during chromosome biorientation. Here we show that histone depletion activates the spindle assembly checkpoint (SAC) and impairs sister chromatid decatenation, leading to chromosome mis-segregation and lethality in the absence of the SAC. We demonstrate that histone depletion impairs chromosome biorientation and activates the Aurora-dependent pathway, which detects tension problems at the kinetochore. Interestingly, SAC activation is suppressed by the absence of Top2 and Smc2, an essential component of condensin. Indeed, smc2-8 suppresses catenanes accumulation, mitotic arrest and growth defects induced by histone depletion at semi-permissive temperature. Remarkably, SAC activation by histone depletion is associated with condensin-mediated alterations of the centromeric chromatin. Therefore, our results reveal the importance of a precise interplay between histone supply and condensin/Top2 for pericentric chromatin structure, precatenanes resolution and centromere biorientation. PMID:25300489

  6. Maternal X chromosome copy number variations are associated with discordant fetal sex chromosome aneuploidies detected by noninvasive prenatal testing.

    PubMed

    Wang, Shaowei; Huang, Shuai; Ma, Linlin; Liang, Lin; Zhang, Junrong; Zhang, Jianguang; Cram, David S

    2015-04-15

    The sensitivity and specificity of noninvasive prenatal testing (NIPT) for detection of sex chromosome aneuploidies (SCAs) compared to common autosomal trisomies are significantly lower. We speculated that in addition to altered maternal X chromosome karyotype, maternal X chromosome copy number variations (CNVs) may also contribute to discordant NIPT SCA results. Clinical NIPT was performed for pregnant women at a single hospital. Copy number variation sequencing (CNV-Seq) was used to identify and quantitate the copy number of maternal X chromosome CNVs for each positive SCA pregnancy. Two out of 25 SCA positive NIPT samples had slightly abnormal ChrX/ChrY z-scores and were referred for invasive test confirmation. However, fetal karyotypes were found to be normal. CNV-Seq analysis of the maternal white blood cell DNA archived from the original two NIPT blood samples identified small CNVs spanning the STS gene, which is associated with X-linked ichthyosis. Correcting for the altered plasma levels of X chromosome DNA caused by the two CNVs and, taking into consideration the phenotypic consequences for X-linked disease, both fetuses were diagnosed as normal. Maternal DNA sequencing is recommended for all positive NIPT SCA results to avoid unnecessary referral for invasive testing and also to evaluate the risk to the fetus of X-linked disease. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Chromosomal alterations in pure nonneoplastic breast lesions: implications for breast cancer progression.

    PubMed

    Ellsworth, Rachel E; Ellsworth, Darrell L; Weyandt, Jamie D; Fantacone-Campbell, Jamie L; Deyarmin, Brenda; Hooke, Jeffrey A; Shriver, Craig D

    2010-06-01

    Columnar cell lesions (CCL) and atypical ductal hyperplasia (ADH) frequently coexist and share molecular changes with in situ and invasive components, suggesting that CCL and ADH may be precursors to breast cancer. These conclusions are largely based on studies examining CCL and/or ADH from patients diagnosed with more advanced disease. We assessed allelic imbalance (AI) in pure CCL or ADH specimens to characterize molecular changes in nonneoplastic breast lesions. DNA samples were obtained from laser-microdissected pure CCL (n = 42) or ADH (n = 31). AI was assessed at 26 chromosomal regions commonly altered in breast cancer. Data were analyzed using Fisher's exact and Student's t-tests using a cutoff of P < 0.05. The average AI frequency was 6.2% in CCL and 6.1% in ADH; approximately 33% of nonneoplastic lesions had no detectable genetic changes. Levels of AI in CCL and ADH were significantly (P < 0.0001) lower than observed in either low- or high-grade ductal carcinoma in situ (DCIS) lesions. Genetic changes characteristic of in situ and invasive disease, especially on chromosomes 16q and 17p, were infrequent in pure nonneoplastic lesions. Pure CCL and ADH lesions demonstrate lower levels of genetic alterations than DCIS, invasive carcinomas or CCL/ADH lesions from cancerous breasts; alterations of chromosomes 16q and 17p were not detected. Pure CCL and ADH lesions are not genetically advanced, and molecular profiles do not support these lesions as obligatory precursors to more advanced disease. Molecular differences between pure and synchronous lesions support re-evaluation of current models of disease initiation, progression, and risk.

  8. Application of flow karyotyping in prenatal detection of chromosome aberrations.

    PubMed Central

    Gray, J W; Trask, B; van den Engh, G; Silva, A; Lozes, C; Grell, S; Schonberg, S; Yu, L C; Golbus, M S

    1988-01-01

    This paper describes the application of bivariate flow karyotyping to (1) classification of chromosomes isolated from cultures of cells taken by amniocentesis and (2) detection of numerical and structural aberrations. Chromosomes were isolated from primary cultures 2-5 wk after amniocentesis, stained with Hoechst 33258 and chromomycin A3, and analyzed using dual beam flow cytometry. Information about chromosome DNA content and DNA base composition was derived from the locations of the peaks in the flow karyotypes, each peak being produced by one or more chromosome types with similar DNA content and DNA base composition. Information about the relative frequency of each chromosome type was determined on the basis of the relative volume of the peak for that chromosome type. Cytogenetic information determined on the basis of flow karyotypes was compared with that obtained by visual analysis following G-banding. Variability among the peak means and volumes in flow karyotypes was determined from analyses of 50 normal amniocyte cultures. Numerical aberrations involving chromosomes 21, 18, and Y were detected correctly in all of 28 analyses, including eight in a blind study. Structural aberrations involving chromosomes 1, 2, 3, 6, 9-12, 13, 14, 15, 21, and 22 were detected in all of seven cultures in a blind study. Flow karyotypes proved to be insensitive to small, normally occurring chromosome polymorphisms detected by banding analysis. In addition, a few samples were erroneously scored as having numerical aberrations. PMID:3337112

  9. Chromosomal alterations associated with the transition from in situ to invasive breast cancer.

    PubMed

    Ellsworth, Rachel E; Vertrees, Amy; Love, Brad; Hooke, Jeffrey A; Ellsworth, Darrell L; Shriver, Craig D

    2008-09-01

    Ductal carcinoma in situ (DCIS) is a preinvasive lesion of the breast with an inherent but nonobligatory tendency for progression to invasive breast cancer. Although the transition from in situ to invasive disease is critical to the development of breast cancer, molecular and biological changes responsible for this transition are not well characterized. Chromosomal alterations at 26 regions were assayed in 66 DCIS lesions and 111 invasive ductal carcinomas. Levels and patterns of allelic imbalance (AI) were compared between grade 1 DCIS and well-differentiated breast carcinomas, and between grade 3 DCIS and poorly differentiated invasive breast carcinomas, using Fisher's exact and Student's t-tests. Levels of AI were significantly lower (P < 0.01) in grade 1 DCIS (11.9%) compared to well-differentiated carcinomas (19.2%), but were not significantly different between grade 3 DCIS and poorly differentiated tumors. No significant differences were detected at any of the 26 chromosomal regions between low-grade DCIS and invasive tumors; however, AI events at chromosomes 1p36, 11q23, and 16q11-q22 could discriminate high-grade in situ from invasive disease. Lower levels of AI in low-grade in situ compared with invasive disease may reflect the protracted time to progression associated with low-grade DCIS. Increased levels of AI at chromosomes 1p36 and 11q23 in poorly differentiated carcinomas may harbor genes associated with invasiveness, while loss of chromosome 16q11-q22 may prevent the transition from in situ to invasive disease. Further characterization of these changes may provide molecular assays to identify DCIS lesions with invasive potential as well as targets for molecular therapeutics.

  10. Assessment of esophageal adenocarcinoma risk using somatic chromosome alterations in longitudinal samples in Barrett’s esophagus

    PubMed Central

    Li, Xiaohong; Paulson, Thomas G.; Galipeau, Patricia C.; Sanchez, Carissa A.; Liu, Karen; Kuhner, Mary K.; Maley, Carlo C.; Self, Steven G.; Vaughan, Thomas L.; Reid, Brian J.; Blount, Patricia L.

    2015-01-01

    Cancers detected at a late stage are often refractory to treatments and ultimately lethal. Early detection can significantly increase survival probability, but attempts to reduce mortality by early detection have frequently increased overdiagnosis of indolent conditions that do not progress over a lifetime. Study designs that incorporate biomarker trajectories in time and space are needed to distinguish patients who progress to an early cancer from those who follow an indolent course. Esophageal adenocarcinoma (EA) is characterized by evolution of punctuated and catastrophic somatic chromosomal alterations and high levels of overall mutations but few recurrently mutated genes aside from TP53. Endoscopic surveillance of Barrett’s esophagus (BE) for early cancer detection provides an opportunity for assessment of alterations for cancer risk in patients who progress to EA compared to nonprogressors. We investigated 1,272 longitudinally collected esophageal biopsies in a 248 Barrett’s patient case-cohort study with 20,425 person-months of follow-up, including 79 who progressed to early-stage EA. Cancer progression risk was assessed for total chromosomal alterations, diversity, and chromosomal region-specific alterations measured with single nucleotide polymorphism arrays in biopsies obtained over esophageal space and time. A model using 29 chromosomal features was developed for cancer risk prediction (Area under receiver operator curve=0.94). The model prediction performance was robust in two independent EA sets and outperformed TP53 mutation, flow cytometric DNA content and histopathologic diagnosis of dysplasia. This study offers a strategy to reduce overdiagnosis in BE and improve early detection of EA and potentially other cancers characterized by punctuated and catastrophic chromosomal evolution. PMID:26130253

  11. Alteration of chromosome behavior and synchronization of parental chromosomes after successive generations in Brassica napus x Orychophragmus violaceus hybrids.

    PubMed

    Zhao, Zhigang; Ma, Ni; Li, Zaiyun

    2007-02-01

    In an earlier study, the progenies of intergeneric hybrids Brassica napus (2n = 38) x Orychophragmus violaceus (2n = 24) were investigated in successive generations (F1-F4) for the cytological phenomenon of parental genome separation during mitotic and meiotic division. In the present study, inbred lines (F5-F8) derived from 1 such hybrid were characterized for morphology, chromosome pairing behaviour, and genome composition. One F5 plant (2n = 31) with slightly yellow petals and 12:19 and 15:16 segregation ratios in its pollen mother cells (PMCs) produced F6 plants with distinct morphological characteristics and wide variations in fertility and chromosome numbers (2n = 25-38). F7 and F8 lines with distinctive morphology and wide ranges in chromsome numbers were established. In PMCs of F7 plants from 4 F6 plants, 0-12 labelled chromosomes from O. violaceus, which predominantly appeared as bivalents, were identified by genomic in situ hybridization. They behaved synchronously with B. napus chromosomes during meiotic division. The results provide molecular cytogenetic evidence of the inclusion of O. violaceus chromosomes in the original hybrids and the cytology in the hybrids documented earlier. They also show that chromosome behaviour was altered and the parental chromosomes became synchronized after successive generations.

  12. Greig cephalopolysyndactyly syndrome: Altered phenotype of a contiguous gene syndrome by the presence of a chromosomal deletion

    SciTech Connect

    Hersh, J.H.; Williams, P.G.; Yen, F.F.

    1994-09-01

    Greig cephalopolysyndactyly syndrome (GCPS) is characterized by craniofacial anomalies, broad thumbs and halluces, polydactyly of the hands and feet, and variable syndactyly. Intellectual abilities are usually normal. Inheritance is in an autosomal dominant fashion. The disorder has been mapped to chromosome 7p13, suggesting that the condition represents a contiguous gene syndrome (CGS). A male infant presented with multiple congenital anomalies, including omphalocele, dysgenesis of the corpus callosum, hydrocephalus, esotropia, broad thumbs and halluces, syndactyly, polydactyly of one foot, hypotonia and developmental delay. A de novo interstitial deletion of chromosome 7p was detected, 46,XY,del(7)(p13p15). Although clinical findings in this case were reminiscent of GCPS, and the chromosomal abnormality included the region assigned to the candidate gene for this syndrome, additional physical abnormalities were present, as well as cognitive deficits. Some of these features have been previously described in patients with chromosomal deletions of 7p. The chromosomal abnormality in our case provides supportive evidence of the gene locus in GCPS, and that GCPS represents a new CGS. However, a larger deletion, extending beyond the limits of the gene, significantly altered the phenotype. Isolation of the gene responsible for GCPS, and identification of additional patients with chromosomal abnormalities in this region of chromosome 7, should help to provide more accurate genotype-phenotype correlations.

  13. Detection of sex chromosomal aneuploidies X-X, Y-Y, and X-Y in human sperm using two-chromosome fluorescence in situ hybridization

    SciTech Connect

    Wyrobek, A.J.; Robbins, W.A. |; Pinkel, D.; Weier, H.U.; Mehraein, Y. |

    1994-10-15

    Sex chromosome aneuploidy is the most common numerical chromosomal abnormality in humans at birth and a substantial portion of these abnormalities involve paternal chromosomes. An efficient method is presented for using air-dried smears of human semen to detect the number of X and Y chromosomes in sperm chromatin using two-chromosome fluorescence in situ hybridization. Air-dried semen smears were pre-treated with dithiothreitol and 3,4-diiodosalicylate salt to decondense the sperm chromatin and then were hybridized with repetitive sequence DNA probes that had been generated by PCR and differentially labeled. Hybridizations with X and Y specific probes showed the expected ratio of 50%X:50%Y bearing sperm. Sperm carrying extra fluorescence domains representing disomy for the X or Y chromosomes occurred at frequencies of {approximately} 4 per 10,000 sperm each. Cells carrying both X and Y fluorescence domains occurred at a frequency of {approximately} 6/10,000. Thus, the overall frequency of sperm that carried an extra sex chromosome was 1.4/1,000. The frequencies of sperm carrying sex chromosome aneuploidies determined by hybridization did not differ statistically from those reported from the same laboratory using the human-sperm/hamster-egg cytogenetic technique. Multi-chromosome fluorescence in situ hybridization to sperm is a promising method for assessing sex-ratio alterations in human semen and for determining the fraction of sperm carrying sex or other chromosome aneuploidies which may be transmissible to offspring. 44 refs., 1 fig., 3 tabs.

  14. Detection of complex genetic alterations in human glioblastoma multiforme using comparative genomic hybridization

    SciTech Connect

    Schlegel, J.; Stumm, G.; Scherthan, H.; Arens, N.

    1996-01-01

    The aim of the present study was to detect complex genetic alterations in human glioblastoma multiforme (GBM) by comparative genomic in situ hybridization (CGH). Of the 24 GBM that were examined, increased fluorescence intensities indicating chromosomal polysomy of chromosome 7 and gene amplification at chromosome 7p were found in 42% of the tumors. In addition, signal enhancement of chromosome 19 was present in 29% and at 12q13-15 in 21% of the tumors. We also detected reduction of fluorescence intensities indicating gross deletions on chromosomes 10 (58%), 9p (46%), and 13 (29%). There was a close correlation of CGH results when compared with Southern analysis of the EGFR gene localized on chromosome 7 and loss of heterozygosity detection of chromosome 9 and 10 by microsatellite PCR. A close correlation was also observed between copy number changes of chromosome 7 and deletions of chromosome 10. Amplification of chromosome 12q and deletions of chromosomes 9p and 13 seemed to be complementary in the tumors investigated in the present study. 44 refs., 3 figs., 1 tab.

  15. GeneBreak: detection of recurrent DNA copy number aberration-associated chromosomal breakpoints within genes.

    PubMed

    van den Broek, Evert; van Lieshout, Stef; Rausch, Christian; Ylstra, Bauke; van de Wiel, Mark A; Meijer, Gerrit A; Fijneman, Remond J A; Abeln, Sanne

    2016-01-01

    Development of cancer is driven by somatic alterations, including numerical and structural chromosomal aberrations. Currently, several computational methods are available and are widely applied to detect numerical copy number aberrations (CNAs) of chromosomal segments in tumor genomes. However, there is lack of computational methods that systematically detect structural chromosomal aberrations by virtue of the genomic location of CNA-associated chromosomal breaks and identify genes that appear non-randomly affected by chromosomal breakpoints across (large) series of tumor samples. 'GeneBreak' is developed to systematically identify genes recurrently affected by the genomic location of chromosomal CNA-associated breaks by a genome-wide approach, which can be applied to DNA copy number data obtained by array-Comparative Genomic Hybridization (CGH) or by (low-pass) whole genome sequencing (WGS). First, 'GeneBreak' collects the genomic locations of chromosomal CNA-associated breaks that were previously pinpointed by the segmentation algorithm that was applied to obtain CNA profiles. Next, a tailored annotation approach for breakpoint-to-gene mapping is implemented. Finally, dedicated cohort-based statistics is incorporated with correction for covariates that influence the probability to be a breakpoint gene. In addition, multiple testing correction is integrated to reveal recurrent breakpoint events. This easy-to-use algorithm, 'GeneBreak', is implemented in R ( www.cran.r-project.org) and is available from Bioconductor ( www.bioconductor.org/packages/release/bioc/html/GeneBreak.html).

  16. Disruption of a conserved CAP-D3 threonine alters condensin loading on mitotic chromosomes leading to chromosome hypercondensation.

    PubMed

    Bakhrebah, Muhammed; Zhang, Tao; Mann, Jeff R; Kalitsis, Paul; Hudson, Damien F

    2015-03-06

    The condensin complex plays a key role in organizing mitotic chromosomes. In vertebrates, there are two condensin complexes that have independent and cooperative roles in folding mitotic chromosomes. In this study, we dissect the role of a putative Cdk1 site on the condensin II subunit CAP-D3 in chicken DT40 cells. This conserved site has been shown to activate condensin II during prophase in human cells, and facilitate further phosphorylation by polo-like kinase I. We examined the functional significance of this phosphorylation mark by mutating the orthologous site of CAP-D3 (CAP-D3(T1403A)) in chicken DT40 cells. We show that this mutation is a gain of function mutant in chicken cells; it disrupts prophase, results in a dramatic shortening of the mitotic chromosome axis, and leads to abnormal INCENP localization. Our results imply phosphorylation of CAP-D3 acts to limit condensin II binding onto mitotic chromosomes. We present the first in vivo example that alters the ratio of condensin I:II on mitotic chromosomes. Our results demonstrate this ratio is a critical determinant in shaping mitotic chromosomes. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Disruption of a Conserved CAP-D3 Threonine Alters Condensin Loading on Mitotic Chromosomes Leading to Chromosome Hypercondensation*

    PubMed Central

    Bakhrebah, Muhammed; Zhang, Tao; Mann, Jeff R.; Kalitsis, Paul; Hudson, Damien F.

    2015-01-01

    The condensin complex plays a key role in organizing mitotic chromosomes. In vertebrates, there are two condensin complexes that have independent and cooperative roles in folding mitotic chromosomes. In this study, we dissect the role of a putative Cdk1 site on the condensin II subunit CAP-D3 in chicken DT40 cells. This conserved site has been shown to activate condensin II during prophase in human cells, and facilitate further phosphorylation by polo-like kinase I. We examined the functional significance of this phosphorylation mark by mutating the orthologous site of CAP-D3 (CAP-D3T1403A) in chicken DT40 cells. We show that this mutation is a gain of function mutant in chicken cells; it disrupts prophase, results in a dramatic shortening of the mitotic chromosome axis, and leads to abnormal INCENP localization. Our results imply phosphorylation of CAP-D3 acts to limit condensin II binding onto mitotic chromosomes. We present the first in vivo example that alters the ratio of condensin I:II on mitotic chromosomes. Our results demonstrate this ratio is a critical determinant in shaping mitotic chromosomes. PMID:25605712

  18. Mutations in g protein encoding genes and chromosomal alterations in primary leptomeningeal melanocytic neoplasms.

    PubMed

    Küsters-Vandevelde, Heidi V N; van Engen-van Grunsven, Ilse A C H; Coupland, Sarah E; Lake, Sarah L; Rijntjes, Jos; Pfundt, Rolph; Küsters, Benno; Wesseling, Pieter; Blokx, Willeke A M; Groenen, Patricia J T A

    2015-04-01

    Limited data is available on the genetic features of primary leptomeningeal melanocytic neoplasms (LMNs). Similarities with uveal melanoma were recently suggested as both entities harbor oncogenic mutations in GNAQ and GNA11. Whether primary LMNs share additional genetic alterations with uveal melanoma including copy number variations is unknown. Twenty primary LMNs ranging from benign and intermediate-grade melanocytomas to melanomas were tested by direct sequencing for hotspot mutations in the genes GNA11, GNAQ, BRAF, NRAS and HRAS. Furthermore, the lesions were tested for copy number variations of chromosomes frequently present in uveal melanoma (1p, 3, 6 and 8q) by multiplex ligation-dependent probe amplification (MLPA). Genome-wide analyses of copy number alterations of two leptomeningeal melanocytic neoplasms were performed using the OncoScan SNP-array. GNAQ(Q209) mutations were present in eleven LMNs, while two of 20 cases carried a GNA11(Q209) mutation. No BRAF, HRAS or NRAS hotspot mutations were detected. Monosomy 3 and gain of 8q were present in one leptomeningeal melanoma, and one intermediate-grade melanocytoma harbored a gain of chromosome 6. With MLPA, the melanocytomas did not show any further gross chromosomal variations. Our data shows that primary LMNs, like uveal melanoma, harbor oncogenic mutations in GNAQ and GNA11 but lack mutations in BRAF, NRAS and HRAS. This finding may help in the differential diagnosis between a primary LMN and a metastasis from a cutaneous melanoma to the central nervous system. Copy number variations in some aggressive LMNs resemble those present in uveal melanoma but their prognostic significance is unclear.

  19. Detection of chromosomal abnormalities by comparative genomic hybridization.

    PubMed

    Lapierre, Jean-Michel; Tachdjian, Gérard

    2005-04-01

    Comparative genomic hybridization (CGH) is a modified in-situ hybridization technique. In this type of analysis, two differentially labeled genomic DNAs (study and reference) are cohybridized to normal metaphase spreads or to microarray. Chromosomal locations of copy number changes in the DNA segments of the study genome are revealed by a variable fluorescence intensity ratio along each target chromosome. Thus, CGH allows detection and mapping of DNA sequence copy differences between two genomes in a single experiment. Since its development, comparative genomic hybridization has been applied mostly as a research tool in the field of cancer cytogenetics to identify genetic changes in many previously unknown regions. It is also a powerful tool for detection and identification of unbalanced chromosomal abnormalities in prenatal, postnatal and preimplantation diagnostics. The development of comparative genomic hybridization and increase in resolution analysis by using the microarray-based technique offer new information on chromosomal pathologies and thus better management of patients.

  20. Chromosome damage and aneuploidy detected by interphase multicolour FISH in benzene-exposed shale oil workers.

    PubMed

    Marcon, F; Zijno, A; Crebelli, R; Carere, A; Veidebaum, T; Peltonen, K; Parks, R; Schuler, M; Eastmond, D

    1999-09-30

    A multicolour tandem-labelling fluorescence in situ hybridization (FISH) procedure was used to detect chromosome alterations in peripheral blood cells of a group of Estonian petrochemistry workers. Twelve workers employed in benzene production and five cokery workers, together with eight unexposed rural controls, were enrolled in the study. The methodology employed, based on the in situ hybridization of adjacent centromeric and pericentromeric regions, allowed the simultaneous detection of both chromosome breakage, involving damage-prone pericentromeric regions, and hyperploidy in interphase cells. Blood smears from all subjects were hybridized with chromosome 1 specific probes, in order to detect genotoxic damage in circulating lymphocytes and granulocytes. Moreover, lymphocyte cultures were established, harvested 48 h following mitogen stimulation and hybridized with the tandem chromosomes 1 and 9 probes. No significant difference in the incidence of breakage was detected in the nucleated cells of blood smears of exposed vs. control subjects. In contrast, modest but significantly increased frequencies of breakage affecting both chromosomes 1 and 9 were observed in the cultured lymphocytes of the benzene-exposed workers compared to the unexposed controls, suggesting an expression of premutagenic lesions during the S-phase in vitro. Across the entire study group, the frequencies of breakage affecting chromosomes 1 and 9 in the stimulated lymphocytes were highly intercorrelated (p < 0.001). No significant difference was found in the incidence of hyperploidy among the study groups, although a tendency to higher values was observed in benzene-exposed workers. Although the relatively small size of the study groups does not allow firm conclusions on the role of occupational exposure, the observed patterns are suggestive of effects in the benzene-exposed workers. This work also shows that tandem labelling FISH can be usefully applied in human biomonitoring, allowing the

  1. High-order chromatin architecture shapes the landscape of chromosomal alterations in cancer

    NASA Astrophysics Data System (ADS)

    Fudenberg, Geoffrey; Getz, Gad; Meyerson, Matthew; Mirny, Leonid

    2012-02-01

    The rapid growth of cancer genome structural information provides an opportunity for a better understanding of the mutational mechanisms of genomic alterations in cancer and the forces of selection that act upon them. Here we test the evidence for two major forces, spatial chromosome structure and purifying (or negative) selection, that shape the landscape of somatic copy-number alterations (SCNAs) in cancer (Beroukhim et al, 2010). Using a maximum likelihood framework we compare SCNA maps and three-dimensional genome architecture as determined by genome-wide chromosome conformation capture (HiC) and described by the proposed fractal-globule (FG) model (Lieberman-Aiden and Van Berkum et al, 2009). This analysis provides evidence that the distribution of chromosomal alterations in cancer is spatially related to three-dimensional genomic architecture and additionally suggests that purifying selection as well as positive selection shapes the landscape of SCNAs during somatic evolution of cancer cells.

  2. Evidence that both genetic instability and selection contribute to the accumulation of chromosome alterations in cancer.

    PubMed

    Gorringe, Kylie L; Chin, Suet-Feung; Pharoah, Paul; Staines, Joanne M; Oliveira, Carla; Edwards, Paul A W; Caldas, Carlos

    2005-05-01

    Cancer cells contain many genetic alterations, and genetic instability may be important in tumourigenesis. We evaluated 58 breast and ovarian cancer cell lines for microsatellite instability (MSI) and chromosomal instability (CIN). MSI was identified in 3/33 breast and 5/25 ovarian cell lines, and 7/8 MSI lines showed an inactivation of mismatch repair. Average ploidy by centromeric fluorescence in situ hybridization (FISH) of MSI (n = 8, average ploidy = 2.65) and microsatellite stable (MSS; n = 7, average ploidy = 3.01) cell lines was not different, due to the presence of three aneuploid MSI lines, and two near-diploid MSS lines. However, the variability of the centromeric FISH data was different between MSI and MSS (P = 0.049). The complexity of structural chromosomal rearrangements was not different between MSI and MSS. Thus, MSI and numerical CIN are not mutually exclusive, and structural CIN occurs independently of MSI or numerical CIN. Dynamic genetic instability was evaluated in three cell lines-MSI diploid (MT-3), MSS diploid (SUM159) and MSS aneuploid (MT-1). Ten clones of each of these cell lines were analysed by centromeric FISH and six-colour chromosome painting. The variation in chromosome number was different among all three cell lines (P < 0.001). MT-3 appeared numerically constant (94% of centromeric FISH signals matched the mode). SUM159 was 88% constant; however, 7% of cells had duplicated chromosomes. MT-1 was 82% constant; most changes were chromosomal losses. The six-colour FISH data showed that SUM159 had more stable structural chromosomal alterations (e.g. chromosomal translocations) compared with MT-3 and MT-1, but had no increase in unstable changes (e.g. chromatid breaks) when compared with MT-3. MT-1 had fewer unstable changes than both MT-3 and SUM159. These data suggest that numerical CIN may contribute to aneuploidy, but that selection plays an important role, particularly for the accumulation of structural chromosomal changes.

  3. Investigating 22q11.2 deletion and other chromosomal aberrations in fetuses with heart defects detected by prenatal echocardiography.

    PubMed

    Bellucco, Fernanda Teixeira da Silva; Belangero, Sintia Iole Nogueira; Farah, Leila Montenegro Silveira; Machado, Maria Virgínia Lima; Cruz, Adriano Pastor; Lopes, Lílian Maria; Lopes, Marco Antonio Borges; Zugaib, Marcelo; Cernach, Mirlene Cecília; Melaragno, Maria Isabel

    2010-11-01

    Congenital heart disease (CHD) is the most common birth defect and the leading cause of mortality in the first year of life. In fetuses with a heart defect, chromosomal abnormalities are very frequent. Besides aneuploidy, 22q11.2 deletion is one of the most recognizable chromosomal abnormalities causing CHD. The frequency of this abnormality varies in nonselected populations. This study aimed to investigate the incidence of the 22q11.2 deletion and other chromosomal alterations in a Brazilian sample of fetuses with structural cardiac anomalies detected by fetal echocardiography. In a prospective study, 68 fetuses with a heart defect were evaluated. Prenatal detection of cardiac abnormalities led to identification of aneuploidy or structural chromosomal anomaly in 35.3% of these cases. None of the fetuses with apparently normal karyotypes had a 22q11.2 deletion. The heart defects most frequently associated with chromosomal abnormalities were atrioventricular septal defect (AVSD), ventricular septal defect (VSD), and tetralogy of Fallot. Autosomal trisomies 18 and 21 were the most common chromosomal abnormalities. The study results support the strong association of chromosome alterations and cardiac malformation, especially in AVSD and VSD, for which a chromosome investigation is indicated. In fetuses with an isolated conotruncal cardiopathy, fluorescence in situ hybridization (FISH) to investigate a 22q11.2 deletion is not indicated.

  4. Modeling the dynamics of chromosomal alteration progression in cervical cancer: A computational model

    PubMed Central

    2017-01-01

    Computational modeling has been applied to simulate the heterogeneity of cancer behavior. The development of Cervical Cancer (CC) is a process in which the cell acquires dynamic behavior from non-deleterious and deleterious mutations, exhibiting chromosomal alterations as a manifestation of this dynamic. To further determine the progression of chromosomal alterations in precursor lesions and CC, we introduce a computational model to study the dynamics of deleterious and non-deleterious mutations as an outcome of tumor progression. The analysis of chromosomal alterations mediated by our model reveals that multiple deleterious mutations are more frequent in precursor lesions than in CC. Cells with lethal deleterious mutations would be eliminated, which would mitigate cancer progression; on the other hand, cells with non-deleterious mutations would become dominant, which could predispose them to cancer progression. The study of somatic alterations through computer simulations of cancer progression provides a feasible pathway for insights into the transformation of cell mechanisms in humans. During cancer progression, tumors may acquire new phenotype traits, such as the ability to invade and metastasize or to become clinically important when they develop drug resistance. Non-deleterious chromosomal alterations contribute to this progression. PMID:28723940

  5. Modeling the dynamics of chromosomal alteration progression in cervical cancer: A computational model.

    PubMed

    Cabrera-Becerril, Augusto; Vargas-De-León, Cruz; Hernández, Sergio; Miramontes, Pedro; Peralta, Raúl

    2017-01-01

    Computational modeling has been applied to simulate the heterogeneity of cancer behavior. The development of Cervical Cancer (CC) is a process in which the cell acquires dynamic behavior from non-deleterious and deleterious mutations, exhibiting chromosomal alterations as a manifestation of this dynamic. To further determine the progression of chromosomal alterations in precursor lesions and CC, we introduce a computational model to study the dynamics of deleterious and non-deleterious mutations as an outcome of tumor progression. The analysis of chromosomal alterations mediated by our model reveals that multiple deleterious mutations are more frequent in precursor lesions than in CC. Cells with lethal deleterious mutations would be eliminated, which would mitigate cancer progression; on the other hand, cells with non-deleterious mutations would become dominant, which could predispose them to cancer progression. The study of somatic alterations through computer simulations of cancer progression provides a feasible pathway for insights into the transformation of cell mechanisms in humans. During cancer progression, tumors may acquire new phenotype traits, such as the ability to invade and metastasize or to become clinically important when they develop drug resistance. Non-deleterious chromosomal alterations contribute to this progression.

  6. Novel, gross chromosomal alterations involving PTEN cooperate with allelic loss in prostate cancer.

    PubMed

    Reid, Alison H M; Attard, Gerhardt; Brewer, Daniel; Miranda, Susana; Riisnaes, Ruth; Clark, Jeremy; Hylands, Lucy; Merson, Sue; Vergis, Roy; Jameson, Charles; Høyer, Søren; Sørenson, Karina Dalsgaard; Borre, Michael; Jones, Chris; de Bono, Johann S; Cooper, Colin S

    2012-06-01

    There is increasing evidence that multiple chromosomal rearrangements occur in prostate cancer. PTEN loss is considered to be a key event in prostate carcinogenesis but the mechanisms of loss remain to be fully elucidated. We hypothesised that gross rearrangements may exist that cause disruption of the PTEN gene in the absence of genomic deletion. We therefore designed a novel fluorescence in situ hybridisation (FISH) assay with probes overlying regions 3' and 5' of PTEN and a third probe overlying the gene. We aimed to identify both genomic deletions and gross rearrangements of PTEN that would be overlooked by previously reported single-probe FISH assays. We proceeded to evaluate a tissue microarray with radical prostatectomy and trans-urethral resection of the prostate specimens from 187 patients. We identified PTEN genomic loss in 45/150 (30%) radical prostatectomy patients and 16/37 (43%) trans-urethral resection of the prostate patients. Importantly, our assay detected novel chromosomal alterations in the PTEN gene (characterised by splitting of FISH signals) in 13 tumours (6.9% of all prostate cancers; 21% of PTEN-lost cancers). All PTEN-rearranged tumours had genomic loss at the other allele and had no expression of PTEN by immunohistochemistry. PTEN-rearranged tumours were significantly more likely to have an underlying ERG rearrangement. Our assay differentiated loss of the probe overlying PTEN in isolation or in combination with either one of or both the probes overlying the 3' and 5' regions. This gave an indication of the size of genomic loss and we observed considerable inter-tumoural heterogeneity in the extent of genomic loss in PTEN-lost tumours. In summary, gross rearrangements of the PTEN locus occur in prostate cancer and can be detected by a 'break-apart' FISH assay. This observation could explain the absence of PTEN protein expression in a subgroup of tumours previously classified as having heterozygous genomic loss using single

  7. Chromosomal and Nuclear Alterations in Root Tip Cells of Allium Cepa L. Induced by Alprazolam

    PubMed Central

    Nefic, Hilada; Musanovic, Jasmin; Metovic, Azra; Kurteshi, Kemajl

    2013-01-01

    ABSTRACT Introduction: Alprazolam is a triazolobenzodiazepine used in panic disorders and other anxiety states. Target organ of Alprazolam is CNS, causing depression of respiration and consciousness. Aim: This study aimed to estimate the genotoxic potential of Alprazolam using Allium cepa test. Methods: Allium cepa is one of the most suitable plants for detecting different types of xenobiotics. The test enables the assessment of different genetic endpoints making possible damage to the DNA of humans to be predicted. Results: Alprazolam induced chromosomal (anaphase bridges, breaks, lagging and stickiness, abnormal spiralisation, multipolarity and polyploidy) and cytological aberrations, especially nuclear alterations (nuclear buds, fragmented nucleus and apoptotic bodies, cells without nucleus, binucleated and micronucleated cells), morphological alterations in shape and size of cells, spindle disturbance and polar deviation in root tip meristem cells of Allium cepa at all tested concentrations. Alprazolam also caused significant inhibition of mitotic index in these cells. Conclusion: These changes in cells are indicators of genotoxic potential of Alprazolam suggesting a need for further in vitro studies on animal and human lymphocytes as well as in vivo studies. PMID:25568504

  8. Precise detection of rearrangement breakpoints in mammalian chromosomes

    PubMed Central

    Lemaitre, Claire; Tannier, Eric; Gautier, Christian; Sagot, Marie-France

    2008-01-01

    Background Genomes undergo large structural changes that alter their organisation. The chromosomal regions affected by these rearrangements are called breakpoints, while those which have not been rearranged are called synteny blocks. We developed a method to precisely delimit rearrangement breakpoints on a genome by comparison with the genome of a related species. Contrary to current methods which search for synteny blocks and simply return what remains in the genome as breakpoints, we propose to go further and to investigate the breakpoints themselves in order to refine them. Results Given some reliable and non overlapping synteny blocks, the core of the method consists in refining the regions that are not contained in them. By aligning each breakpoint sequence against its specific orthologous sequences in the other species, we can look for weak similarities inside the breakpoint, thus extending the synteny blocks and narrowing the breakpoints. The identification of the narrowed breakpoints relies on a segmentation algorithm and is statistically assessed. Since this method requires as input synteny blocks with some properties which, though they appear natural, are not verified by current methods for detecting such blocks, we further give a formal definition and provide an algorithm to compute them. The whole method is applied to delimit breakpoints on the human genome when compared to the mouse and dog genomes. Among the 355 human-mouse and 240 human-dog breakpoints, 168 and 146 respectively span less than 50 Kb. We compared the resulting breakpoints with some publicly available ones and show that we achieve a better resolution. Furthermore, we suggest that breakpoints are rarely reduced to a point, and instead consist in often large regions that can be distinguished from the sequences around in terms of segmental duplications, similarity with related species, and transposable elements. Conclusion Our method leads to smaller breakpoints than already published ones

  9. Transgene integration and chromosome alterations in two transgenic lines of tritordeum.

    PubMed

    Barro, F; Martín, A; Cabrera, A

    2003-01-01

    Plants from two transgenic lines of tritordeum (an amphiploid between Triticum turgidum cv. durum and Hordeumn chilense) have been analyzed by fluorescence in-situ hybridization (FISH) to characterize the transgene integration sites and chromosome rearrangements. Transgenic lines were transformed in two different events with the genes encoding for the high-molecular-weight glutenin subunits (HMW-GS), 1Ax1 and/or 1Dx5. Three integration sites and four translocations were detected. All three integration sites were located on chromosome segments of Hordeum chilense translocated into wheat chromosomes. No translocations from wheat into H. chilense chromosomes were observed. Both HMW-GS transgenes were expressed at high levels in the endosperm of transgenic plants. The analysis by FISH of transgenic plants allowed the early detection of homozygous and heterozygous plants. The consequences and implications of translocations on breeding are discussed.

  10. X Chromosome Inactivation and Breast Cancer: Epigenetic Alteration in Tumor Initiation and Progression

    DTIC Science & Technology

    2007-09-01

    types of mammary tumors, but not others. For instance, X chromosomal abnormalities appear to be associated with basal -like human breast cancer (BLC...with the Xi (Richardson et al., 2006). Seventeen of thirty-eight BRCA1WT basal -like (BLC) and non-BLC tumor samples lacked detectable XIST RNA...chromosome, suggesting loss of the XIST-expressing Xi in these cells. Twelve of the sporadic, basal -like tumor samples also showed a loss of heterozygous X

  11. Chromosomal mosaicism of extraembryonic cells detected by prenatal diagnosis

    SciTech Connect

    Zolotukhina, T.V.; Shilova, N.V.

    1995-09-01

    Data on detection of chromosomal mosaicism in amniotic cells and chorionic villi obtained by prenatal cytogenetic diagnosis are presented. The frequency of chromosomal mosaicism in preparations of amniotic fluid cell culture was 2.6% (6 out of 226), and that in {open_quotes}direct{close_quotes} villus preparations was 1.6% (13 out of 774). The necessity to perform an additional analysis of other fetal cells or neonatal lymphocytes to specify the diagnosis was shown. The analysis of the outcome of pregnancies during which chromosomal mosaicism in the extraembryonic cells was detected indicates that these women form a high-risk group, both genetically and obstetrically; in only 8 out of 19 cases did pregnancies end in normal deliveries at term; in three cases, spontaneous abortions occurred at 16-31 weeks of gestation; in three cases, the pregnancies were terminated due to fetal chromosomal aberrations in nonmosaic form; the outcome of pregnancy in five cases was preterm delivery of an underweight newborn. 26 refs., 1 tab.

  12. Telomere shortening relaxes X chromosome inactivation and forces global transcriptome alterations.

    PubMed

    Schoeftner, Stefan; Blanco, Raquel; Lopez de Silanes, Isabel; Muñoz, Purificación; Gómez-López, Gonzalo; Flores, Juana M; Blasco, Maria A

    2009-11-17

    Telomeres are heterochromatic structures at chromosome ends essential for chromosomal stability. Telomere shortening and the accumulation of dysfunctional telomeres are associated with organismal aging. Using telomerase-deficient TRF2-overexpressing mice (K5TRF2/Terc(-/-)) as a model for accelerated aging, we show that telomere shortening is paralleled by a gradual deregulation of the mammalian transcriptome leading to cumulative changes in a defined set of genes, including up-regulation of the mTOR and Akt survival pathways and down-regulation of cell cycle and DNA repair pathways. Increased DNA damage from dysfunctional telomeres leads to reduced deposition of H3K27me3 onto the inactive X chromosome (Xi), impaired association of the Xi with telomeric transcript accumulations (Tacs), and reactivation of an X chromosome-linked K5TRF2 transgene that is subjected to X-chromosome inactivation in female mice with sufficiently long telomeres. Exogenously induced DNA damage also disrupts Xi-Tacs, suggesting DNA damage at the origin of these alterations. Collectively, these findings suggest that critically short telomeres activate a persistent DNA damage response that alters gene expression programs in a nonstochastic manner toward cell cycle arrest and activation of survival pathways, as well as impacts the maintenance of epigenetic memory and nuclear organization, thereby contributing to organismal aging.

  13. The clinical application of array CGH for the detection of chromosomal defects in 20,126 unselected newborns

    PubMed Central

    2013-01-01

    Background Array comparative genomic hybridization (CGH) is a powerful tool for detecting unbalanced chromosomal alterations. To validate the usefulness of array CGH in newborn screening, we examined 20,126 unselected infants. In addition, the number of newborns analyzed with array CGH is the largest one ever reported. Findings A total of 20,126 unselected newborns were investigated with array CGH and cytogenetic analyses. The analyses revealed 87 cases with chromosome abnormalities. Of these, 53 cases had significant chromosome aneuploidies, including trisomy 13, trisomy 21, 47,XXY or 45,X, and the other 34 cases presented partial chromosomal deletions or duplications. Conclusions In this study, we show that array CGH is an appropriate tool for the screening of chromosomal abnormalities in newborns, especially for the infants without distinct clinical features. PMID:23725218

  14. Chromosome-specific staining to detect genetic rearrangements associated with chromosome 3 and/or chromosome 17

    DOEpatents

    Gray, Joe W [San Francisco, CA; Pinkel, Daniel [Lafayette, CA; Kallioniemi, Olli-Pekka [Turku, FI; Kallioniemi, Anne [Tampere, FI; Sakamoto, Masaru [Tokyo, JP

    2008-09-09

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  15. Chromosome-specific staining to detect genetic rearrangements associated with chromosome 3 and/or chromosome 17

    DOEpatents

    Gray, Joe W [San Francisco, CA; Pinkel, Daniel [Lafayette, CA; Kallioniemi, Olli-Pekka [Turku, FI; Kallioniemi, Anne [Tampere, FI; Sakamoto, Masaru [Tokyo, JP

    2009-10-06

    Methods and compositions for staining based upon nucleic acid sequence that employ .[.nudeic.]. .Iadd.nucleic .Iaddend.acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  16. Chromosome-specific staining to detect genetic rearrangements associated with chromosome 3 and/or chromosome 17

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel; Kallioniemi, Olli-Pekka; Kallioniemi, Anne; Sakamoto, Masaru

    2002-01-01

    Methods and compositions for staining based upon nucleic acid sequence that employ nudeic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  17. Chromosomal Rainbows detect Oncogenic Rearrangements of Signaling Molecules in Thyroid Tumors

    SciTech Connect

    O'Brien, Benjamin; Jossart, Gregg H.; Ito, Yuko; Greulich-Bode, Karin M.; Weier, Jingly F.; Munne, Santiago; Clark, Orlo H.; Weier, Heinz-Ulrich G.

    2010-08-19

    Altered signal transduction can be considered a hallmark of many solid tumors. In thyroid cancers the receptor tyrosine kinase (rtk) genes NTRK1 (Online Mendelian Inheritance in Man = OMIM *191315, also known as 'TRKA'), RET ('Rearranged during Transfection protooncogene', OMIM *164761) and MET (OMIM *164860) have been reported as activated, rearranged or overexpressed. In many cases, a combination of cytogenetic and molecular techniques allows elucidation of cellular changes that initiate tumor development and progression. While the mechanisms leading to overexpression of the rtk MET gene remain largely unknown, a variety of chromosomal rearrangements of the RET or NTKR1 gene could be demonstrated in thyroid cancer. Abnormal expressions in these tumors seem to follow a similar pattern: the rearrangement translocates the 3'-end of the rtk gene including the entire catalytic domain to an expressed gene leading to a chimeric RNA and protein with kinase activity. Our research was prompted by an increasing number of reports describing translocations involving ret and previously unknown translocation partners. We developed a high resolution technique based on fluorescence in situ hybridization (FISH) to allow rapid screening for cytogenetic rearrangements which complements conventional chromosome banding analysis. Our technique applies simultaneous hybridization of numerous probes labeled with different reporter molecules which are distributed along the target chromosome allowing the detection of cytogenetic changes at near megabase-pair (Mbp) resolution. Here, we report our results using a probe set specific for human chromosome 10, which is altered in a significant portion of human thyroid cancers (TC's). While rendering accurate information about the cytogenetic location of rearranged elements, our multi-locus, multi-color analysis was developed primarily to overcome limitations of whole chromosome painting (WCP) and chromosome banding techniques for fine mapping of

  18. Alteration of chromosome arm 6p is characteristic of primary mediastinal B-cell lymphoma, as identified by genome-wide allelotyping.

    PubMed

    Rigaud, G; Moore, P S; Taruscio, D; Scardoni, M; Montresor, M; Menestrina, F; Scarpa, A

    2001-06-01

    Five cases of primary mediastinal B-cell lymphoma (PMBL) each have been studied with 375 microsatellite markers from all 22 autosomes. Of the 151 genomic alterations among the 1,875 assays, only five were allelic losses. The remainder of the microsatellite alterations consisted of 114 allelic imbalances and 32 instabilities. Microsatellite alterations were found in all cases on chromosomal arms 6p and 9p. These allelic imbalances most likely are indicative of genetic amplification, a finding agreeing well with those of studies using either comparative genomic hybridization or arbitrarily primed polymerase chain reaction, in which amplification of chromosome arm 9p in PMBL has been found. The allelic imbalances on chromosome arm 6p always included marker D6S276 located at 6p21.3-p22.3, where the MHC class I genes reside. These allelic imbalances may be reflective of alterations in the expression of the MHC gene products, characteristic of PMBL. Allelic anomalies close to the MYB gene locus on 6q were detected in two cases and prompted the analysis of MYB rearrangements in a series of 30 lymphomas. One rearrangement was detected in one of 18 cases of PMBL and in none of 10 diffuse, large B-cell lymphomas and two T-cell lymphomas. Our genome-wide microsatellite analysis provides independent confirmation that PMBL is characterized by infrequent chromosomal losses and by frequent genetic alterations involving chromosomal arm 9p. For the first time, chromosomal arm 6p has been identified as a highly frequent target of genetic alterations in this tumor type. Finally, MYB may also be involved occasionally in PMBL pathogenesis. Copyright 2001 Wiley-Liss, Inc.

  19. Chromosomal abnormalities in fetuses with ultrasonographically detected neural tube defects.

    PubMed

    Kanit, Hakan; Özkan, Azra Arici; Öner, Soner Recai; Ispahi, Ciğdem; Endrikat, Jan Siegfried; Ertan, Kubilay

    2011-10-01

    We analyzed the karyotype of fetuses with ultrasonographically detected neural tube defects (NTDs). In our study, we included a total of 194 fetuses with NTDs. We analyzed the type of NTD, the karyotype, maternal age, fetal gestational age at diagnosis, and fetal sex. Of the 194 fetuses with NTDs, 87 were anencephalic and 107 had other, nonanencephalic, NTDs. A total of 12 fetuses were shown to have chromosomal abnormalities. Three of 87 anencephalic fetuses (3.45%) had chromosomal abnormalities. The sex ratio for anencephalic fetuses was 65.5% : 34.5% for female and male fetuses. Nine of 107 fetuses with other NTDs (8.41%) had chromosomal abnormalities. Seven fetuses had isolated NTDs and a further seven fetuses had additional ultrasonographic anomalies. Two of the latter had abnormal karyotypes. The sex ratio of all other NTD cases was 67.3% : 32.7% for female and male fetuses. The high number of chromosomal abnormalities justifies prenatal karyotyping in all fetuses with ultrasonographically diagnosed NTDs.

  20. Chromosome

    MedlinePlus

    Chromosomes are structures found in the center (nucleus) of cells that carry long pieces of DNA. DNA ... is the building block of the human body. Chromosomes also contain proteins that help DNA exist in ...

  1. Single-cell chromosomal imbalances detection by array CGH

    PubMed Central

    Le Caignec, Cedric; Spits, Claudia; Sermon, Karen; De Rycke, Martine; Thienpont, Bernard; Debrock, Sophie; Staessen, Catherine; Moreau, Yves; Fryns, Jean-Pierre; Van Steirteghem, Andre; Liebaers, Inge; Vermeesch, Joris R.

    2006-01-01

    Genomic imbalances are a major cause of constitutional and acquired disorders. Therefore, aneuploidy screening has become the cornerstone of preimplantation, prenatal and postnatal genetic diagnosis, as well as a routine aspect of the diagnostic workup of many acquired disorders. Recently, array comparative genomic hybridization (array CGH) has been introduced as a rapid and high-resolution method for the detection of both benign and disease-causing genomic copy-number variations. Until now, array CGH has been performed using a significant quantity of DNA derived from a pool of cells. Here, we present an array CGH method that accurately detects chromosomal imbalances from a single lymphoblast, fibroblast and blastomere within a single day. Trisomy 13, 18, 21 and monosomy X, as well as normal ploidy levels of all other chromosomes, were accurately determined from single fibroblasts. Moreover, we showed that a segmental deletion as small as 34 Mb could be detected. Finally, we demonstrated the possibility to detect aneuploidies in single blastomeres derived from preimplantation embryos. This technique offers new possibilities for genetic analysis of single cells in general and opens the route towards aneuploidy screening and detection of unbalanced translocations in preimplantation embryos in particular. PMID:16698960

  2. Crowding-Induced Structural Alterations of Random-Loop Chromosome Model

    NASA Astrophysics Data System (ADS)

    Kim, Jun Soo; Backman, Vadim; Szleifer, Igal

    2011-04-01

    We investigate structural alterations of random-loop polymers due to changes in the crowding condition, as a model to study environmental effects on the structure of chromosome subcompartments. The polymer structure is changed in a nonmonotonic fashion with an increasing density of crowders: condensed at small volume fractions; decondensed at high crowding volume fractions. The nonmonotonic behavior is a manifestation of the nontrivial distance dependence of the depletion interactions. We also show that crowding-induced structural alterations affect the access of binding proteins to the surface of polymer segments and are distinguished from structural changes due to the increased number of specific polymer loops.

  3. Chromosomal changes detected by fluorescence in situ hybridization in patients with acute lymphoblastic leukemia.

    PubMed

    Zhang, Lijun; Parkhurst, J B; Kern, W F; Scott, K V; Niccum, D; Mulvihill, J J; Li, Shibo

    2003-09-01

    -T or T cell lineage and the six adults were negative for TEL/AML1 fusion. One patient had double Philadelphia chromosomes, three had a rearrangement or a deletion of the MLL gene, one had t (4;11) and two had a deletion of the MLL. One of the patients with an MLL deletion also had a large ring of chromosome 21, and r (21) was caused by AML1 gene tandemly duplicated at least five times. The second case with the MLL deletion was also unique, the patient had a t (12;21) as well. A total of 20 patients had numerical changes (gain or loss) of chromosomes 4, 10, 17 and 21. Eight patients were found to have trisomies of three or four different chromosomes. Interestingly, seven of these patients did not have TEL/AML1, BCR/ABL or the MLL gene rearrangement; one did have the TEL/AML1 gene fusion. Eleven patients with pro-B cell or B cell type ALL (9 children with ALL, 2 adults with ALL) had numerical changes of chromosome 21 (gain 1 or 2 chromosome 21), among them, 10 patients had no structural alteration of chromosome 21, and one was combined by t (12; 21). Four patients had a monosomy of chromosome 17 and three out of these patients with monosomy 17 also had a fusion signal of TEL/AML1. FISH plays an important role in detecting chromosome changes, especially in some cryptic chromosome translocations and patients with culture failures. This study found a trend towards a division between patients who had structural changes such as t (12;21) or a ring chromosome 21 and those who had numerical changes of chromosome 21 as well as the patients with TEL/AML1 fusion and patients with the coexistence of numerical chromosomal changes of chromosomes 4, 10 and 17. In our opinion there are two separate mechanisms which lead to the development or progression of leukemia.

  4. Automating dicentric chromosome detection from cytogenetic biodosimetry data.

    PubMed

    Rogan, Peter K; Li, Yanxin; Wickramasinghe, Asanka; Subasinghe, Akila; Caminsky, Natasha; Khan, Wahab; Samarabandu, Jagath; Wilkins, Ruth; Flegal, Farrah; Knoll, Joan H

    2014-06-01

    We present a prototype software system with sufficient capacity and speed to estimate radiation exposures in a mass casualty event by counting dicentric chromosomes (DCs) in metaphase cells from many individuals. Top-ranked metaphase cell images are segmented by classifying and defining chromosomes with an active contour gradient vector field (GVF) and by determining centromere locations along the centreline. The centreline is extracted by discrete curve evolution (DCE) skeleton branch pruning and curve interpolation. Centromere detection minimises the global width and DAPI-staining intensity profiles along the centreline. A second centromere is identified by reapplying this procedure after masking the first. Dicentrics can be identified from features that capture width and intensity profile characteristics as well as local shape features of the object contour at candidate pixel locations. The correct location of the centromere is also refined in chromosomes with sister chromatid separation. The overall algorithm has both high sensitivity (85 %) and specificity (94 %). Results are independent of the shape and structure of chromosomes in different cells, or the laboratory preparation protocol followed. The prototype software was recoded in C++/OpenCV; image processing was accelerated by data and task parallelisation with Message Passaging Interface and Intel Threading Building Blocks and an asynchronous non-blocking I/O strategy. Relative to a serial process, metaphase ranking, GVF and DCE are, respectively, 100 and 300-fold faster on an 8-core desktop and 64-core cluster computers. The software was then ported to a 1024-core supercomputer, which processed 200 metaphase images each from 1025 specimens in 1.4 h.

  5. Chromosome-specific staining to detect genetic rearrangements associated with chromosome 3 and/or chromosome 17

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel; Kallioniemi, Olli-Pekka; Kallioniemi, Anne; Sakamoto, Masaru

    2009-10-06

    Methods and compositions for staining based upon nucleic acid sequence that employ nudeic nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  6. Method of detecting genetic deletions identified with chromosomal abnormalities

    SciTech Connect

    Gray, Joe W; Pinkel, Daniel; Tkachuk, Douglas

    2013-11-26

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyzes. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acids probes are typically of a complexity greater tha 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particlularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML) and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar ut genetically different diseases, and for many prognostic and diagnostic applications.

  7. Method of detecting genetic translocations identified with chromosomal abnormalities

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel; Tkachuk, Douglas

    2001-01-01

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML) and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  8. Chromosome-specific staining to detect genetic rearrangements

    SciTech Connect

    Gray, Joe W.; Pinkel, Daniel; Tkachuk, Douglas; Westbrook, Carol

    2013-04-09

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyzes. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML) and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  9. Detection of chromosomal imbalances in transitional cell carcinoma of the bladder by comparative genomic hybridization.

    PubMed Central

    Voorter, C.; Joos, S.; Bringuier, P. P.; Vallinga, M.; Poddighe, P.; Schalken, J.; du Manoir, S.; Ramaekers, F.; Lichter, P.; Hopman, A.

    1995-01-01

    Comparative genomic hybridization (CGH) was applied for a comprehensive screening of chromosomal aberrations in 14 transitional cell carcinomas of the bladder of different grade and stage. The results were compared in a number of selected cases with those obtained by restriction fragment length polymorphism analyses and targeted fluorescence in situ hybridization. Distinct amplifications, found with CGH, were located on 3p22-24, 10p13-14, 12q13-15, 17q22-23, 18p11, and 22q11-13. These high copy number amplifications and the frequency of imbalances involving chromosome 5, occurring in 4 of 14 cases, have not yet been identified in transitional cell carcinomas. Apart from these new aberrations, imbalances were detected in 3 or more cases for chromosomes 9 and 11, as already described previously in the literature. In four tumors, the copy number of specific chromosomal regions was also analyzed by interphase cytogenetics. Although in most instances the CGH data were confirmed, in one tumor, distinct differences were observed, possibly a result of heterogeneity of the tumor cell population. Furthermore, the CGH data were compared with loss of heterozygosity as revealed by restriction fragment length polymorphism analysis in the same tumors. In 80% of informative cases, no loss was detected by restriction fragment length polymorphism or by CGH. Of the 15 cases of loss of heterozygosity, 7 showed a loss also with CGH, whereas in 8 cases no loss was observed. In summary, CGH is a fast method to obtain a comprehensive picture of chromosomal imbalances in transitional cell carcinomas, including a number of previously unknown genomic alterations such as high level amplifications. Images Figure 2 Figure 2 Figure 5 PMID:7778674

  10. Microarray-based comparative genomic hybridization analysis in neonates with congenital anomalies: detection of chromosomal imbalances.

    PubMed

    Emy Dorfman, Luiza; Leite, Júlio César L; Giugliani, Roberto; Riegel, Mariluce

    2015-01-01

    To identify chromosomal imbalances by whole-genome microarray-based comparative genomic hybridization (array-CGH) in DNA samples of neonates with congenital anomalies of unknown cause from a birth defects monitoring program at a public maternity hospital. A blind genomic analysis was performed retrospectively in 35 stored DNA samples of neonates born between July of 2011 and December of 2012. All potential DNA copy number variations detected (CNVs) were matched with those reported in public genomic databases, and their clinical significance was evaluated. Out of a total of 35 samples tested, 13 genomic imbalances were detected in 12/35 cases (34.3%). In 4/35 cases (11.4%), chromosomal imbalances could be defined as pathogenic; in 5/35 (14.3%) cases, DNA CNVs of uncertain clinical significance were identified; and in 4/35 cases (11.4%), normal variants were detected. Among the four cases with results considered causally related to the clinical findings, two of the four (50%) showed causative alterations already associated with well-defined microdeletion syndromes. In two of the four samples (50%), the chromosomal imbalances found, although predicted as pathogenic, had not been previously associated with recognized clinical entities. Array-CGH analysis allowed for a higher rate of detection of chromosomal anomalies, and this determination is especially valuable in neonates with congenital anomalies of unknown etiology, or in cases in which karyotype results cannot be obtained. Moreover, although the interpretation of the results must be refined, this method is a robust and precise tool that can be used in the first-line investigation of congenital anomalies, and should be considered for prospective/retrospective analyses of DNA samples by birth defect monitoring programs. Copyright © 2014 Sociedade Brasileira de Pediatria. Published by Elsevier Editora Ltda. All rights reserved.

  11. Gyramides prevent bacterial growth by inhibiting DNA gyrase and altering chromosome topology.

    PubMed

    Rajendram, Manohary; Hurley, Katherine A; Foss, Marie H; Thornton, Kelsey M; Moore, Jared T; Shaw, Jared T; Weibel, Douglas B

    2014-06-20

    Antibiotics targeting DNA gyrase have been a clinical success story for the past half-century, and the emergence of bacterial resistance has fueled the search for new gyrase inhibitors. In this paper we demonstrate that a new class of gyrase inhibitors, the gyramides, are bacteriostatic agents that competitively inhibit the ATPase activity of Escherichia coli gyrase and produce supercoiled DNA in vivo. E. coli cells treated with gyramide A have abnormally localized, condensed chromosomes that blocks DNA replication and interrupts chromosome segregation. The resulting alterations in DNA topology inhibit cell division through a mechanism that involves the SOS pathway. Importantly, gyramide A is a specific inhibitor of gyrase and does not inhibit the closely related E. coli enzyme topoisomerase IV. E. coli mutants with reduced susceptibility to gyramide A do not display cross-resistance to ciprofloxacin and novobiocin. The results demonstrate that the gyramides prevent bacterial growth by a mechanism in which the topological state of chromosomes is altered and halts DNA replication and segregation. The specificity and activity of the gyramides for inhibiting gyrase makes these compounds important chemical tools for studying the mechanism of gyrase and the connection between DNA topology and bacterial cell division.

  12. Gyramides Prevent Bacterial Growth by Inhibiting DNA Gyrase and Altering Chromosome Topology

    PubMed Central

    2015-01-01

    Antibiotics targeting DNA gyrase have been a clinical success story for the past half-century, and the emergence of bacterial resistance has fueled the search for new gyrase inhibitors. In this paper we demonstrate that a new class of gyrase inhibitors, the gyramides, are bacteriostatic agents that competitively inhibit the ATPase activity of Escherichia coli gyrase and produce supercoiled DNA in vivo. E. coli cells treated with gyramide A have abnormally localized, condensed chromosomes that blocks DNA replication and interrupts chromosome segregation. The resulting alterations in DNA topology inhibit cell division through a mechanism that involves the SOS pathway. Importantly, gyramide A is a specific inhibitor of gyrase and does not inhibit the closely related E. coli enzyme topoisomerase IV. E. coli mutants with reduced susceptibility to gyramide A do not display cross-resistance to ciprofloxacin and novobiocin. The results demonstrate that the gyramides prevent bacterial growth by a mechanism in which the topological state of chromosomes is altered and halts DNA replication and segregation. The specificity and activity of the gyramides for inhibiting gyrase makes these compounds important chemical tools for studying the mechanism of gyrase and the connection between DNA topology and bacterial cell division. PMID:24712739

  13. Micro RNAs and DNA methylation are regulatory players in human cells with altered X chromosome to autosome balance

    PubMed Central

    Rajpathak, Shriram N.; Deobagkar, Deepti D.

    2017-01-01

    The gene balance hypothesis predicts that an imbalance in the dosage sensitive genes affects the cascade of gene networks that may influence the fitness of individuals. The phenotypes associated with chromosomal aneuploidies demonstrate the importance of gene dosage balance. We have employed untransformed human fibroblast cells with different number of X chromosomes to assess the expression of miRNAs and autosomal genes in addition to the DNA methylation status. High throughput NGS analysis using illumina Next seq500 has detected several autosomal as well as X linked miRNAs as differentially expressed in X monosomy and trisomy cells. Two of these miRNAs (hsa-miR-125a-5p and 335-5p) are likely to be involved in regulation of the autosomal gene expression. Additionally, our data demonstrates altered expression and DNA methylation signatures of autosomal genes in X monosomy and trisomy cells. In addition to miRNAs, expression of DNMT1 which is an important epigenetic player involved in many processes including cancer, is seen to be altered. Overall, present study provides a proof for regulatory roles of micro RNAs and DNA methylation in human X aneuploidy cells opening up possible new ways for designing therapeutic strategies. PMID:28233878

  14. Molecular cytogenetic detection of chromosome 15 deletions in patients with Prader-Willi and Angelman syndromes

    SciTech Connect

    Chadwick, D.E.; Weksberg, R.; Shuman, C.

    1994-09-01

    Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are clinically distinct genetic disorders involving alterations of chromosome 15q11-q13. Approximately 75% of individuals with PWS and AS have deletions within 15q11-q13 by molecular analysis. We have evaluated fluorescence in situ hybridization (FISH) for the clinical laboratory detection of del(15)(q11q13) using the cosmid probes D15S11 and GABRB3 (ONCOR, Gaithersburg, NY). 4/4 PWS and 1/1 AS patients previously identified as having cytogenetic deletions were deleted for both probes. In a prospectively ascertained series of 54 patient samples referred to rule out either PWS or AS, 8 were deleted for D15S11 and GABRB3. In addition, an atypical deletion patient with PWS was also identified who was found to be deleted for GABRB3 but not D15S11. The SNRPN locus was also deleted in this patient. Only 4 of the 9 patient samples having molecular cytogenetic deletions were clearly deleted by high resolution banding (HRB) analysis. The microscopic and submicroscopic deletions have been confirmed by dinucleotide (CA) repeat analysis. Microsatellite polymorphism analysis was also used to demonstrate that five non-deletion patients in this series had biparental inheritance of chromosome 15, including region q11-q13. Deletions were not detected by either HRB, FISH or microsatellite polymorphism analysis in samples obtained from parents of the deletion patients. Methylation studies of chromosome 15q11-q13 are in progress for this series of PWS and AS families. FISH analysis of chromosome 15q11-q13 in patients with PWS and AS is a rapid, sensitive and reliable method for deletion detection.

  15. A 11.7-Mb Paracentric Inversion in Chromosome 1q Detected in Prenatal Diagnosis Associated with Familial Intellectual Disability.

    PubMed

    Rigola, Maria A; Baena, Neus; Català, Vicenç; Lozano, Iris; Gabau, Elisabet; Guitart, Miriam; Fuster, Carmen

    2015-01-01

    Most apparent balanced chromosomal inversions are usually clinically asymptomatic; however, infertility, miscarriages, and mental retardation have been reported in inversion carriers. We present a small family with a paracentric inversion 1q42.13q43 detected in routine prenatal diagnosis. Molecular cytogenetic methods defined the size of the inversion as 11.7 Mb and excluded other unbalanced chromosomal alterations in the patients. Our findings suggest that intellectual disability is caused by dysfunction, disruption, or position effects of genes located at or near the breakpoints involved in this inversion.

  16. X-ray induced visible alterations in the giant chromosomes of Phryne cincta (Nematocera, Diptera): relation of radiation sensitivity to pronuclear chromosome structure.

    PubMed

    Israelewski, N

    1975-12-10

    In order to induce chromosomal rearrangements, males were exposed to x-rays and then mated to non-irradiated females. The number of each type of structural alteration was determined by examination of the polytene chromosomes of the F1 progeny. -- A comparison of the results with similar studies made on Drosophila revealed a significantly greater sensitivity in Phryne. Parallel to that an extremely high frequency of small inversions was ascertained in Phryne, and the observed ratio of inversions to translocations was the inverse of that which would be expected from purely mathematical considerations based on the lengths of the different chromosomes. These facts allow the conclusion that the paternal pronuclear chromosomes in Phryne are highly spiralized. Besides, the kinetochore-to-translocation-breakpoint distance was measured in both of the chromosomes involved in each reciprocal translocation and the differences (kinetochore-break distance differences) were registered and from them the arrangement of the chromosomes in the pronucleus of Phryne deduced. The data obtained support the assumption of an ordered, polar-field type of orientation. In Drosophila, in contrast, the comparable data showed that the pronuclear chromosomes are not spiralized and are randomly arranged (Bauer, 1939). -- These results seem to indicate that a close correlation exists between the different radiation sensitivities of Drosophila and Phryne and the different states of spiralisation and arrangements of their chromosomes in the pronucleus stage. It is hypothesized that the influence of the maternal genome on the degree of spiralization of the paternal chromosomes could account for differences in the pronuclear chromosome structure of both species.

  17. High chromosome conservation detected by comparative chromosome painting in chicken, pigeon and passerine birds.

    PubMed

    Derjusheva, Svetlana; Kurganova, Anna; Habermann, Felix; Gaginskaya, Elena

    2004-01-01

    Chicken chromosome paints for macrochromosomes 1-10, Z, and the nine largest microchromosomes (Griffin et al. 1999) were used to analyze chromosome homologies between chicken (Gallus gallus domesticus: Galliformes), domestic pigeon (Columba livia: Columbiformes), chaffinch (Fringilla coelebs Passeriformes), and redwing (Turdus iliacus: Passeriformes). High conservation of syntenies was revealed. In general, both macro- and microchromosomes in these birds showed very low levels of interchromosomal rearrangements. Only two cases of rearrangements were found. Chicken chromosome 1 corresponds to chromosome 1 in pigeon, but to chromosomes 3 and 4 in chaffinch and chromosomes 2 and 5 in redwing. Chicken chromosome 4 was shown to be homologous to two pairs of chromosomes in the karyotypes of pigeon and both passerine species. Comparative analysis of chromosome painting data and the results of FISH with (TTAGGG)n probe did not reveal any correlation between the distribution of interstitial telomere sites (ITSs) and chromosome rearrangements in pigeon, chaffinch and redwing. In chaffinch, ITSs were found to co-localize with a tandem repeat GS (Liangouzov et al. 2002), monomers of which contain an internal TTAGGG motif.

  18. Comparative analyses of chromosome alterations in soft-tissue metastases within and across patients with castration-resistant prostate cancer

    PubMed Central

    Holcomb, Ilona N.; Young, Janet M.; Coleman, Ilsa M.; Salari, Keyan; Grove, Douglas I.; Hsu, Li; True, Lawrence D.; Roudier, Martine P.; Morrissey, Colm M.; Higano, Celestia S.; Nelson, Peter S.; Vessella, Robert L.; Trask, Barbara J.

    2009-01-01

    Androgen deprivation is the mainstay of therapy for progressive prostate cancer. Despite initial and dramatic tumor inhibition, most men eventually fail therapy and die of metastatic castration-resistant (CR) disease. Here, we characterize the profound degree of genomic alteration found in CR tumors using array CGH, gene expression arrays, and FISH. By cluster analysis, we show that the similarity of the genomic profiles from primary and metastatic tumors is driven by the patient. Using data adjusted for this similarity, we identify numerous high-frequency alterations in the CR tumors, such as 8p loss and chromosome 7 and 8q gain. By integrating array CGH and expression array data, we reveal genes whose correlated values suggest they are relevant to prostate cancer biology. We find alterations that are significantly associated with the metastases of specific organ sites, and others with CR tumors versus the tumors of patients with localized prostate cancer not treated with androgen deprivation. Within the high-frequency sites of loss in CR metastases, we find an over-representation of genes involved in cellular lipid metabolism, including PTEN. Finally, using FISH we verify the presence of a gene fusion between TMPRSS2 and ERG suggested by chromosome-21 deletions detected by array CGH. We find the fusion in 54% of our CR tumors, and 81% of the fusion-positive tumors contain cells with multiple copies of the fusion. Our investigation lays the foundation for a better understanding of and possible therapeutic targets for CR disease, the poorly responsive and final stage of prostate cancer. PMID:19773449

  19. Centromeric association of small supernumerary marker chromosomes with their sister-chromosomes detected by three dimensional molecular cytogenetics

    PubMed Central

    2012-01-01

    Background Small supernumerary marker chromosomes (sSMC) are detected in 0.043% of general population and can be characterized for their chromosomal origin, genetic content and shape by molecular cytogenetic approaches. Even though recently progress was achieved towards genotype-phenotype-correlations of sSMC, nothing is known on the influence that an additional derivative extra chromosome has on the nuclear architecture. Results Here we present the first three-dimensional interphase fluorescence in situ hybridization (FISH) studies for the nuclear architecture of sSMC. It could be shown that sSMC derived from chromosomes 15, 16 or 18 preferentially colocalized with one of their corresponding sister chromosomes. This was true in B- and T-lymphocytes as well as in skin fibroblasts. Additionally, a case with a complex sSMC with a karyotype 47,XY,+der(18)t(8;18)(8p23.2 ~ 23.1;18q11.1) was studied. Here the sSMC co-localized with one homologous chromosome 8 instead of 18. Conclusion Overall, there is a kind of "attraction" between an sSMC and one of its homologous sister chromosomes. This seems to be transmitted by the euchromatic part of the sSMC rather than its heterochromatic one. PMID:22413994

  20. Enhanced expression of epidermal growth factor receptor correlates with alterations of chromosome 7 in human pancreatic cancer.

    PubMed Central

    Korc, M; Meltzer, P; Trent, J

    1986-01-01

    Recently, the gene for the epidermal growth factor (EGF) receptor has been mapped to chromosome 7p, the short arm of chromosome 7 [Shimizu, N., Kondo, I., Gamou, M. A., Behzadian, A. & Shimizu, Y. (1984) Somatic Cell Mol. Genet. 10, 45-53]. Utilizing EGF binding in saturation studies, karyology, and cDNA hybridization experiments, we have sought to determine whether there is a correlation between dosage or alteration of chromosome 7 and enhanced expression of EGF receptor in cultured human pancreatic carcinoma cells. Saturation binding studies with 125I-labeled EGF were performed at 4 degrees C with four established human pancreatic cancer cell lines: T3M4, PANC-1, COLO 357, and UACC-462. Analysis of binding data revealed enhanced numbers of EGF receptors in all four cell lines. Chromosome banding analysis revealed clonal structural alterations of chromosome 7p in the cell lines T3M4, PANC-1, and COLO 357, whereas UACC-462 displayed multiple copies of chromosome 7. Hybridization studies using a radiolabeled EGF receptor cDNA probe failed to demonstrate DNA sequence amplification in any cell line but confirmed the presence of EGF receptor mRNA in these cells in approximate proportion to EGF receptor number. Our results suggest that enhanced expression of EGF receptor in human pancreatic cancer can be associated with either structural or numerical alterations of chromosome 7. Images PMID:3014534

  1. Method for detecting a pericentric inversion in a chromosome

    DOEpatents

    Lucas, Joe N.

    2000-01-01

    A method is provided for determining a clastogenic signature of a sample of chromosomes by quantifying a frequency of a first type of chromosome aberration present in the sample; quantifying a frequency of a second, different type of chromosome aberration present in the sample; and comparing the frequency of the first type of chromosome aberration to the frequency of the second type of chromosome aberration. A method is also provided for using that clastogenic signature to identify a clastogenic agent or dosage to which the cells were exposed.

  2. Nonrandom chromosomal imbalances in primary mediastinal B-cell lymphoma detected by arbitrarily primed PCR fingerprinting.

    PubMed

    Scarpa, A; Taruscio, D; Scardoni, M; Iosi, F; Paradisi, S; Ennas, M G; Rigaud, G; Moore, P S; Menestrina, F

    1999-11-01

    We used arbitrarily primed polymerase chain reaction (AP-PCR) fingerprinting to identify chromosomal imbalances in six primary mediastinal B-cell lymphomas (PMBLs). Seventy-four chromosomal imbalances were detected, consisting of 49 sequence gains and 25 losses. Amplifications on chromosome X were seen in five cases, four of which involved the same chromosomal locus. Nonrandom gains at the same locus were also identified on chromosomes 2 and 7 in four cases and on chromosomes 5, 9, and 12 in three cases. Five PMBLs were also analyzed by comparative genomic hybridization (CGH), which found chromosome arm 9p amplification as the only nonrandom imbalance. Our data demonstrate that chromosomal amplifications outnumber losses in PMBL. These mainly involve chromosomes 9 and X and may reflect more complex phenomena, such as translocations or other chromosomal rearrangements, as AP-PCR found coexistent gains and losses on these chromosomes. Comparison between AP-PCR and CGH suggests that anomalies affecting the same chromosomal regions may occur at much higher frequencies than expected by CGH, suggesting that genomic amplifications are usually confined to DNA segments smaller than the megabase long segments required for detection in CGH. Modest increases in genetic material may be as effective as higher-level amplifications when affecting sites where a proto-oncogene resides. Copyright 1999 Wiley-Liss, Inc.

  3. New Insights in the Cytogenetic Practice: Karyotypic Chaos, Non-Clonal Chromosomal Alterations and Chromosomal Instability in Human Cancer and Therapy Response.

    PubMed

    Rangel, Nelson; Forero-Castro, Maribel; Rondón-Lagos, Milena

    2017-06-03

    Recently, non-clonal chromosomal alterations previously unappreciated are being proposed to be included in cytogenetic practice. The aim of this inclusion is to obtain a greater understanding of chromosomal instability (CIN) and tumor heterogeneity and their role in cancer evolution and therapy response. Although several genetic assays have allowed the evaluation of the variation in a population of cancer cells, these assays do not provide information at the level of individual cells, therefore limiting the information of the genomic diversity within tumors (heterogeneity). The karyotype is one of the few available cytogenetic techniques that allow us not only to identify the chromosomal alterations present within a single cell, but also allows us to profile both clonal (CCA) and non-clonal chromosomal alterations (NCCAs). A greater understanding of CIN and tumor heterogeneity in cancer could not only improve existing therapeutic regimens but could also be used as targets for the design of new therapeutic approaches. In this review we indicate the importance and significance of karyotypic chaos, NCCAs and CIN in the prognosis of human cancers.

  4. A new momenclature for structural aberrations detected by chromosome painting

    SciTech Connect

    Tucker, J.D.; Morgan, W.F.; Awa, A.A.; Bauchinger, M.; Blakey, D.; Cornforth, N.N.; Littlefield, L.G.; Natarajan, A.T.; Shasserre, C.

    1994-12-31

    The advent of chromosome painting has brought the realization that structural aberrations can be far more complex than previously imagined. Different laboratories have devised their own nomenclature systems to deal with this complexity, with the result that the terminology has become inconsistent and confusing. Recently, an international group of cytogeneticists experienced with chromosome painting convened to address this issue. The result is a systematic nomenclature capable of describing chromosome aberrations occurring between painted and unpainted chromosomes, as well as aberrations involving only painted chromosomes. The nomenclature is flexible enough to describe accurately even the most extensively rearranged chromosomes. As a consequence of this flexibility, the scheme upon which the nomenclature is based differs substantially from other systems of aberration classification. We call this system the Protocol for Aberration Identification and Nomenclature Terminology (PAINT).

  5. Altered segregation pattern and numerical chromosome abnormalities interrelate in spermatozoa from Robertsonian translocation carriers.

    PubMed

    Godo, Anna; Blanco, Joan; Vidal, Francesca; Sandalinas, Mireia; Garcia-Guixé, Elena; Anton, Ester

    2015-07-01

    The aim of this study was to assess whether there is a relationship between numerical chromosome abnormalities and certain segregation modes in spermatozoa from Robertsonian translocation carriers. A sequential fluorescence in-situ hybridization protocol based on two successive hybridization rounds was performed on sperm samples from one t(13;22) and ten t(13;14) carriers. Patient inclusion criteria included the presence of a positive interchromosomal effect (ICE). In the first round, numerical abnormalities for chromosomes 15/22, 18, 21, X and Y were analysed. In the second round, the segregation outcome of the rearranged chromosomes was evaluated in the numerically abnormal spermatozoa detected in the first round, as well as in randomly assessed spermatozoa. Aneuploid spermatozoa showed statistical differences in all segregation modes when compared with randomly assessed spermatozoa: alternate (50.7% versus 84.3%), adjacent (36.6% versus 14.6%) and 3:0 (10.2% versus 1%). Diploid/multiple disomic spermatozoa showed differences in alternate (3.7% versus 84.3%) and 3:0 (67.6% versus 1%). We concluded that in Robertsonian translocation carriers that exhibit ICE, numerically abnormal spermatozoa preferentially contain unbalanced segregation products. This might be explained by heterosynapsis acting as a rescue mechanism that would lead to aberrant recombination, which is a predisposing factor for non-disjunction events.

  6. Structural chromosomal anomalies detected by prenatal genetic diagnosis: our experience.

    PubMed

    Farcaş, Simona; Crişan, C D; Andreescu, Nicoleta; Stoian, Monica; Motoc, A G M

    2013-01-01

    The prenatal diagnosis is currently widely spread and facilitates the acquiring of important genetic information about the fetus by a rate extremely accelerate and considered without precedent. In this paper, we like to present our experience concerning the genetic diagnosis and counseling offered for pregnancies in which a structural chromosomal aberration was found. The study group is formed by 528 prenatal samples of amniotic fluid and chorionic villi, received by our laboratory from 2006 through October 2012 for cytogenetic diagnosis. The appropriate genetic investigation was selected based on the indications for prenatal diagnosis. The cases with structural chromosomal anomalies and polymorphic variants were analyzed as regard to the maternal age, gestational age, referral indications and type of chromosomal anomaly found. A total number of 21 structural chromosomal anomalies and polymorphic variants were identified in the study group. Out of 21 structural chromosomal anomalies and polymorphic variants, six deletions and microdeletions, four situations with abnormal long "p" arm of acrocentric chromosomes, two duplications, two reciprocal translocations, two inversions, two additions, one Robertsonian translocation associating trisomy 13, one 9q heteromorphism and one complex chromosome rearrangement were noticed. To the best of our knowledge, this is the first Romanian study in which the diagnostic strategies and the management of the prenatal cases with structural rearrangements are presented. The data provided about the diagnosis strategy and the management of the prenatal cases with structural chromosomal anomalies represents a useful tool in genetic counseling of pregnancies diagnosed with rare structural chromosomal anomalies.

  7. Therapeutic strategies in male breast cancer: clinical implications of chromosome 17 gene alterations and molecular subtypes.

    PubMed

    Schildhaus, Hans-Ulrich; Schroeder, Lars; Merkelbach-Bruse, Sabine; Binot, Elke; Büttner, Reinhard; Kuhn, Walther; Rudlowski, Christian

    2013-12-01

    Male breast cancer (MBC) is a rare disease. To date, therapy is mainly based on studies and clinical experiences with breast cancer in women. Only little is known about molecular typing of MBC, particularly with regard to potential biological predictors for adjuvant therapy. In female breast cancer tumors with chromosome 17 centromere (CEP17) duplication, HER2 and/or Topoisomerase II alpha (Topo II-α) gene alterations have been suggested to be associated with poor prognosis and increased sensitivity to anthracycline-containing regimens. In a well characterized cohort of 96 primary invasive MBC, we studied CEP17, HER2 and Topo II-α alterations by fluorescence in-situ hybridization (FISH), and expression of hormone receptors (HR), HER2 and Ki67 by immunohistochemistry to define molecular subtypes. Tumor characteristics and follow-up data were available and correlated with molecular findings. HER2 amplification and Topo II-α amplification/deletion were exceptionally rare in MBC (6.3% and 3.1%, respectively). CEP17 polysomy were found in 9.4% of tumors. HER2, Topo II-α and CEP17 gene alterations were not correlated to patients outcome. 96.9% of our cases were HR positive. Triple negative tumors were found in only 3.1% of the cases. In nodal negative tumors luminal A subtypes were significantly associated with better overall survival. Our results provide evidence for a predominant male breast cancer phenotype, characterized by HR expression and a lack of HER2/Topo II-α alterations and CEP17 duplicates. Therefore, the impact of anthracycline sensitivity linked to HER2/Topo II-α alterations as found in female breast cancer has low clinical significance for this specific male breast cancer phenotype. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Altered interphase fluorescence in situ hybridization profiles of chromosomes 4, 8q24, and 9q34 in pancreatic ductal adenocarcinoma are associated with a poorer patient outcome.

    PubMed

    Gutiérrez, María L; Muñoz-Bellvis, Luis; Sarasquete, María E; Hernández-Mejía, David G; Abad, María del Mar; Bengoechea, Oscar; Corchete, Luis; González-González, María; García-García, Jacinto; Gonzalez, Marcos; Mota, Ines; Orfao, Alberto; Sayagues, José M

    2014-11-01

    Most patients with pancreatic ductal adenocarcinoma (PDAC) die within 6 months of diagnosis. However, 20% to 25% patients undergoing total tumor resection remain alive and disease-free 5 years after diagnostic surgery. Few studies on tumor markers have predicted patient prognosis and/or survival. We evaluated the effect of tumor cytogenetic copy number changes detected by interphase fluorescence in situ hybridization on overall survival (OS) of 55 PDAC patients. The prognostic value of copy number changes showing an effect on OS was validated in an external cohort of 44 surgically resected PDAC patients by comparative genomic hybridization arrays, and the genes coded in altered chromosomes with prognostic value were identified by high-density single-nucleotide polymorphism arrays in 20 cases. Copy number changes of chromosomes 4 and 9q34 with gains of 8q24 were independently associated with shorter OS. On the basis of these three chromosomal alterations, a score is proposed that identifies patients with significantly different (P < 0.001) 5-year OS rates: 60% ± 20%, 16% ± 8%, and 0% ± 0%, respectively. Our results show an association between tumor cytogenetics and OS of PDAC patients and provide the basis for further prognostic stratification of patients undergoing complete tumor resection. Further studies to identify specific genes coded in these chromosomes and their functional consequences are necessary to understand the clinical effect of these changes.

  9. Analysis of protein gene products in cells with altered chromosome sets for the purpose of genetic mapping

    SciTech Connect

    Shishkin, S.S.; Zakharov, S.F.; Gromov, P.S.; Shcheglova, M.V.; Kukharenko, V.I.; Shilov, A.G.; Matveeva, N.M.; Zhdanova, N.S.; Efimochkin, A.S.; Krokhina, T.B. |

    1994-12-01

    Two-dimensional electrophoresis was used for analyzing proteins in hybrid cells that contained single human chromosomes (chromosome 5, chromosome 21, or chromosomes 5 and 21) against the background of the mouse genome. By comparing the protein patterns of hybrid and parent cells (about 1000 protein fractions for each kind of cell), five fractions among proteins of hybrid cells were supposedly identified as human proteins. The genes of two of them are probably located on chromosome 5, and those of the other three on chromosome 21. Moreover, analysis of proteins in fibroblasts of patients with the cri-du-chat syndrome (5p-) revealed a decrease in the content of two proteins as compared with those in preparations of diploid fibroblasts. This fact was regarded as evidence that two corresponding genes are located on the short arm of chromosome 5. Methodological problems associated with the use of protein pattern analysis in cells with altered chromosome sets for the purposes of genetic mapping are discussed.

  10. Genomic alterations in cervical carcinoma: Losses of chromosome heterozygosity (LOH) correlated with cytogenetic, HPV, and p53 evaluations

    SciTech Connect

    Klinger, H.P.; Mullokandov, M.; Khollodilov, N.G.

    1994-09-01

    This study was undertaken to obtain indications of chromosomes likely to carry tumorigenicity suppressor genes the loss of function of which play a role in the origin or progression of cervical carcinomas. PCR and electrophoresis with primers for 73 highly polymorphic microsatellite chromosome markers were used to determine the incidence of LOH of all of the autosomes in 38 cervical carcinomas. According to these criteria 14 of the autosomes are involved in LOH in 24% to 42% of the tumors. This involves chromosomes 1, 2, 3, 4, 5, 6, 7, 8, 9, 11, 13, 16, 18 and 19. Most frequently involved are chromosomes 3 and 6 with LOH in 42% of the tumors. The chromosomes next most frequently involved are 4, 7, 11 and 18, with LOH in 31-32% of cases. Chromosomes 1, 2, 5, 8 and 16 each had LOH in 29% of the tumors; 9 and 13 each in 26%; and 19 in 24% of the tumors. All other autosomes had LOH in 18% or fewer of the tumors. Cytogenetic analyses performed on direct preparations from many of the same tumors agreed well with the molecular LOH assays. Correlation of the information obtained with both of these methods provides considerable insight into the mechanisms involved in the occurrence of these chromosome alterations. Chromosome 3 is the third most frequent chromosome involved in LOH in all types of cancer. In cervical carcinomas the region most frequently involved is 3p13-p25, which is a segment within which suppressors have been implicated in several other types of malignancies. Chromosome 6 on the other hand is rarely involved in other neoplasias and this appears to be unique to cervical carcinomas. Of interest was the finding that many of the HPV-negative tumors had LOH of chromosome 17 and many of these expressed mutant p53. The latter tumors occur in older women and are on the average much more aggressive than the HPV-positive tumors.

  11. Detection and Automated Scoring of Dicentric Chromosomes in Nonstimulated Lymphocyte Prematurely Condensed Chromosomes After Telomere and Centromere Staining

    SciTech Connect

    M'kacher, Radhia; El Maalouf, Elie; Terzoudi, Georgia; Ricoul, Michelle; Heidingsfelder, Leonhard; Karachristou, Ionna; Laplagne, Eric; Hempel, William M.; Colicchio, Bruno; Dieterlen, Alain; Pantelias, Gabriel; Sabatier, Laure

    2015-03-01

    Purpose: To combine telomere and centromere (TC) staining of premature chromosome condensation (PCC) fusions to identify dicentrics, centric rings, and acentric chromosomes, making possible the realization of a dose–response curve and automation of the process. Methods and Materials: Blood samples from healthy donors were exposed to {sup 60}Co irradiation at varying doses up to 8 Gy, followed by a repair period of 8 hours. Premature chromosome condensation fusions were carried out, and TC staining using peptide nucleic acid probes was performed. Chromosomal aberration (CA) scoring was carried out manually and automatically using PCC-TCScore software, developed in our laboratory. Results: We successfully optimized the hybridization conditions and image capture parameters, to increase the sensitivity and effectiveness of CA scoring. Dicentrics, centric rings, and acentric chromosomes were rapidly and accurately detected, leading to a linear-quadratic dose–response curve by manual scoring at up to 8 Gy. Using PCC-TCScore software for automatic scoring, we were able to detect 95% of dicentrics and centric rings. Conclusion: The introduction of TC staining to the PCC fusion technique has made possible the rapid scoring of unstable CAs, including dicentrics, with a level of accuracy and ease not previously possible. This new approach can be used for biological dosimetry in radiation emergency medicine, where the rapid and accurate detection of dicentrics is a high priority using automated scoring. Because there is no culture time, this new approach can also be used for the follow-up of patients treated by genotoxic therapy, creating the possibility to perform the estimation of induced chromosomal aberrations immediately after the blood draw.

  12. Detection of unbalanced chromosome segregations in preimplantation genetic diagnosis of translocations by short comparative genomic hibridization.

    PubMed

    Rius, Mariona; Obradors, Albert; Daina, Gemma; Ramos, Laia; Pujol, Aïda; Martínez-Passarell, Olga; Marquès, Laura; Oliver-Bonet, Maria; Benet, Jordi; Navarro, Joaquima

    2011-07-01

    To apply a comprehensive chromosomal screening through short comparative genomic hybridization (CGH) in the preimplantation genetic diagnosis (PGD) of translocations. Clinical research study. A PGD laboratory and two IVF clinics. Three Robertsonian translocation carriers, two reciprocal translocation carriers, and a double-translocation carrier. After using the short-CGH approach in the reanalysis of two unbalanced embryos, discarded from a PGD for a reciprocal translocation carrier, the same method was applied in the PGD of day-3 embryos of translocation carriers. Ability of short CGH to detect partial chromosomal abnormalities in unbalanced embryos, translocation segregation proportions, and proportion of embryos carrying chromosomal abnormalities not related to the translocations. The short-CGH technique detected errors resulting from the meiotic segregation of the chromosomes involved in the translocations and other abnormalities affecting the remaining chromosomes. Alternate segregation was detected most frequently among Robertsonian translocation cases, whereas unbalanced chromosome segregations were found predominantly in reciprocal ones. Aneuploidy and structural chromosome errors were found more frequently in Robertsonian than in reciprocal translocation carriers. Application of short-CGH PGD achieved pregnancy in two cases. Short CGH is a reliable approach for PGD of translocations, as it is capable of detecting partial chromosome errors caused by unbalanced segregations simultaneously to the screening of all chromosomes, and it may improve the results after PGD for translocation carriers. Copyright © 2011 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  13. X-chromosome tiling path array detection of copy number variants in patients with chromosome X-linked mental retardation

    PubMed Central

    Madrigal, I; Rodríguez-Revenga, L; Armengol, L; González, E; Rodriguez, B; Badenas, C; Sánchez, A; Martínez, F; Guitart, M; Fernández, I; Arranz, JA; Tejada, MI; Pérez-Jurado, LA; Estivill, X; Milà, M

    2007-01-01

    Background Aproximately 5–10% of cases of mental retardation in males are due to copy number variations (CNV) on the X chromosome. Novel technologies, such as array comparative genomic hybridization (aCGH), may help to uncover cryptic rearrangements in X-linked mental retardation (XLMR) patients. We have constructed an X-chromosome tiling path array using bacterial artificial chromosomes (BACs) and validated it using samples with cytogenetically defined copy number changes. We have studied 54 patients with idiopathic mental retardation and 20 controls subjects. Results Known genomic aberrations were reliably detected on the array and eight novel submicroscopic imbalances, likely causative for the mental retardation (MR) phenotype, were detected. Putatively pathogenic rearrangements included three deletions and five duplications (ranging between 82 kb to one Mb), all but two affecting genes previously known to be responsible for XLMR. Additionally, we describe different CNV regions with significant different frequencies in XLMR and control subjects (44% vs. 20%). Conclusion This tiling path array of the human X chromosome has proven successful for the detection and characterization of known rearrangements and novel CNVs in XLMR patients. PMID:18047645

  14. Chromosome alterations in breast carcinomas: frequent involvement of DNA losses including chromosomes 4q and 21q.

    PubMed Central

    Schwendel, A.; Richard, F.; Langreck, H.; Kaufmann, O.; Lage, H.; Winzer, K. J.; Petersen, I.; Dietel, M.

    1998-01-01

    Comparative genomic hybridization was applied to map DNA gains and losses in 39 invasive ductal breast carcinomas. Frequent abnormalities included gains on chromosomal regions 1q, 8q, 11q12-13, 16p, 19, 20q and X as well as frequent losses on 1p, 5q, 6q, 9p, 11q, 13q and 16q. Furthermore, frequent losses on 4q (20 cases) and 21q (14 cases) were found for the first time in this tumour type. High copy number amplifications were observed at 8q12-24, 11q11-13 and 20q13-ter. Highly differentiated tumours were associated with gains on 1q and 11q12-13 along with losses on 1p21-22, 4q, 13q, 11q21-ter. Undifferentiated breast carcinomas were characterized by additional DNA imbalances, i.e. deletions of 5q13-23, all of chromosome 9, the centromeric part of chromosome 13 including band 13q14 and the overrepresentation of chromosome X. We speculate that these changes are associated with tumour progression of invasive ductal breast cancer. Images Figure 2 Figure 3 PMID:9743305

  15. Chromosomal rearrangements detected by FISH and G-banding.

    PubMed

    Hou, J W; Wang, T R

    1996-09-01

    Fluorescence in situ hybridization (FISH) using chromosome-specific DNA libraries as painting probes, locus-specific unique sequence (cosmid) probes, and Y-specific repetitive sequences was applied in the analysis of eighteen cases of chromosomal rearrangements of undetermined nature. FISH clarified the origin of the extra or translocated chromosome segments in seventeen patients, one with 2q+, two with 4q+, one each with 6p+, 7p+, 9q+, 10p+, 11q+ and 12p+, two with 13q+, and one each with 15q+, 17p+, 18p+, 20p+, 21p+ and Yq+, as well as the nature of a de novo supernumerary chromosome marker in a previously reported case. By G-banding and molecular cytogenetic studies of the family members, six cases were determined to have unbalanced translocations inherited from the carrier parent. The extra translocated genetic material may cause specific trisomic syndromes, including partial 6p21.3-p23, 9q32-q34.3, 13q32-q34, 15q24-q26, and 17p11.2-p13 trisomies in those patients. A translocated 21q segment on 12p was shown by a painting probe in a patient with Down features. A patient with cat cry syndrome resulting from a loss of the terminal segment of the short arm of chromosome 5 was confirmed by a cosmid probe showing de novo reciprocal translocation between chromosomes 5 and 18:t(5;18) (p13.3;p11.31). With FISH, the extra material on the rearranged chromosome could also be identified as duplicated or translocated. The FISH technique thus provides a method for the analysis of extra structurally abnormal chromosomes (especially in de novo cases), recognizable syndromes (contiguous gene syndromes) caused by translocated deletion from parental balanced chromosome rearrangements, and supernumerary marker chromosomes. FISH subsequent to G-banding is also of great help in the confirmation of preliminary abnormal G-banded karyotypes after a modified destaining procedure. In conclusion, the combination of G-banding and FISH is very useful in the accurate diagnosis of chromosomal

  16. Sympatric predator detection alters cutaneous respiration in Lymnaea.

    PubMed

    Orr, Mike; Lukowiak, Ken

    2010-01-01

    The ability of an organism to detect a predator and then to take the appropriate vigilance actions is paramount for survival of the species. Lab-reared snails (>250 generations) maintain their ability to detect predators and alter both aerial and cutaneous respiration. However, only the scent of a sympatric predator altered aerial respiration in freshly collected 'wild' snails. Here we test the hypothesis that the detection of a sympatric predator but not an allopatric predator will alter cutaneous respiration in freshly collected 'wild' snails. We find that Alberta snails while altering their cutaneous respiration to the scent of a sympatric predator (tiger salamander) do not alter respiration to the scent of a crayfish (an allopatric predator). In Dutch snails there is a greater alteration to the scent of crayfish (sympatric predator) than to an allopatric predator (tiger salamander).

  17. FISH detection of chromosome 15 deletions in Prader-Willi and Angelman syndromes

    SciTech Connect

    Teshima, I.; Chadwick, D.; Chitayat, D.

    1996-03-29

    We have evaluated fluorescence in situ hybridization (FISH) analysis for the clinical laboratory detection of the 15q11-q13 deletion seen in Prader-Willi syndrome (PWS) and Angelman syndrome (AS) using probes for loci D15S11, SNRPN, D15S10, and GABRB3. In a series of 118 samples from patients referred for PWS or AS, 29 had deletions by FISH analysis. These included two brothers with a paternally transmitted deletion detectable with the probe for SNRPN only. G-banding analysis was less sensitive for deletion detection but useful in demonstrating other cytogenetic alterations in four cases. Methylation and CA-repeat analyses of 15q11-q13 were used to validate the FISH results. Clinical findings of patients with deletions were variable, ranging from newborns with hypotonia as the only presenting feature to children who were classically affected. We conclude that FISH analysis is a rapid and reliable method for detection of deletions within 15q11-q13 and whenever a deletion is found, FISH analysis of parental chromosomes should also be considered. 41 refs., 4 figs., 2 tabs.

  18. Detection of Y Chromosome DNA as Evidence of Semen in Cervicovaginal Secretions of Sexually Active Women

    PubMed Central

    Chomont, Nicolas; Grésenguet, Gérard; Lévy, Michel; Hocini, Hakim; Becquart, Pierre; Matta, Mathieu; Tranchot-Diallo, Juliette; Andreoletti, Laurent; Carreno, Marie-Paule; Kazatchkine, Michel D.; Bélec, Laurent

    2001-01-01

    The detection of traces of semen in cervicovaginal secretions (CVS) from sexually active women practicing unprotected sex is a prerequisite for the accurate study of cervicovaginal immunity. Two semen markers, the prostatic-specific antigen (PSA) and the Y chromosome, were detected in parallel in CVS obtained by a standardized vaginal washing of consecutive women attending the principal medical center for sexually transmitted diseases of Bangui, Central African Republic. PSA was detected by immunoenzymatic capture assay in the cell-free fraction of CVS, and the Y chromosome was detected by a single PCR assay of DNA extracted by silica from the cell fraction (Y PCR). Fifty (19%) cell-free fractions of the 264 β-globin-positive CVS samples were positive for PSA, and 100 (38%) cell fractions of the CVS samples were positive for the Y chromosome. All the 50 (19%) PSA-containing CVS samples were also positive for the Y chromosome. Fifty (19%) CVS samples were positive only for the Y chromosome, with no detectable PSA. The remaining 164 (62%) CVS samples were both PSA and Y chromosome negative. These findings demonstrate that CVS from sexually active women may contain cell-associated semen residues unrecognized by conventional immunoenzymatic assays used to detect semen components. The detection of cell-associated male DNA with a highly sensitive and specific procedure such as Y PCR constitutes a method of choice to detect semen traces in female genital secretions. PMID:11527810

  19. Origin of nuclear and chromosomal alterations derived from the action of an aneugenic agent--Trifluralin herbicide.

    PubMed

    Fernandes, Thaís Cristina Casimiro; Mazzeo, Dânia Elisa Christofoletti; Marin-Morales, Maria Aparecida

    2009-09-01

    Trifluralin is a herbicide capable of interfering in mitotic cell division due to either microtubule depolymerization or alteration in the concentration of calcium ions within the cell. The aim of this study was to investigate the effects of trifluralin in Allium cepa meristematic cells, evaluating the induction mechanisms of the chromosomal and nuclear aberrations. In this study, A. cepa root tips were submitted for 24h treatment to several concentrations of this herbicide and 48 h recovery post-treatment. The results showed that some concentrations of trifluralin can lead to a mitotic index inhibition, besides inducing chromosomal and nuclear alterations throughout the mitotic cycle. Some of the alterations found seem to be resulting from the herbicide action in different phases and in more than one consecutive cell cycle.

  20. Chromosome segregation regulation in human zygotes: altered mitotic histone phosphorylation dynamics underlying centromeric targeting of the chromosomal passenger complex.

    PubMed

    van de Werken, C; Avo Santos, M; Laven, J S E; Eleveld, C; Fauser, B C J M; Lens, S M A; Baart, E B

    2015-10-01

    Are the kinase feedback loops that regulate activation and centromeric targeting of the chromosomal passenger complex (CPC), functional during mitosis in human embryos? Investigation of the regulatory kinase pathways involved in centromeric CPC targeting revealed normal phosphorylation dynamics of histone H2A at T120 (H2ApT120) by Bub1 kinase and subsequent recruitment of Shugoshin, but phosphorylation of histone H3 at threonine 3 (H3pT3) by Haspin failed to show the expected centromeric enrichment on metaphase chromosomes in the zygote. Human cleavage stage embryos show high levels of chromosomal instability. What causes this high error rate is unknown, as mechanisms used to ensure proper chromosome segregation in mammalian embryos are poorly described. In this study, we investigated the pathways regulating CPC targeting to the inner centromere in human embryos. We characterized the distribution of the CPC in relation to activity of its two main centromeric targeting pathways: the Bub1-H2ApT120-Sgo-CPC and Haspin-H3pT3-CPC pathways. The study was conducted between May 2012 and March 2014 on human surplus embryos resulting from in vitro fertilization treatment and donated for research. In zygotes, nuclear envelope breakdown was monitored by time-lapse imaging to allow timed incubations with specific inhibitors to arrest at prometaphase and metaphase, and to interfere with Haspin and Aurora B/C kinase activity. Functionality of the targeting pathways was assessed through characterization of histone phosphorylation dynamics by immunofluorescent analysis, combined with gene expression by RT-qPCR and immunofluorescent localization of key pathway proteins. Immunofluorescent analysis of the CPC subunit Inner Centromere Protein revealed the pool of stably bound CPC proteins was not strictly confined to the inner centromere of prometaphase chromosomes in human zygotes, as observed in later stages of preimplantation development and somatic cells. Investigation of the

  1. Gene alterations at Drosophila inversion breakpoints provide prima facie evidence for natural selection as an explanation for rapid chromosomal evolution

    PubMed Central

    2012-01-01

    Background Chromosomal inversions have been pervasive during the evolution of the genus Drosophila, but there is significant variation between lineages in the rate of rearrangement fixation. D. mojavensis, an ecological specialist adapted to a cactophilic niche under extreme desert conditions, is a chromosomally derived species with ten fixed inversions, five of them not present in any other species. Results In order to explore the causes of the rapid chromosomal evolution in D. mojavensis, we identified and characterized all breakpoints of seven inversions fixed in chromosome 2, the most dynamic one. One of the inversions presents unequivocal evidence for its generation by ectopic recombination between transposon copies and another two harbor inverted duplications of non-repetitive DNA at the two breakpoints and were likely generated by staggered single-strand breaks and repair by non-homologous end joining. Four out of 14 breakpoints lay in the intergenic region between preexisting duplicated genes, suggesting an adaptive advantage of separating previously tightly linked duplicates. Four out of 14 breakpoints are associated with transposed genes, suggesting these breakpoints are fragile regions. Finally two inversions contain novel genes at their breakpoints and another three show alterations of genes at breakpoints with potential adaptive significance. Conclusions D. mojavensis chromosomal inversions were generated by multiple mechanisms, an observation that does not provide support for increased mutation rate as explanation for rapid chromosomal evolution. On the other hand, we have found a number of gene alterations at the breakpoints with putative adaptive consequences that directly point to natural selection as the cause of D. mojavensis rapid chromosomal evolution. PMID:22296923

  2. Gene alterations at Drosophila inversion breakpoints provide prima facie evidence for natural selection as an explanation for rapid chromosomal evolution.

    PubMed

    Guillén, Yolanda; Ruiz, Alfredo

    2012-02-01

    Chromosomal inversions have been pervasive during the evolution of the genus Drosophila, but there is significant variation between lineages in the rate of rearrangement fixation. D. mojavensis, an ecological specialist adapted to a cactophilic niche under extreme desert conditions, is a chromosomally derived species with ten fixed inversions, five of them not present in any other species. In order to explore the causes of the rapid chromosomal evolution in D. mojavensis, we identified and characterized all breakpoints of seven inversions fixed in chromosome 2, the most dynamic one. One of the inversions presents unequivocal evidence for its generation by ectopic recombination between transposon copies and another two harbor inverted duplications of non-repetitive DNA at the two breakpoints and were likely generated by staggered single-strand breaks and repair by non-homologous end joining. Four out of 14 breakpoints lay in the intergenic region between preexisting duplicated genes, suggesting an adaptive advantage of separating previously tightly linked duplicates. Four out of 14 breakpoints are associated with transposed genes, suggesting these breakpoints are fragile regions. Finally two inversions contain novel genes at their breakpoints and another three show alterations of genes at breakpoints with potential adaptive significance. D. mojavensis chromosomal inversions were generated by multiple mechanisms, an observation that does not provide support for increased mutation rate as explanation for rapid chromosomal evolution. On the other hand, we have found a number of gene alterations at the breakpoints with putative adaptive consequences that directly point to natural selection as the cause of D. mojavensis rapid chromosomal evolution.

  3. Detection of skewed X-chromosome inactivation in Fragile X syndrome and X chromosome aneuploidy using quantitative melt analysis.

    PubMed

    Godler, David E; Inaba, Yoshimi; Schwartz, Charles E; Bui, Quang M; Shi, Elva Z; Li, Xin; Herlihy, Amy S; Skinner, Cindy; Hagerman, Randi J; Francis, David; Amor, David J; Metcalfe, Sylvia A; Hopper, John L; Slater, Howard R

    2015-07-01

    Methylation of the fragile X mental retardation 1 (FMR1) exon 1/intron 1 boundary positioned fragile X related epigenetic element 2 (FREE2), reveals skewed X-chromosome inactivation (XCI) in fragile X syndrome full mutation (FM: CGG > 200) females. XCI skewing has been also linked to abnormal X-linked gene expression with the broader clinical impact for sex chromosome aneuploidies (SCAs). In this study, 10 FREE2 CpG sites were targeted using methylation specific quantitative melt analysis (MS-QMA), including 3 sites that could not be analysed with previously used EpiTYPER system. The method was applied for detection of skewed XCI in FM females and in different types of SCA. We tested venous blood and saliva DNA collected from 107 controls (CGG < 40), and 148 FM and 90 SCA individuals. MS-QMA identified: (i) most SCAs if combined with a Y chromosome test; (ii) locus-specific XCI skewing towards the hypomethylated state in FM females; and (iii) skewed XCI towards the hypermethylated state in SCA with 3 or more X chromosomes, and in 5% of the 47,XXY individuals. MS-QMA output also showed significant correlation with the EpiTYPER reference method in FM males and females (P < 0.0001) and SCAs (P < 0.05). In conclusion, we demonstrate use of MS-QMA to quantify skewed XCI in two applications with diagnostic utility.

  4. Heritable alteration of DNA methylation induced by whole-chromosome aneuploidy in wheat.

    PubMed

    Gao, Lihong; Diarso, Moussa; Zhang, Ai; Zhang, Huakun; Dong, Yuzhu; Liu, Lixia; Lv, Zhenling; Liu, Bao

    2016-01-01

    Aneuploidy causes changes in gene expression and phenotypes in all organisms studied. A previous study in the model plant Arabidopsis thaliana showed that aneuploidy-generated phenotypic changes can be inherited to euploid progenies and implicated an epigenetic underpinning of the heritable variations. Based on an analysis by amplified fragment length polymorphism and methylation-sensitive amplified fragment length polymorphism markers, we found that although genetic changes at the nucleotide sequence level were negligible, extensive changes in cytosine DNA methylation patterns occurred in all studied homeologous group 1 whole-chromosome aneuploid lines of common wheat (Triticum aestivum), with monosomic 1A showing the greatest amount of methylation changes. The changed methylation patterns were inherited by euploid progenies derived from the aneuploid parents. The aneuploidy-induced DNA methylation alterations and their heritability were verified at selected loci by bisulfite sequencing. Our data have provided empirical evidence supporting earlier suggestions that heritability of aneuploidy-generated, but aneuploidy-independent, phenotypic variations may have an epigenetic basis. That at least one type of aneuploidy - monosomic 1A - was able to cause significant epigenetic divergence of the aneuploid plants and their euploid progenies also lends support to recent suggestions that aneuploidy may have played an important and protracted role in polyploid genome evolution.

  5. HIGH-RESOLUTION GENOMIC ARRAYS FACILITATE DETECTION OF NOVEL CRYPTIC CHROMOSOMAL LESIONS IN MYELODYSPLASTIC SYNDROMES

    PubMed Central

    O’Keefe, Christine L.; Tiu, Ramon; Gondek, Lukasz P.; Powers, Jennifer; Theil, Karl S.; Kalaycio, Matt; Lichtin, Alan; Sekeres, Mikkael A.; Maciejewski, Jaroslaw P.

    2008-01-01

    Objective Unbalanced chromosomal aberrations are common in myelodysplastic syndromes, and have prognostic implications. An increased frequency of cytogenetic changes may reflect an inherent chromosomal instability due to failure of DNA repair. Therefore, it is likely that chromosomal defects in myelodysplastic syndromes may be more frequent than predicted by metaphase cytogenetics and new cryptic lesions may be revealed by precise analysis methods. Methods We used a novel high-resolution karyotyping technique, array-based comparative genomic hybridization, to investigate the frequency of cryptic chromosomal lesions in a cohort of 38 well-characterized myelodysplastic syndromes patients; results were confirmed by microsatellite quantitative PCR or single nucleotide polymorphism analysis. Results As compared to metaphase karyotyping, chromosomal abnormalities detected by array-based analysis were encountered more frequently and in a higher proportion of patients. For example, chromosomal defects were found in patients with a normal karyotype by traditional cytogenetics. In addition to verifying common abnormalities, previously cryptic defects were found in new regions of the genome. Cryptic changes often overlapped chromosomes and regions frequently identified as abnormal by metaphase cytogenetics. Conclusion The results underscore the instability of the myelodysplastic syndromes genome and highlight the utility of array-based karyotyping to study cryptic chromosomal changes which may provide new diagnostic information. PMID:17258073

  6. Molecular detection of cryptic Y-chromosomal material in patients with Turner syndrome.

    PubMed

    Cortés-Gutiérrez, Elva I; Herrera-Bartolo, Rosalba; Dávila-Rodríguez, Martha I; Palacios-Saucedo, Gerardo C; Vargas-Villarreal, Javier; Romero-Villarreal, Juana B

    2012-10-01

    A systematic search for a hidden Y-chromosome mosaicism, in Turner syndrome (TS) patients is justified by the evaluation of the risk of development of germ cell tumors. In this study, we analyzed cryptic Y-chromosome derivatives by polymerase chain reaction (PCR) coupled with fluorescence in situ hybridization (FISH) using Y-specific sequences in patients with TS, and validated this methodology. Unrelated patients with TS (n=32) of Mexican mestizo ethnic origin were diagnosed using cytogenetic analysis. Clinical assessment, endocrine evaluation, karyotyping, FISH and PCR analysis of the Y-chromosomal loci were performed. We found that 9.4% (3 out of 32) patients with TS had Y-chromosome material. Two patients showed Y-chromosome by conventional cytogenetics. One patient had no Y-chromosome by initial karyotyping (45, X) but was positive by lymphocyte PCR DNA analysis of the Y-sequence-specific sex-determining region Y (SRY) gene. Our results suggest that the detection of the Y-chromosome material using sensitive methods, such as PCR coupled with FISH, should be carried out in all patients with TS and should not be limited to TS patients with cytogenetically identifiable Y-chromosome and/or virilization.

  7. Replication Banding Patterns in Human Chromosomes Detected Using 5-ethynyl-2'-deoxyuridine Incorporation

    PubMed Central

    Hoshi, Osamu; Ushiki, Tatsuo

    2011-01-01

    A novel technique using the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) into replicating DNA is described for the analysis of replicating banding patterns of human metaphase chromosomes. Human lymphocytes were synchronized with excess thymidine and treated with EdU during the late S phase of the cell cycle. The incorporated EdU was then detected in metaphase chromosomes using Alexa Fluor® 488 azides, through the 1,3-dipolar cycloaddition reaction of organic azides with the terminal acetylene group of EdU. Chromosomes with incorporated EdU showed a banding pattern similar to G-banding of normal human chromosomes. Imaging by atomic force microscopy (AFM) in liquid conditions showed that the structure of the chromosomes was well preserved even after EdU treatment. Comparison between fluorescence microscopy and AFM images of the same chromosome 1 indicated the presence of ridges and grooves in the chromatid arm, features that have been previously reported in relation to G-banding. These results suggest an intimate relationship between EdU-induced replication bands and G- or R-bands in human chromosomes. This technique is thus useful for analyzing the structure of chromosomes in relation to their banding patterns following DNA replication in the S phase. PMID:22096263

  8. Replication Banding Patterns in Human Chromosomes Detected Using 5-ethynyl-2'-deoxyuridine Incorporation.

    PubMed

    Hoshi, Osamu; Ushiki, Tatsuo

    2011-10-26

    A novel technique using the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) into replicating DNA is described for the analysis of replicating banding patterns of human metaphase chromosomes. Human lymphocytes were synchronized with excess thymidine and treated with EdU during the late S phase of the cell cycle. The incorporated EdU was then detected in metaphase chromosomes using Alexa Fluor® 488 azides, through the 1,3-dipolar cycloaddition reaction of organic azides with the terminal acetylene group of EdU. Chromosomes with incorporated EdU showed a banding pattern similar to G-banding of normal human chromosomes. Imaging by atomic force microscopy (AFM) in liquid conditions showed that the structure of the chromosomes was well preserved even after EdU treatment. Comparison between fluorescence microscopy and AFM images of the same chromosome 1 indicated the presence of ridges and grooves in the chromatid arm, features that have been previously reported in relation to G-banding. These results suggest an intimate relationship between EdU-induced replication bands and G- or R-bands in human chromosomes. This technique is thus useful for analyzing the structure of chromosomes in relation to their banding patterns following DNA replication in the S phase.

  9. A system for the detection of chromosomal rearrangements using Sordaria macrospora.

    PubMed

    Arnaise, S; Leblon, G; Lares, L

    1984-01-01

    A system is described for the detection and diagnosis of induced chromosomal rearrangement using Sordaria macrospora. The system uses the property of the rearrangement to produce defective white ascospores as meiotic progeny from heterozygous crosses. Two reconstruction experiments have shown that this system is able to give reliable quantitative measures of rearrangement frequencies. Evidence for a photoreactivation process was obtained, suggesting that pyrimidine dimers may well be an important lesion in UV-induced chromosomal rearrangement. No evidence of induction of chromosomal rearrangement was obtained in experiments with the powerful chemical mutagen N-methyl-N'-nitro-N-nitrosoguanidine.

  10. Statistical methods for detecting genomic alterations through array-based comparative genomic hybridization (CGH).

    PubMed

    Wang, Yuedong; Guo, Sun-Wei

    2004-01-01

    Array-based comparative genomic hybridization (ABCGH) is an emerging high-resolution and high-throughput molecular genetic technique that allows genome-wide screening for chromosome alterations associated with tumorigenesis. Like the cDNA microarrays, ABCGH uses two differentially labeled test and reference DNAs which are cohybridized to cloned genomic fragments immobilized on glass slides. The hybridized DNAs are then detected in two different fluorochromes, and the significant deviation from unity in the ratios of the digitized intensity values is indicative of copy-number differences between the test and reference genomes. Proper statistical analyses need to account for many sources of variation besides genuine differences between the two genomes. In particular, spatial correlations, the variable nature of the ratio variance and non-Normal distribution call for careful statistical modeling. We propose two new statistics, the standard t-statistic and its modification with variances smoothed along the genome, and two tests for each statistic, the standard t-test and a test based on the hybrid adaptive spline (HAS). Simulations indicate that the smoothed t-statistic always improves the performance over the standard t-statistic. The t-tests are more powerful in detecting isolated alterations while those based on HAS are more powerful in detecting a cluster of alterations. We apply the proposed methods to the identification of genomic alterations in endometrium in women with endometriosis.

  11. Detection of chromosomal anomalies in endometrial atypical hyperplasia and carcinoma by using fluorescence in situ hybridization.

    PubMed

    Qian, Junqi; Weber, Deena; Cochran, Richard; Hossain, Deloar; Bostwick, David G

    2010-04-25

    Endometrial cancer is the most common pelvic gynecological malignancy. The diagnosis of well-differentiated endometrial adenocarcinoma, atypical hyperplasia, and hyperplasia is often challenging. The authors sought to investigate the utility of chromosomal anomalies for the detection of endometrial hyperplasia and carcinoma using multitarget fluorescence in situ hybridization (FISH). Samples were collected by endometrial Tao brush and processed by liquid-based cytological preparation protocol from consecutive cases to include 50 benign, 50 hyperplasia without atypia, 47 atypical hyperplasia, and 53 endometrial cancers. Each was hybridized using fluorescence-labeled DNA probes to chromosomes 1, 8, and 10. The FISH signals were enumerated in 100 cells per case, and the chromosomal anomalies were correlated with pathologic findings, including histologic diagnoses on matched endometrial tissue samples. Numeric chromosomal anomalies were found in 0% (0 of 50) of benign, 20% (10 of 50) of hyperplasia, 74% (35 of 47) of atypical hyperplasia, and 87% (46 of 53) of carcinoma specimens. The mean percentage of cells with chromosomal changes was 55% in cancer specimens, which was significantly higher than that in hyperplasia without atypia (13%, P < .0001) and atypical hyperplasia (32%, P = .003). The most frequent chromosomal anomaly was gain of chromosome 1. FISH anomalies had an overall sensitivity of 81% and specificity of 90% for the detection of atypical hyperplasia and/or endometrial carcinoma. There was no association with grade of endometrial carcinoma. Multitarget FISH appears to be useful for the differential diagnosis of hyperplasia, atypical hyperplasia, and endometrial adenocarcinoma, with a high level of sensitivity and specificity. It is also a potential tool for the early detection of neoplastic cells in endometrial cytology specimens. Endometrial hyperplasia with FISH-detected chromosomal anomalies may represent a clinically significant subset of cases that

  12. A novel method for sex determination by detecting the number of X chromosomes.

    PubMed

    Nakanishi, Hiroaki; Shojo, Hideki; Ohmori, Takeshi; Hara, Masaaki; Takada, Aya; Adachi, Noboru; Saito, Kazuyuki

    2015-01-01

    A novel method for sex determination, based on the detection of the number of X chromosomes, was established. Current methods, based on the detection of the Y chromosome, can directly identify an unknown sample as male, but female gender is determined indirectly, by not detecting the Y chromosome. Thus, a direct determination of female gender is important because the quality (e.g., fragmentation and amelogenin-Y null allele) of the Y chromosome DNA may lead to a false result. Thus, we developed a novel sex determination method by analyzing the number of X chromosomes using a copy number variation (CNV) detection technique (the comparative Ct method). In this study, we designed a primer set using the amelogenin-X gene without the CNV region as the target to determine the X chromosome copy number, to exclude the influence of the CNV region from the comparative Ct value. The number of X chromosomes was determined statistically using the CopyCaller software with real-time PCR. All DNA samples from participants (20 males, 20 females) were evaluated correctly using this method with 1-ng template DNA. A minimum of 0.2-ng template DNA was found to be necessary for accurate sex determination with this method. When using ultraviolet-irradiated template DNA, as mock forensic samples, the sex of the samples could not be determined by short tandem repeat (STR) analysis but was correctly determined using our method. Thus, we successfully developed a method of sex determination based on the number of X chromosomes. Our novel method will be useful in forensic practice for sex determination.

  13. Chromosome-Specific Staining To Detect Genetic Rearrangements Associated With Chromosome 3 And/Or Chromosone 17

    DOEpatents

    Gray; Joe W.; Pinkel; Daniel; Kallioniemi; Olli-Pekka; Kallioniemi; Anne; Sakamoto; Masaru

    2002-02-05

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  14. Alterations of chromosome 11q13 in cervical carcinoma cell lines

    SciTech Connect

    Popescu, N.C.; Zimonjic, D.B.

    1996-02-01

    In cervical cancer, evidence for the existence of a tumor-suppressor gene on chromosome 11 has been generated from studies with somatic cell hybrids, chromosome microcell transfer, or deletion analysis of DNA markers. As suggested by somatic cell hybrids analysis, chromosome 11 harbors at least three distinctive tumor-suppressor genes, two on the short arm and one on the long arm. Loss of heterozygosity (LOH) analysis using 16 markers, 10 of which were microsatellite-based, placed the region of a putative tumor-suppressor gene to 11q22-24. Recently, 11q13 was assigned as another possible site on the basis of molecular rearrangements, deletions, and translocations, nonrandomly involving this region in four of eight cervical carcinoma cell lines. Abnormal chromosomes 11 were found in HeLa, SiHa, and Caski lines and in C33A, a human papilloma virus-negative cell line. 18 refs.

  15. Altered cohesin gene dosage affects Mammalian meiotic chromosome structure and behavior.

    PubMed

    Murdoch, Brenda; Owen, Nichole; Stevense, Michelle; Smith, Helen; Nagaoka, So; Hassold, Terry; McKay, Michael; Xu, Huiling; Fu, Jun; Revenkova, Ekaterina; Jessberger, Rolf; Hunt, Patricia

    2013-01-01

    Based on studies in mice and humans, cohesin loss from chromosomes during the period of protracted meiotic arrest appears to play a major role in chromosome segregation errors during female meiosis. In mice, mutations in meiosis-specific cohesin genes cause meiotic disturbances and infertility. However, the more clinically relevant situation, heterozygosity for mutations in these genes, has not been evaluated. We report here evidence from the mouse that partial loss of gene function for either Smc1b or Rec8 causes perturbations in the formation of the synaptonemal complex (SC) and affects both synapsis and recombination between homologs during meiotic prophase. Importantly, these defects increase the frequency of chromosomally abnormal eggs in the adult female. These findings have important implications for humans: they suggest that women who carry mutations or variants that affect cohesin function have an elevated risk of aneuploid pregnancies and may even be at increased risk of transmitting structural chromosome abnormalities.

  16. Association testing to detect gene-gene interactions on sex chromosomes in trio data.

    PubMed

    Lee, Yeonok; Ghosh, Debashis; Zhang, Yu

    2013-01-01

    Autism Spectrum Disorder (ASD) occurs more often among males than females in a 4:1 ratio. Among theories used to explain the causes of ASD, the X chromosome and the Y chromosome theories attribute ASD to the X-linked mutation and the male-limited gene expressions on the Y chromosome, respectively. Despite the rationale of the theory, studies have failed to attribute the sex-biased ratio to the significant linkage or association on the regions of interest on X chromosome. We further study the gender biased ratio by examining the possible interaction effects between two genes in the sex chromosomes. We propose a logistic regression model with mixed effects to detect gene-gene interactions on sex chromosomes. We investigated the power and type I error rates of the approach for a range of minor allele frequencies and varying linkage disequilibrium between markers and QTLs. We also evaluated the robustness of the model to population stratification. We applied the model to a trio-family data set with an ASD affected male child to study gene-gene interactions on sex chromosomes.

  17. Detection of chromosomal abnormalities and the 22q11 microdeletion in fetuses with congenital heart defects.

    PubMed

    Lv, Wei; Wang, Shuyu

    2014-11-01

    Chromosomal abnormalities and the 22q11 microdeletion are implicated in congenital heart defects (CHDs). This study was designed to detect these abnormalities in fetuses and determine the effect of genetic factors on CHD etiology. Between January 2010 and December 2011, 113 fetuses with CHD treated at the Beijing Obstetrics and Gynecology Hospital were investigated, using chromosome karyotyping of either amniotic fluid cell or umbilical cord blood cell samples. Fetuses with a normal result were then investigated for the 22q11 microdeletion by fluorescence in situ hybridization. Of the 113 patients, 12 (10.6%) exhibited chromosomal abnormalities, while 6 (5.3%) of the remaining 101 cases presented with a 22q11 microdeletion. The incidence of chromosomal abnormalities was significantly higher in the group of fetuses presenting with extracardiac malformations in addition to CHD (P<0.001), although the detection of the 22q11 microdeletion was not significantly different between the two groups (P=0.583). In addition, all fetuses with the 22q11 microdeletion occurred de novo. In conclusion, genetic factors are important in the etiology of CHD. Where fetuses present with cardiac defects, additional chromosomal analysis is required to detect extracardiac abnormalities. Fetuses with heart defects should also be considered for 22q11 microdeletion detection to evaluate fetal prognosis, particularly prior to surgery.

  18. Genomic Imbalances in Neonates With Birth Defects: High Detection Rates by Using Chromosomal Microarray Analysis

    PubMed Central

    Lu, Xin-Yan; Phung, Mai T.; Shaw, Chad A.; Pham, Kim; Neil, Sarah E.; Patel, Ankita; Sahoo, Trilochan; Bacino, Carlos A.; Stankiewicz, Pawel; Lee Kang, Sung-Hae; Lalani, Seema; Chinault, A. Craig; Lupski, James R.; Cheung, Sau W.; Beaudet, Arthur L.

    2009-01-01

    OBJECTIVES Our aim was to determine the frequency of genomic imbalances in neonates with birth defects by using targeted array-based comparative genomic hybridization, also known as chromosomal microarray analysis. METHODS Between March 2006 and September 2007, 638 neonates with various birth defects were referred for chromosomal microarray analysis. Three consecutive chromosomal microarray analysis versions were used: bacterial artificial chromosome-based versions V5 and V6 and bacterial artificial chromosome emulated oligonucleotide-based version V6 Oligo. Each version had targeted but increasingly extensive genomic coverage and interrogated >150 disease loci with enhanced coverage in genomic rearrangement-prone pericentromeric and subtelomeric regions. RESULTS Overall, 109 (17.1%) patients were identified with clinically significant abnormalities with detection rates of 13.7%, 16.6%, and 19.9% on V5, V6, and V6 Oligo, respectively. The majority of these abnormalities would not be defined by using karyotype analysis. The clinically significant detection rates by use of chromosomal microarray analysis for various clinical indications were 66.7% for “possible chromosomal abnormality” ± “others” (other clinical indications), 33.3% for ambiguous genitalia ± others, 27.1% for dysmorphic features + multiple congenital anomalies ± others, 24.6% for dysmorphic features ± others, 21.8% for congenital heart disease ± others, 17.9% for multiple congenital anomalies ± others, and 9.5% for the patients referred for others that were different from the groups defined. In all, 16 (2.5%) patients had chromosomal aneuploidies, and 81 (12.7%) patients had segmental aneusomies including common microdeletion or microduplication syndromes and other genomic disorders. Chromosomal mosaicism was found in 12 (1.9%) neonates. CONCLUSIONS Chromosomal microarray analysis is a valuable clinical diagnostic tool that allows precise and rapid identification of genomic imbalances

  19. SKY detection of chromosome rearrangements in two cases of tMDS with a complex karyotype.

    PubMed

    Cohen, Ninette; Trakhtenbrot, Luba; Yukla, Mona; Manor, Yosef; Gaber, Elena; Yosef, Gabi; Amariglio, Ninette; Rechavi, Gideon; Amiel, Aliza

    2002-10-15

    In this study, we used spectral karyotyping (SKY) and fluorescence in situ hybridization (FISH) as complementary techniques for the analysis of two therapy-related secondary myelodysplastic syndrome (t-MDS) cases with complex karyotypes, previously analyzed by G-banding. Different types of SKY's cytogenetic contributions include confirmation of G-banding results, identification of partially characterized rearrangements, identification of marker chromosomes unidentified by G-banding, and detection of cryptic reciprocal translocations. In particular, the ability of SKY to clarify a number of markers led to the comprehension of clonal evolution. The common aberration found in these two t-MDS cases was the fragility of chromosome 5 and monosomy of chromosome 18. We clearly present that the use of SKY combined with conventional G-banding analysis and FISH has assisted in the identification of important chromosomal events that may play a key role in the development of t-MDS.

  20. Endometriosis-induced alterations in mouse metaphase II oocyte microtubules and chromosomal alignment: a possible cause of infertility.

    PubMed

    Mansour, Gihan; Sharma, Rakesh K; Agarwal, Ashok; Falcone, Tommaso

    2010-10-01

    To examine the effect of peritoneal fluid (PF) of patients with endometriosis on the cytoskeleton of metaphase II oocytes and correlate the results with the stage of endometriosis and the duration of infertility. Prospective-controlled study. Center for reproductive medicine at a tertiary-care hospital. Women with endometriosis (n=23) and tubal ligation/reversal (n=15). Peritoneal fluid obtained from 38 women (23 with endometriosis and 15 tubal ligation/reversal) after laparoscopy. Four hundred metaphase II oocytes were used: 165 frozen metaphase II oocytes were incubated in the PF of patients with endometriosis, 135 oocytes incubated in the PF of nonendometriosis patients (control subjects) and 100 oocytes incubated in human tubal fluid (HTF) media. Spindle abnormalities (microtubule and chromosomal) were evaluated by confocal imaging. In the endometriosis group, the cytoskeleton had a higher frequency of abnormal meiotic spindle and chromosomal misalignment (score ≥3), indicating severe damage compared with the control groups. The proportions of abnormalities in microtubule and chromosome alterations in endometriosis (67.9% and 63.6%, respectively) were significantly higher than for oocytes incubated with PF of the nonendometriosis group (24.4% and 14.8%) as well as the HTF group (13% and 13%). Oocyte cytoskeleton damage positively correlated with the duration of infertility and the stage of endometriosis. Alteration of oocyte cytoskeleton might be one of the causes of poor oocyte quality in patients with endometriosis. Copyright © 2010. Published by Elsevier Inc.

  1. Deletion of DXZ4 on the human inactive X chromosome alters higher-order genome architecture

    PubMed Central

    Darrow, Emily M.; Huntley, Miriam H.; Dudchenko, Olga; Stamenova, Elena K.; Durand, Neva C.; Sun, Zhuo; Huang, Su-Chen; Sanborn, Adrian L.; Machol, Ido; Shamim, Muhammad; Seberg, Andrew P.; Lander, Eric S.; Chadwick, Brian P.; Aiden, Erez Lieberman

    2016-01-01

    During interphase, the inactive X chromosome (Xi) is largely transcriptionally silent and adopts an unusual 3D configuration known as the “Barr body.” Despite the importance of X chromosome inactivation, little is known about this 3D conformation. We recently showed that in humans the Xi chromosome exhibits three structural features, two of which are not shared by other chromosomes. First, like the chromosomes of many species, Xi forms compartments. Second, Xi is partitioned into two huge intervals, called “superdomains,” such that pairs of loci in the same superdomain tend to colocalize. The boundary between the superdomains lies near DXZ4, a macrosatellite repeat whose Xi allele extensively binds the protein CCCTC-binding factor. Third, Xi exhibits extremely large loops, up to 77 megabases long, called “superloops.” DXZ4 lies at the anchor of several superloops. Here, we combine 3D mapping, microscopy, and genome editing to study the structure of Xi, focusing on the role of DXZ4. We show that superloops and superdomains are conserved across eutherian mammals. By analyzing ligation events involving three or more loci, we demonstrate that DXZ4 and other superloop anchors tend to colocate simultaneously. Finally, we show that deleting DXZ4 on Xi leads to the disappearance of superdomains and superloops, changes in compartmentalization patterns, and changes in the distribution of chromatin marks. Thus, DXZ4 is essential for proper Xi packaging. PMID:27432957

  2. Parental decisions of prenatally detected sex chromosome abnormality.

    PubMed Central

    Kim, Yon-Ju; Park, So-Yeon; Han, Jung-Heol; Kim, Moon-Young; Yang, Jae-Hyug; Choi, Kyu-Hong; Kim, Young-Mi; Kim, Jin-Mee; Ryu, Hyun-Mee

    2002-01-01

    Because of the widespread use of amniocentesis, the prenatal recognition of sex chromosome abnormality (SCA) has become increasingly common. Recent literature provided an insight into the understanding of the natural history and prognosis for individuals with SCA. Our study was designed to review the parental decision on pregnancy with SCA. Over the last 10 yr, we diagnosed 38 cases (0.50%) with SCA out of 7,498 prenatal cases. We reviewed the records and the results of the pregnancies. We included the cases (n=25) of apparently normal anatomic fetus to analyze the factors influencing parental decision. We excluded 13 cases with obvious anomaly or presumably bad outcome. Fifteen (60%) couples continued their pregnancies and ten (40%) terminated theirs. Nine couples (64%) out of fourteen mosaicism cases continued their pregnancies. All five pregnancies assisted by reproductive technique continued their pregnancies. More pregnancies were continued when counseling was done by an MD geneticist rather than by an obstetrician. A significant trend was observed with a higher rate of pregnancy continuation in recent years. The genetic counseling is important to give appropriate information to the parents. Establishing guidelines and protocols will help both obstetricians and parents to make a decision. PMID:11850589

  3. Marker chromosomes.

    PubMed

    Rao, Kiran Prabhaker; Belogolovkin, Victoria

    2013-04-01

    Marker chromosomes are a morphologically heterogeneous group of structurally abnormal chromosomes that pose a significant challenge in prenatal diagnosis. Phenotypes associated with marker chromosomes are highly variable and range from normal to severely abnormal. Clinical outcomes are very difficult to predict when marker chromosomes are detected prenatally. In this review, we outline the classification, etiology, cytogenetic characterization, and clinical consequences of marker chromosomes, as well as practical approaches to prenatal diagnosis and genetic counseling.

  4. Y chromosome microdeletions and alterations of spermatogenesis, patient approach and genetic counseling.

    PubMed

    Rives, Nathalie

    2014-05-01

    Infertility affects 15% of couples at reproductive age and human male infertility appears frequently idiopathic. The main genetic causes of spermatogenesis defect responsible for non-obstructive azoospermia and severe oligozoospermia are constitutional chromosomal abnormalities and microdeletions in the azoospermia factor region of the Y chromosome. The improvement of the Yq microdeletion screening method gave new insights in the mechanism responsible for the genesis of Yq microdeletions and for the consequences of the management of male infertility and genetic counselling in case of assisted reproductive technology. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  5. Influence of incorporated bromodeoxyuridine on the induction of chromosomal alterations by ionizing radiation and long-wave UV in CHO cells.

    PubMed

    Zwanenburg, T S; van Zeeland, A A; Natarajan, A T

    1985-01-01

    Incorporation of BrdUrd into nuclear DNA sensitizes CHO cells (1) to the induction of chromosomal aberrations by X-rays and 0.5 MeV neutrons and (2) to induction of chromosomal aberrations and SCEs by lw-UV. We have attempted to establish a correlation between induced chromosomal alterations and induced single- or double-strand breaks in DNA. The data show that while DSBs correlate very well with X-ray-induced aberrations, no clear correlation could be established between lw-UV induced SSBs (including alkali-labile sites) and chromosomal alterations. In addition the effect of 3-aminobenzamide (3AB) on the induction of chromosomal aberrations and SCEs induced by lw-UV has been determined. It is shown that 3AB is without any effect when lw-UV-irradiated cells are posttreated with this inhibitor. The significance of these results is discussed.

  6. No excess gene movement is detected off the avian or lepidopteran Z chromosome.

    PubMed

    Toups, Melissa A; Pease, James B; Hahn, Matthew W

    2011-01-01

    Most of our knowledge of sex-chromosome evolution comes from male heterogametic (XX/XY) taxa. With the genome sequencing of multiple female heterogametic (ZZ/ZW) taxa, we can now ask whether there are patterns of evolution common to both sex chromosome systems. In all XX/XY systems examined to date, there is an excess of testis-biased retrogenes moving from the X chromosome to the autosomes, which is hypothesized to result from either sexually antagonistic selection or escape from meiotic sex chromosome inactivation (MSCI). We examined RNA-mediated (retrotransposed) and DNA-mediated gene movement in two independently evolved ZZ/ZW systems, birds (chicken and zebra finch) and lepidopterans (silkworm). Even with sexually antagonistic selection likely operating in both taxa and MSCI having been identified in the chicken, we find no evidence for an excess of genes moving from the Z chromosome to the autosomes in either lineage. We detected no excess for either RNA- or DNA-mediated duplicates, across a range of approaches and methods. We offer some potential explanations for this difference between XX/XY and ZZ/ZW sex chromosome systems, but further work is needed to distinguish among these hypotheses. Regardless of the root causes, we have identified an additional, potentially inherent, difference between XX/XY and ZZ/ZW systems.

  7. Detection of Y chromosome microdeletions and mitochondrial DNA mutations in male infertility patients.

    PubMed

    Güney, A I; Javadova, D; Kırac, D; Ulucan, K; Koc, G; Ergec, D; Tavukcu, H; Tarcan, T

    2012-04-27

    Infertility affects about 10-15% of all couples attempting pregnancy with infertility attributed to the male partner in approximately half of the cases. Proposed causes of male infertility include sperm motility disturbances, Y chromosome microdeletions, chromosomal abnormalities, single gene mutations, and sperm mitochondrial DNA (mtDNA) rearrangements. To investigate the etiology of decreased sperm fertility and motility of sperm and to develop an appropriate therapeutic strategy, the molecular basis of these defects must be elucidated. In this study, we aimed to reveal the relationships between the genetic factors including sperm mtDNA mutations, Y chromosome microdeletions, and sperm parameters that can be regarded as candidate factors for male infertility. Thirty men with a history of infertility and 30 fertile men were recruited to the study. Y chromosome microdeletions were analyzed by multiplex PCR. Mitochondrial genes ATPase6, Cytb, and ND1, were amplified by PCR and then analyzed by direct sequencing. No Y chromosome microdeletions were detected in either group. However, a total of 38 different nucleotide substitutions were identified in the examined mitochondrial genes in both groups, all of which are statistically non-significant. Fifteen substitutions caused an amino acid change and 12 were considered novel mutations. As a conclusion, mtDNA mutations and Y chromosome microdeletions in male infertility should be examined in larger numbers in order to clarify the effect of genetic factors.

  8. Alterations in glycosylation as biomarkers for cancer detection.

    PubMed

    Reis, Celso A; Osorio, Hugo; Silva, Luisa; Gomes, Catarina; David, Leonor

    2010-04-01

    Glycoconjugates constitute a major class of biomolecules which include glycoproteins, glycosphingolipids and proteoglycans. Glycans are involved in several physiological and pathological conditions, such as host-pathogen interactions, cell differentiation, migration, tumour invasion and metastisation, cell trafficking and signalling. Cancer is associated with glycosylation alterations in glycoproteins and glycolipids. This review describes various aspects of protein glycosylation with the focus on alterations associated with human cancer. The application of these glycosylation modifications as biomarkers for cancer detection in tumour tissues and serological assays is summarised.

  9. Chromosome Specific Substitution Lines of Aegilops geniculata Alter Parameters of Bread Making Quality of Wheat

    PubMed Central

    Tsujimoto, Hisashi; Gupta, Raj Kumar; Kumar, Aman; Kaur, Navneet; Kumar, Rohit; Chunduri, Venkatesh; Sharma, Nand Kishor; Chawla, Meenakshi; Sharma, Saloni; Mundey, Jaspreet Kaur

    2016-01-01

    Wheat cultivars with wide introgression have strongly impacted global wheat production. Aegilops geniculata (MgUg) is an important wild relative with several useful traits that can be exploited for wheat improvement. Screening of Ae. geniculata addition lines indicated a negative effect of 1Ug and the positive effect of 1Mg chromosome on wheat dough strength. Negative effect of 1Ug is probably associated with variation in number and position of the tripeptide repeat motif in the high molecular weight glutenin (HMW-G) gene. To utilize the positive potential of 1Mg chromosome, three disomic substitution lines (DSLs) 1Mg(1A), 1Mg(1B) and 1Mg(1D) were created. These lines were characterized for morphological, cytogenetic properties and biochemical signatures using FISH, 1D-, 2D-PAGE and RP-HPLC. Contribution of wheat 1A, 1B and 1D chromosomes towards dough mixing and baking parameters, chapatti quality, Fe/Zn content and glume color were identified. Observed order of variation in the dough mixing and baking parameters {1Mg(1D) ≤wheat ≤1Mg(1B) ≤1Mg(1A)} indicated that chromosome specific introgression is desirable for best utilization of wild species’ potential. PMID:27755540

  10. Ionizing irradiation-induced radical stress stalls live meiotic chromosome movements by altering the actin cytoskeleton

    PubMed Central

    Illner, Doris; Scherthan, Harry

    2013-01-01

    Meiosis generates haploid cells or spores for sexual reproduction. As a prelude to haploidization, homologous chromosomes pair and recombine to undergo segregation during the first meiotic division. During the entire meiotic prophase of the yeast Saccharomyces cerevisiae, chromosomes perform rapid movements that are suspected to contribute to the regulation of recombination. Here, we investigated the impact of ionizing radiation (IR) on movements of GFP–tagged bivalents in live pachytene cells. We find that exposure of sporulating cultures with >40 Gy (4-krad) X-rays stalls pachytene chromosome movements. This identifies a previously undescribed acute radiation response in yeast meiosis, which contrasts with its reported radioresistance of up to 1,000 Gy in survival assays. A modified 3′-end labeling assay disclosed IR-induced dsDNA breaks (DSBs) in pachytene cells at a linear dose relationship of one IR-induced DSB per cell per 5 Gy. Dihydroethidium staining revealed formation of reactive oxygen species (ROS) in irradiated cells. Immobility of fuzzy-appearing irradiated bivalents was rescued by addition of radical scavengers. Hydrogen peroxide-induced ROS did reduce bivalent mobility similar to 40 Gy X IR, while they failed to induce DSBs. IR- and H2O2-induced ROS were found to decompose actin cables that are driving meiotic chromosome mobility, an effect that could be rescued by antioxidant treatment. Hence, it appears that the meiotic actin cytoskeleton is a radical-sensitive system that inhibits bivalent movements in response to IR- and oxidant-induced ROS. This may be important to prevent motility-driven unfavorable chromosome interactions when meiotic recombination has to proceed in genotoxic environments. PMID:24046368

  11. SNP-based non-invasive prenatal testing detects sex chromosome aneuploidies with high accuracy

    PubMed Central

    Samango-Sprouse, Carole; Banjevic, Milena; Ryan, Allison; Sigurjonsson, Styrmir; Zimmermann, Bernhard; Hill, Matthew; Hall, Megan P.; Westemeyer, Margaret; Saucier, Jennifer; Demko, Zachary; Rabinowitz, Matthew

    2013-01-01

    Objective To develop a single nucleotide polymorphism- and informatics-based non-invasive prenatal test that detects sex chromosome aneuploidies early in pregnancy. Methods Fifteen aneuploid samples, including thirteen 45,X, two 47,XXY, and one 47,XYY, along with 185 euploid controls, were analyzed. Cell-free DNA was isolated from maternal plasma, amplified in a single multiplex PCR assay that targeted 19,488 polymorphic loci covering chromosomes 13, 18, 21, X, and Y, and sequenced. Sequencing results were analyzed using a Bayesian-based maximum likelihood statistical method to determine copy number of interrogated chromosomes, calculating sample-specific accuracies. Results Of the samples that passed a stringent quality control metric (93%), the algorithm correctly identified copy number at all five chromosomes in all 187 samples, for 934/935 correct calls as early as 9.4 weeks of gestation. We detected 45,X with 91.7% sensitivity (CI: 61.5-99.8%) and 100% specificity (CI: 97.9-100%), and 47,XXY and 47,XYY. The average calculated accuracy was 99.78%. Conclusion This method non-invasively detected 45,X, 47,XXY, and 47,XYY fetuses from cfDNA isolated from maternal plasma with high calculated accuracies, and thus offers a non-invasive method with the potential to function as a routine screen allowing for early prenatal detection of rarely diagnosed yet commonly occurring sex aneuploidies. PMID:23712453

  12. Genetic effects of individual chromosomes in cotton cultivars detected by using chromosome substitution lines as genetic probes.

    PubMed

    Wu, Jixiang; Jenkins, Johnie N; McCarty, Jack C; Saha, Sukumar

    2010-12-01

    Determination of chromosomes or chromosome arms with desirable genes in different inbred lines and/or crosses should provide useful genetic information for crop improvement. In this study, we applied a modified additive-dominance model to analyze a data set of 13 cotton chromosome substitution lines and their recurrent parent TM-1, five commercial cultivars, and their 70 F(2) hybrids. The chromosome additive and dominance variance components for eight agronomic and fiber traits were determined. On average, each chromosome or chromosome arm was associated with 6.5 traits in terms of additive and/or dominance effects. The chromosomes or chromosome arms, which contributed significant additive variances for the traits investigated, included 2, 16, 18, 25, 5sh (short arm), 14sh, 15sh, 22sh, and 22Lo (long arm). Chromosome additive effects were also predicted in this study. The results showed that CS-B 25 was favorably associated with several fiber traits, while FM966 was favorably associated with both yield and fiber traits with alleles on multiple chromosomes or chromosome arms. Thus, this study should provide valuable genetic information on pure line development for several improved traits such as yield and fiber quality.

  13. Array-based detection of genetic alterations associated with disease

    SciTech Connect

    Pinkel, Daniel; Albertson, Donna G.; Gray, Joe W.

    2007-09-11

    The present invention relates to DNA sequences from regions of copy number change on chromosome 20. The sequences can be used in hybridization methods for the identification of chromosomal abnormalities associated with various diseases.

  14. Traditional karyotyping vs copy number variation sequencing for detection of chromosomal abnormalities associated with spontaneous miscarriage.

    PubMed

    Liu, S; Song, L; Cram, D S; Xiong, L; Wang, K; Wu, R; Liu, J; Deng, K; Jia, B; Zhong, M; Yang, F

    2015-10-01

    To compare the performance of traditional G-banding karyotyping with that of copy number variation sequencing (CNV-Seq) for detection of chromosomal abnormalities associated with miscarriage. Products of conception (POC) were collected from spontaneous miscarriages. Chromosomal abnormalities were detected using high-resolution G-banding karyotyping and CNV sequencing. Quantitative fluorescent polymerase chain reaction analysis of maternal and POC DNA for short tandem repeat (STR) markers was used to both monitor maternal cell contamination and confirm the chromosomal status and sex of the miscarriage tissue. A total of 64 samples of POC, comprising 16 with an abnormal and 48 with a normal karyotype, were selected and coded for analysis by CNV-Seq. CNV-Seq results were concordant for 14 (87.5%) of the 16 gross chromosomal abnormalities identified by karyotyping, including 11 autosomal trisomies and three sex chromosomal aneuploidies (45,X). Of the two discordant results, a 69,XXX polyploidy was missed by CNV-Seq, although supporting STR marker analysis confirmed the triploidy. In contrast, CNV-Seq identified a sample with 45,X karyotype as a 45,X/46,XY mosaic. In the remaining 48 samples of POC with a normal karyotype, CNV-Seq detected a 2.58-Mb 22q deletion associated with DiGeorge syndrome and nine different smaller CNVs of no apparent clinical significance. CNV-Seq used in parallel with STR profiling is a reliable and accurate alternative to karyotyping for identifying chromosome copy number abnormalities associated with spontaneous miscarriage. Copyright © 2015 ISUOG. Published by John Wiley & Sons Ltd.

  15. CAFE: an R package for the detection of gross chromosomal abnormalities from gene expression microarray data.

    PubMed

    Bollen, Sander; Leddin, Mathias; Andrade-Navarro, Miguel A; Mah, Nancy

    2014-05-15

    The current methods available to detect chromosomal abnormalities from DNA microarray expression data are cumbersome and inflexible. CAFE has been developed to alleviate these issues. It is implemented as an R package that analyzes Affymetrix *.CEL files and comes with flexible plotting functions, easing visualization of chromosomal abnormalities. CAFE is available from https://bitbucket.org/cob87icW6z/cafe/ as both source and compiled packages for Linux and Windows. It is released under the GPL version 3 license. CAFE will also be freely available from Bioconductor. sander.h.bollen@gmail.com or nancy.mah@mdc-berlin.de Supplementary data are available at Bioinformatics online.

  16. Copy number alterations of chromosomal regions enclosing protein tyrosine phosphatase receptor-like genes in colorectal cancer.

    PubMed

    Laczmanska, Izabela; Karpinski, Pawel; Kozlowska, Joanna; Bebenek, Marek; Ramsey, David; Sedziak, Tomasz; Ziolkowski, Piotr; Sasiadek, Maria M

    2014-12-01

    Protein tyrosine phosphatases that act in different cellular pathways are described most commonly as tumor suppressors, but also as oncogenes. Their role has previously been described in colorectal cancer, as well as in gastric, breast, thyroid, prostate, ovarian, pancreatic, glioma, liver, leukemia and many other cancers. In a previous study, we have described protein tyrosine phosphatase receptor type T, M, Z1 and Q genes (PTPRT, PTPRM, PTPRZ1 and PTPRQ) hypermethylated in sporadic colorectal cancer. Thus, in this study, we examined the relation of unbalanced chromosomal alterations within regions covering these four protein tyrosine phosphatase genes with this cancer. One hundred and two cancer tissues were molecularly characterized, including analysis of the BRAF and K-ras mutations and methylator phenotype. The analysis of chromosomal aberrations was performed using Comparative Genomic Hybridization. We observed amplification of three regions containing genes coding for PTPs, such as PTPRZ1 (7q31.3, amplified in 23.5% of cases), PTPRQ (12q21.2, amplified in 5.9% of cases), PTPRT (20q12, amplified in 29.4% of cases), along with deletions in the region of PTPRM (18p11.2, deleted in 21.6% of cases). These data may suggest that in sporadic colorectal cancer PTPRZ1, PTPRT, PTPRQ probably act as oncogenes, while PTPRM acts as a tumor suppressor gene. Our study also revealed that gains on chromosome 20q12 and losses on chromosome 18p11.2 are connected with the absence of the BRAF mutation and the conventional adenocarcinoma pathway. Copyright © 2014 Elsevier GmbH. All rights reserved.

  17. Chromosome 7 abnormalities in prostate cancer detected by dual-color fluorescence in situ hybridization.

    PubMed

    Cui, J; Deubler, D A; Rohr, L R; Zhu, X L; Maxwell, T M; Changus, J E; Brothman, A R

    1998-11-01

    Aneusomy of chromosome 7 and loss at 7q (especially 7q31.1) have been reported in prostate cancer. To further investigate abnormalities of 7q and the relationship with whole chromosome 7 changes, we have conducted a dual-color fluorescence in situ hybridization (FISH) analysis on isolated nuclei from 28 primary prostate cancers. A pericentromeric probe for chromosome 7, five newly isolated sequence-specific bacterial artificial chromosome (BAC) probes from 7q31.1, and one BAC for the epidermal growth factor receptor (EGFR) gene at 7p12 were used in dual color hybridizations. Pericentromeric probes for chromosomes X and 4 were also used as controls. Sixteen (57.1%) of the 28 tumors showed clonal aberrations. Nine of them were trisomy 7 and four were hypertetrasomy for chromosome 7. Deletions at 7q31.1 were found in two of the high grade tumors. With the exception of these two cases, all other cases showed concordant results using all probes. These findings confirm previous studies that aneusomy of 7 is associated with prostate cancer progression, and there may be a tumor suppressor gene (TSG) at 7q31.1 which is associated with tumor progression. In addition, our study indicates: (1) the deletion pattern of individual nuclei infers that deletions at 7q31.1 precede reduplications of chromosome 7; and (2) the amplification of EGFR was not detected at the DNA level, suggesting that activation of this oncogene may play a minor role in prostate cancer.

  18. Nicotine-induced Disturbances of Meiotic Maturation in Cultured Mouse Oocytes: Alterations of Spindle Integrity and Chromosome Alignment.

    PubMed

    Zenzes, Maria Teresa; Bielecki, Ryszard

    2004-09-15

    We investigated whether nicotine exposure in vitro of mouse oocytes affects spindle and chromosome function during meiotic maturation (M-I and M-II). Oocytes in germinal vesicle (GV) stage were cultured in nicotine for 8 h or for 16 h, to assess effects in M-I and in metaphase II (M-II). The latter culture setting used the three protocols: 8 h nicotine then 8 h medium (8N + 8M); 16 h nicotine (16N); 8 h medium then 8 h nicotine (8M + 8N). Non-toxic concentrations of nicotine at 1.0, 2.5, 5.0 and 10.0 mmol/L were used. Spindle-chromosome configurations were analyzed with wide-field optical sectioning microscopy. In 8 h cultures, nicotine exposure resulted in dose-related increased proportions of M-I oocytes with defective spindle-chromosome configurations. A dose-related delayed entry into anaphase I was also detected. In 16 h cultures, nicotine exposure for the first 8 h (8N + 8M), or for 16 h (16N), resulted in dose- and time-related increased proportions of oocytes arrested in M-I (10 mmol/L; 8 h: 53.2%, controls 9.6%; 16 h: 87.6%, controls 8.5%). Defects in M-I spindles and chromosomes caused M-I arrest leading to dose-related decreased proportions of oocytes that reached metaphase-II (10 mmol/L 8 h: 46.8%, controls 90.4%;16 h: 12.4%, controls 91.5%). A delayed anaphase-I affected the normal timing of M-II, leading to abnormal oocytes with dispersed chromosomes, or with double spindles and no polar body. Nicotine exposure during the second 8 h (8M + 8N) resulted in dose-related, increased proportions of M-II oocytes with defective spindles and chromosomes (10 mmol/L: 42.9%, controls 2.0%). Nicotine has no adverse effects on GV break down, but induces spindle and chromosome defects compromising oocyte meiotic maturation and development.

  19. Method for detection and identification of multiple chromosomal integration sites in transgenic animals created with lentivirus.

    PubMed

    Bryda, Elizabeth C; Pearson, Michael; Agca, Yuksel; Bauer, Beth A

    2006-12-01

    Transgene delivery systems, particularly those involving retroviruses, often result in the integration of multiple copies of the transgene throughout the host genome. Since site-specific silencing of trangenes can occur; it becomes important to identify the number and chromosomal location of the multiple copies of the transgenes in order to correlate inheritance of the transgene at a particular chromosomal site with a specific and robust phenotype. Using a technique that combines restriction endonuclease digest and several rounds of PCR amplification followed by nucleotide sequencing, it is possible to identify multiple chromosomal integration sites in transgenic founder animals. By designing genotyping assays to detect each individual integration site in the offspring of these founders, the inheritance of transgenes integrated at specific chromosomal locations can be followed efficiently as the transgenes randomly segregate in subsequent generations. Phenotypic characteristics can then be correlated with inheritance of a transgene integrated at a particular chromosomal location to allow rational selection of breeding animals in order to establish the transgenic line.

  20. Experimental approaches for the detection of chromosomal malsegregation occurring in the germline of mammals

    SciTech Connect

    Russell, L.B.

    1985-01-01

    Existing and newly proposed methods to detect the induction of heritable aneuploidy are summarized, and their favorable and unfavorable features discussed. Among the tests involving direct chromosomal examination, that involving study of pronuclear chromosome at first cleavage is judged to be the most universally informative and reliable, provided tertiary trisomy can be ruled out. Measurement of post-midterm (fetal) death is proposed as a trisomy prescreen that can be readily combined with a dominant-lethal test. Among the genetic procedures for identifying the results of malsegregation events, direct detection of aneuploids has a number of advantages over complementation methods, in which only a fraction of the products of aneuploid gametes is detectable. Direct detection of aneuploids must, however, be restricted to sex-chromosomes, if postnatal animals are examined. A genetic marker system to detect autosomal trisomies in fetuses is proposed. An examination of experimental parameters that might maximize induction of malsegregation leads to the recommendation to include preleptotene among exposed germ-cell stages. 64 refs., 3 figs., 5 tabs.

  1. Manipulation Detection and Preference Alterations in a Choice Blindness Paradigm

    PubMed Central

    Taya, Fumihiko; Gupta, Swati; Farber, Ilya; Mullette-Gillman, O'Dhaniel A.

    2014-01-01

    Objectives It is commonly believed that individuals make choices based upon their preferences and have access to the reasons for their choices. Recent studies in several areas suggest that this is not always the case. In choice blindness paradigms, two-alternative forced-choice in which chosen-options are later replaced by the unselected option, individuals often fail to notice replacement of their chosen option, confabulate explanations for why they chose the unselected option, and even show increased preferences for the unselected-but-replaced options immediately after choice (seconds). Although choice blindness has been replicated across a variety of domains, there are numerous outstanding questions. Firstly, we sought to investigate how individual- or trial-factors modulated detection of the manipulations. Secondly, we examined the nature and temporal duration (minutes vs. days) of the preference alterations induced by these manipulations. Methods Participants performed a computerized choice blindness task, selecting the more attractive face between presented pairs of female faces, and providing a typewritten explanation for their choice on half of the trials. Chosen-face cue manipulations were produced on a subset of trials by presenting the unselected face during the choice explanation as if it had been selected. Following all choice trials, participants rated the attractiveness of each face individually, and rated the similarity of each face pair. After approximately two weeks, participants re-rated the attractiveness of each individual face online. Results Participants detected manipulations on only a small proportion of trials, with detections by fewer than half of participants. Detection rates increased with the number of prior detections, and detection rates subsequent to first detection were modulated by the choice certainty. We show clear short-term modulation of preferences in both manipulated and non-manipulated explanation trials compared to choice

  2. A simple cytogenetic method to detect chromosomally integrated human herpesvirus-6.

    PubMed

    Ohye, Tamae; Kawamura, Yoshiki; Inagaki, Hidehito; Yoshikawa, Akiko; Ihira, Masaru; Yoshikawa, Tetsushi; Kurahashi, Hiroki

    2016-02-01

    Some healthy individuals carry human herpesvirus-6 (HHV-6) within a host chromosome, which is called inherited chromosomally integrated human herpesvirus-6 (iciHHV-6). Because iciHHV-6 is generally considered a non-pathogenic condition, it is important to distinguish iciHHV-6 from HHV-6 reactivation in immunocompromised hosts because both conditions manifest high copy numbers of the HHV-6 in peripheral blood mononuclear cells. Although fluorescent in situ hybridization (FISH) is a reliable method for the diagnosis of iciHHV-6, HHV-6-specific FISH probes are not commercially available. In our present study, we established a simple PCR-based method for producing FISH probes that can detect the chromosomal integration site of iciHHV-6 at high sensitivity. Using these probes, we confirmed that HHV-6 signals were consistently located at the telomeric region in all of the 13 iciHHV-6 individuals examined. Interestingly, in all seven Japanese iciHHV-6A patients, signals were detected exclusively on chromosome 22q. This method provides a simple and fast approach for iciHHV-6 diagnosis in the clinical laboratory.

  3. Flow cytometry may allow microscope-independent detection of holocentric chromosomes in plants

    PubMed Central

    Zedek, František; Veselý, Pavel; Horová, Lucie; Bureš, Petr

    2016-01-01

    Two chromosomal structures, known as monocentric and holocentric chromosomes, have evolved in eukaryotes. Acentric fragments of monocentric chromosomes are unequally distributed to daughter cells and/or lost, while holocentric fragments are inherited normally. In monocentric species, unequal distribution should generate chimeras of cells with different nuclear DNA content. We investigated whether such differences in monocentric species are detectable by flow cytometry (FCM) as (i) a decreased nuclear DNA content and (ii) an increased coefficient of variance (CV) of the G1 peak after gamma radiation-induced fragmentation. We compared 13 monocentric and 9 holocentric plant species. Unexpectedly, monocentrics and holocentrics did not differ with respect to parameters (i) and (ii) in their response to gamma irradiation. However, we found that the proportion of G2 nuclei was highly elevated in monocentrics after irradiation, while holocentrics were negligibly affected. Therefore, we hypothesize that DNA-damaging agents induce cell cycle arrest leading to endopolyploidy only in monocentric and not (or to much lesser extent) in holocentric plants. While current microscope-dependent methods for holocentrism detection are unreliable for small and numerous chromosomes, which are common in holocentrics, FCM can use somatic nuclei. Thus, FCM may be a rapid and reliable method of high-throughput screening for holocentric candidates across plant phylogeny. PMID:27255216

  4. [FREQUENCIES OF FETAL CHROMOSOMAL ABERRATIONS DETECTED BY AMNIOCENTESIS: OUR 15-YEARS EXPERIENCE].

    PubMed

    Stoyanova, V; Ivanov, H; Linev, A; Vachev, T

    2015-01-01

    Amniocentesis is the most common and reliable prenatal diagnostic method for chromosomopathies. The purpose of the present study is to retrospectively evaluate our 15-year experience with prenatal cytogenetic diagnosis by amniocentesis, focusing on the indications and rates of chromosome abnormalities. The current study involve prenatal cytogenetic analysis from 564 amniocentesis performed at the Department of Medical Genetics, St. George University Hospital, Plovdiv between January 2000 and December 2014. Among clinical indications, abnormal maternal serum screening results (54.96%; 310/564) have been the most common indication for amniocentesis. Chromosomal abnormalities were detected in 5.5% (31/546) of cases. Structural rearrangements were the most common abnormality found (16/3 1;51,61%) with prevalence of balanced aberrations--11 cases. The highest detection rate of chromosome aberrations was in cases undergoing amniocentesis due to known family history of chromosomal abnormality (15.1%), followed by abnormal fetal ultrasound finding group (7.69%), increasing-risk maternal prenatal screening results (4.52%), and advanced maternal age (3.28%). This study provides important information for prenatal genetic counseling of families at risk with aim of prenatal care and prevention during pregnancies.

  5. Melatonin prevents hypochlorous acid-induced alterations in microtubule and chromosomal structure in metaphase-II mouse oocytes.

    PubMed

    Banerjee, Jashoman; Maitra, Dhiman; Diamond, Michael P; Abu-Soud, Husam M

    2012-09-01

    Hypochlorous acid (HOCl) is generated by myeloperoxidase, using chloride and hydrogen peroxide as substrates. Here we demonstrate that HOCl alters metaphase-II mouse oocyte microtubules and chromosomal (CH) alignment which can be prevented by melatonin. Metaphase-II mouse oocytes, obtained commercially, were grouped as: control, melatonin (150, 200nmol/mL), HOCl (10, 20, 50, and 100nmol/mL), and HOCl (50nmol/mL) pretreated with 150 and 200 nmol/mL of melatonin. Microtubule and CH alignment was studied utilizing an indirect immunofluorescence technique and scored by two observers. Pearson chi-square test and Fisher's exact test were used to compare outcomes between controls and treated groups and also among each group. Poor scores for the spindle and chromosomes increased significantly at 50nmol/mL of HOCl (P<0.001). Oocytes treated with melatonin only at 150 and 200 nmol/mL showed no changes; significant differences (P<0.001) were observed when oocytes exposed to 50nmol/mL of HOCl were compared to oocytes pretreated with 200 nmol/mL melatonin. Fifty percent of the oocytes demonstrated good scores, both in microtubule and CH alterations, when pretreated with melatonin at 150 nmol/mL compared to 0% in the HOCl-only group. HOCl alters the metaphase-II mouse oocyte spindle and CH alignment in a dose-dependant manner, which might be a potential cause of poor oocyte quality (e.g., in patients with endometriosis). Melatonin prevented the HOCl-mediated spindle and CH damage, and therefore, may be an attractive therapeutic option to prevent oocyte damage in endometriosis or inflammatory diseases where HOCl levels are known to be elevated. © 2012 John Wiley & Sons A/S.

  6. Multicolor FISHs for simultaneous detection of genes and DNA segments on human chromosomes.

    PubMed

    Shimizu, Nobuyoshi; Maekawa, Masahiko; Asai, Satoko; Shimizu, Yoshiko

    2015-12-01

    We have developed a convenient multicolor fluorescent in situ hybridization (FISH) (five-, four-, three-, and two-color FISHs) for detecting specific genes/DNA segments on the human chromosomes. As a foundation of multicolor FISH, we first isolated 80 bacterial artificial chromosome (BAC) probes that specifically detect the peri-centromeres (peri-CEN) and subtelomeres (subTEL) of 24 different human chromosomes (nos. 1~22, X, and Y) by screening our homemade BAC library (Keio BAC library) consisting of 200,000 clones. Five-color FISH was performed using human DNA segments specific for peri-CEN or subTEL, which were labeled with five different fluorescent dyes [7-diethylaminocoumarin (DEAC): blue, fluorescein isothiocyanate (FITC): green, rhodamine 6G (R6G): yellow, TexRed: red, and cyanine5 (Cy5): purple]. To observe FISH signals under a fluorescence microscope, five optic filters were carefully chosen to avoid overlapping fluorescence emission. Five-color FISH and four-color FISH enabled us to accurately examine the numerical anomaly of human chromosomes. Three-color FISH using two specific BAC clones, that distinguish 5' half of oncogene epidermal growth factor receptor (EGFR) from its 3' half, revealed the amplification and truncation of EGFR in EGFR-overproducing cancer cells. Moreover, two-color FISH readily detected a fusion gene in leukemia cells such as breakpoint cluster region (BCR)/Abelson murine leukemia viral oncogene homologue (ABL) on the Philadelphia (Ph') chromosome with interchromosomal translocation. Some other successful cases such as trisomy 21 of Down syndrome are presented. Potential applications of multicolor FISH will be discussed.

  7. Ensemble spontaneous activity alterations detected by CISA approach.

    PubMed

    Boudaoud, Sofiane; Rix, Hervé; Meste, Olivier; Cazals, Yves

    2007-01-01

    In this paper, we propose a method for detecting alterations in the Ensemble Spontaneous Activity (ESA), a random signal representing the composite spontaneous contribution of the auditory nerve recorded on the round window. The proposed method is based on shape analysis of the ESA amplitude histogram. For this task, we use a recent approach, the Corrected Integral Shape Averaging (CISA). Using this approach, a shape clustering algorithm is proposed to classify healthy and pathological ESA signals generated by a recent ESA model. This model allows a precise simulation of neural mechanisms occurring in the auditory nerve. The obtained results demonstrate that this shape analysis is very sensitive for detecting a small number of fibers with correlated firing, supposed to occur during a particular type of tinnitus. In comparison, the classical spectral index fails in this detection.

  8. Chromosomal abnormalities not currently detected by cell-free fetal DNA: a retrospective analysis at a single center.

    PubMed

    Shani, Hagit; Goldwaser, Tamar; Keating, Jennifer; Klugman, Susan

    2016-06-01

    Cell-free fetal DNA analysis is used as a screening test to identify pregnancies that are at risk for common autosomal and sex chromosome aneuploidies. The purpose of this study was to investigate the chromosomal abnormalities that would not be detected by cell-free fetal DNA in a single medical center. This was a retrospective cohort analysis of 3182 consecutive invasive diagnostic procedures that were performed at Montefiore Medical Center's Division of Reproductive and Medical Genetics from January 1, 2009 to August 31, 2014. All patients underwent cytogenetic analysis; one-third of the patients (1037/3182) went through chromosomal microarray analysis. Clinically significant chromosomal abnormalities were detected in 220 of 3140 cases (7%) after we excluded multiple gestation pregnancies (n = 42). Of these 125 cases (57%) were diagnosed with the common autosomal trisomies that involved chromosomes 21, 18, and 13 and with sex chromosome aneuploidies. There were 23 mosaic karyotypes; 8 of them involved trisomy in chromosomes 21 and 13; 5 of them were sex chromosome mosaics, and 10 of them were other mosaic cases. Five cases of triploidy were detected. Additionally, 19 unbalanced chromosomal rearrangements, a rare autosomal trisomy, and 47 clinically significant findings on chromosomal microarray analysis were diagnosed. Based on the published detection rates of cell-free fetal DNA testing and considering the "no-results" rate, we calculated that 99 of 220 chromosomal changes (45%) could not have been detected by cell-free fetal DNA testing: 16 of the 125 common aneuploidies and sex chromosome aneuploidies, 1 of the 5 triploidy cases, 15 of the 23 mosaic cases, all cases of unbalanced chromosomal rearrangements (n = 19), rare autosomal trisomy (n = 1), and 47 clinically significant chromosomal microarray abnormalities. Current cell-free DNA testing could not detect up to one-half of the clinically significant chromosomal abnormalities that were found, which

  9. High Resolution X Chromosome-Specific Array-CGH Detects New CNVs in Infertile Males

    PubMed Central

    Krausz, Csilla; Giachini, Claudia; Lo Giacco, Deborah; Daguin, Fabrice; Chianese, Chiara; Ars, Elisabet; Ruiz-Castane, Eduard; Forti, Gianni; Rossi, Elena

    2012-01-01

    Context The role of CNVs in male infertility is poorly defined, and only those linked to the Y chromosome have been the object of extensive research. Although it has been predicted that the X chromosome is also enriched in spermatogenesis genes, no clinically relevant gene mutations have been identified so far. Objectives In order to advance our understanding of the role of X-linked genetic factors in male infertility, we applied high resolution X chromosome specific array-CGH in 199 men with different sperm count followed by the analysis of selected, patient-specific deletions in large groups of cases and normozoospermic controls. Results We identified 73 CNVs, among which 55 are novel, providing the largest collection of X-linked CNVs in relation to spermatogenesis. We found 12 patient-specific deletions with potential clinical implication. Cancer Testis Antigen gene family members were the most frequently affected genes, and represent new genetic targets in relationship with altered spermatogenesis. One of the most relevant findings of our study is the significantly higher global burden of deletions in patients compared to controls due to an excessive rate of deletions/person (0.57 versus 0.21, respectively; p = 8.785×10−6) and to a higher mean sequence loss/person (11.79 Kb and 8.13 Kb, respectively; p = 3.435×10−4). Conclusions By the analysis of the X chromosome at the highest resolution available to date, in a large group of subjects with known sperm count we observed a deletion burden in relation to spermatogenic impairment and the lack of highly recurrent deletions on the X chromosome. We identified a number of potentially important patient-specific CNVs and candidate spermatogenesis genes, which represent novel targets for future investigations. PMID:23056185

  10. Alteration of wheat vernalization requirement by alien chromosome-mediated transposition of MITE

    PubMed Central

    Gorafi, Yasir Serag Alnor; Eltayeb, Amin Elsadig; Tsujimoto, Hisashi

    2016-01-01

    Under the changing climate, early flowering is advantageous to escape terminal heat and drought. Previously during evaluation of 14 chromosome introgression lines (ILs), we found three ILs that flowered a month earlier than their wheat background Chinese Spring (CS). This paper describes the cause of the early flowering in the ILs and provides insight into the evolution of spring wheat from the winter wheat. We used specific molecular markers for Vrn genes to determine its allelic composition. Phenotypic evaluations carried out under field conditions and in a growth chamber. Unlike the winter vrn-A1 allele of CS, the spring Vrn-A1 allele of the ILs had insertions of 222 and 131-bp miniature inverted-repeat transposable element (MITE) in the promoter region. Sequence analysis indicated that the 222-bp insertion is similar to an insertion in the spring genotype, Triple Dirk D. Our results ruled out any possibility of outcrossing or contamination. Without vernalization, Vrn-A1 is highly expressed in the ILs compared to CS. We attribute the early flowering of the ILs to the insertion of the MITE in the promoter of Vrn-A1. The alien chromosome might mediate this insertion. PMID:27162490

  11. Alteration of wheat vernalization requirement by alien chromosome-mediated transposition of MITE.

    PubMed

    Gorafi, Yasir Serag Alnor; Eltayeb, Amin Elsadig; Tsujimoto, Hisashi

    2016-03-01

    Under the changing climate, early flowering is advantageous to escape terminal heat and drought. Previously during evaluation of 14 chromosome introgression lines (ILs), we found three ILs that flowered a month earlier than their wheat background Chinese Spring (CS). This paper describes the cause of the early flowering in the ILs and provides insight into the evolution of spring wheat from the winter wheat. We used specific molecular markers for Vrn genes to determine its allelic composition. Phenotypic evaluations carried out under field conditions and in a growth chamber. Unlike the winter vrn-A1 allele of CS, the spring Vrn-A1 allele of the ILs had insertions of 222 and 131-bp miniature inverted-repeat transposable element (MITE) in the promoter region. Sequence analysis indicated that the 222-bp insertion is similar to an insertion in the spring genotype, Triple Dirk D. Our results ruled out any possibility of outcrossing or contamination. Without vernalization, Vrn-A1 is highly expressed in the ILs compared to CS. We attribute the early flowering of the ILs to the insertion of the MITE in the promoter of Vrn-A1. The alien chromosome might mediate this insertion.

  12. Presence of an extra chromosome alters meiotic double-stranded break repair dynamics and MLH1 foci distribution in human oocytes.

    PubMed

    Robles, P; Roig, I; Garcia, R; Brieño-Enríquez, M; Martin, M; Cabero, Ll; Toran, N; Garcia Caldés, M

    2013-03-01

    Studies performed on human trisomic 21 oocytes have revealed that during meiosis, the three homologues 21 synapse and, in some cases, achieve what looks like a trivalent. This implies that meiotic recombination takes place among the three homologous chromosomes 21, and to some extent, crossovers form between them. To see how meiotic recombination is in the presence of an extra chromosome 21, we analyzed the distribution of three recombination markers (γH2AX, RPA, and MLH1) on trisomic 21 oocytes at pachynema and, in particular, on chromosomes 21. Results clearly show how the presence of an extra chromosome 21 alters meiotic recombination progression, leading to the presence of a higher number of early recombination markers at pachynema. Moreover, the distribution on these chromosomes 21 of some of these markers is different in aneuploid oocytes. Finally, there is a substantial increase in the number of MLH1 foci, a marker of most crossovers in mammals, which is related to the number of synapsed chromosomes in pachynema. Thus, bivalents 21 had fewer MLH1 foci than partial or total trivalents, suggesting a close relationship between synapsis and crossover designation. All of the data presented suggest that the presence of an extra chromosome alters meiotic recombination globally in aneuploid human oocytes.

  13. Detection of Turner syndrome using X-chromosome inactivation specific differentially methylated CpG sites: A pilot study.

    PubMed

    Zhang, Qiang; Guo, Xiaohong; Tian, Tian; Wang, Teng; Li, Qiaoli; Wang, Lei; Liu, Yun; Xing, Qinghe; He, Lin; Zhao, Xinzhi

    2017-05-01

    Early diagnosis of Turner syndrome (TS) may improve preventive measures and treatment. X-chromosome inactivation specific differentially methylated CpG sites (XIDMSs) that are high methylated in inactive X chromosomes (Xi) and unmethylated in active X chromosomes (Xa) may be potential makers for TS detection. The candidate XIDMSs were screened from 9 male and 12 female DNA samples with normal karyotypes using the Illumina 450k array and validated by bisulfite sequencing PCR and pyrosequencing assay. X chromosome dosage was calculated according to the methylation level of multiple XIDMSs. Overall, 108 candidate XIDMSs were screened by the 450k array. Validations indicated that XIDMSs gathered and formed the X-chromosome inactivation specific differentially methylated regions (XIDMRs). Using 3 XIDMRs at SAT1, UXT and UTP14A loci, 36 TS, 22 normal female and 6 male samples were analyzed. Methylation levels of the 20 XIDMSs in the XIDMRs could distinguish between TS and normal female DNA samples, the X chromosome dosage was consistent with karyotyping data. Analyzing samples of 2 triple X syndrome and 3 Klinefelter syndrome patients suggested that this method could be used to detect X chromosome aneuploids other than TS. XIDMSs are widely spread along the X chromosome and might be effective markers for detection of TS and other X chromosome aneuploids. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Alteration of the sex determining system resulting from structural change of the sex chromosomes in the frog Rana rugosa.

    PubMed

    Ohtani, H; Miura, I; Hanada, H; Ichikawa, Y

    2000-02-15

    Rana rugosa in Japan is divided into four geographical races on the basis of the karyotype of the sex chromosomes: one in which heteromorphic sex chromosomes occur in the female sex (ZW/ZZ-system), another in which they are present in males (XX/XY-system), and the remaining two in which no heteromorphism is seen in either sex. The last two inherit the XX/XY sex determining system. Y and Z chromosomes in the former two are of the same karyotype as the no. 7 chromosomes seen in one of the latter two, whereas X and W are caused by two inversions that occurred in the original Xs (no. 7). In this study, we first attempted to detect the structural difference between the resulting X and W by examining their chiasma formation. The chiasma distribution between X and W was closely similar to that between two Xs, suggesting that the W and X are identical in structure. Regarding the change from XX/XY- to ZW/ZZ-system, the simplest explanation is that the putative female-determining gene(s) on the W grew functionally stronger by inversions. Next, we examined the sex of triploids having two Xs and one Z. The data showed that the triploids with two original Xs and a Z were all male, whereas most of those with two resulting Xs and a Z developed into females as expected. We speculated that the female-determining gene(s) on the resulting X grew mildly stronger functionally by position effect, whereas those on the W grew much stronger for some other reason (e.g., duplication). J. Exp. Zool. 286:313-319, 2000. Copyright 2000 Wiley-Liss, Inc.

  15. Detection of homologous recombination between yeast artificial chromosomes with overlapping inserts.

    PubMed Central

    Cellini, A; Lacatena, R M; Tocchini-Valentini, G P

    1991-01-01

    We have developed a system which facilitates the detection of recombination between Yeast Artificial Chromosomes (YAC's) carrying homologous inserts. The system consists of a classical YAC vector, a new YAC vector and two appropriately labelled yeast strains of opposite mating type. The new YAC vector differs in markers from the canonical YAC vector. To test whether homologous recombination takes place, phage lambda DNA was cloned in the two vectors to provide a region of homology. The two constructs were then introduced into yeast strains of opposite mating type in which the endogenous genes for the selective markers present in the vectors are not expressed. Artificial chromosomes obtained by meiotic recombination are detected in the spores resulting from the mating. PMID:1826951

  16. 45,X/46,XY chromosome mosaicism detected by midtrimester amniocentesis in amniocyte clones.

    PubMed

    Hecht, F; Hecht, B K

    1982-07-01

    Amniocyte clones from a mild-trimester pregnancy disclosed 45,X/46,XY sex chromosome mosaicism. Because of the uncertainty concerning the phenotype of the fetus, the parents elected to terminate the pregnancy. Mixed (asymmetrical) gonadal dysgenesis was not found. The fetus appeared to have a normal male uro-genital system. No malformations of any type were detected, although as expected, the fetus did have 45,X/46,XY mosaicism.

  17. The NACP/synuclein gene: Chromosomal assignment and screening for alterations in Alzheimer disease

    SciTech Connect

    Campion, D.; Martin, C.; Charbonnier, F.

    1995-03-20

    The major component of the vascular and plaque amyloid deposits in Alzheimer disease is the amyloid {beta} peptide (A{beta}). A second intrinsic component of amyloid, the NAC (non-A{beta} component of amyloid) peptide, has recently been identified, and its precursor protein was named NACP. A computer homology search allowed us to establish that the human NACP gene was homologous to the rat synuclein gene. We mapped the NACP/synuclein gene to chromosome 4 and cloned three alternatively spliced transcripts in lymphocytes derived from a normal subject. We analyzed by RT-PCR and direct sequencing the entire coding region of the NACP/synuclein gene in a group of patients with familial early onset Alzheimer disease. No mutation was found in 26 unrelated patients. Further studies are required to investigate the implication of the NACP/synuclein gene in Alzheimer disease. 21 refs., 3 tabs.

  18. Detection of X chromosome aneuploidy using Southern blot analysis during routine population-based screening for fragile X syndrome.

    PubMed

    Adir, Vardit; Shahak, Elena; Dar, Hanna; Borochowitz, Zvi U

    2003-01-01

    We report herein two cases where detection of X chromosome aneuploidy (cytogenetically proved 45,X/46XX and 47,XXX) was made possible by molecular diagnosis during population-based carrier screening for Fragile X syndrome, using Southern blot analysis. This study emphasizes the value of molecular analysis for gene dosage to suggest chromosomal aneuploidy.

  19. A new direction for prenatal chromosome microarray testing: software-targeting for detection of clinically significant chromosome imbalance without equivocal findings

    PubMed Central

    Bint, Susan; Irving, Melita D.; Kyle, Phillipa M.; Akolekar, Ranjit; Mohammed, Shehla N.; Mackie Ogilvie, Caroline

    2014-01-01

    Purpose. To design and validate a prenatal chromosomal microarray testing strategy that moves away from size-based detection thresholds, towards a more clinically relevant analysis, providing higher resolution than G-banded chromosomes but avoiding the detection of copy number variants (CNVs) of unclear prognosis that cause parental anxiety. Methods. All prenatal samples fulfilling our criteria for karyotype analysis (n = 342) were tested by chromosomal microarray and only CNVs of established deletion/duplication syndrome regions and any other CNV >3 Mb were detected and reported. A retrospective full-resolution analysis of 249 of these samples was carried out to ascertain the performance of this testing strategy. Results. Using our prenatal analysis, 23/342 (6.7%) samples were found to be abnormal. Of the remaining samples, 249 were anonymized and reanalyzed at full-resolution; a further 46 CNVs were detected in 44 of these cases (17.7%). None of these additional CNVs were of clear clinical significance. Conclusion. This prenatal chromosomal microarray strategy detected all CNVs of clear prognostic value and did not miss any CNVs of clear clinical significance. This strategy avoided both the problems associated with interpreting CNVs of uncertain prognosis and the parental anxiety that are a result of such findings. PMID:24795849

  20. Multicolor detection of every chromosome as a means of detecting mosaicism and nuclear organization in human embryonic nuclei.

    PubMed

    Turner, Kara; Fowler, Katie; Fonseka, Gothami; Griffin, Darren; Ioannou, Dimitrios

    2016-06-01

    Fluorescence in-situ hybridization (FISH) revolutionized cytogenetics using fluorescently labelled probes with high affinity with target (nuclear) DNA. By the early 1990s FISH was adopted as a means of preimplantation genetic diagnosis (PGD) sexing for couples at risk of transmitting X-linked disorders and later for detection of unbalanced translocations. Following a rise in popularity of PGD by FISH for sexing and the availability of multicolor probes (5-8 colors), the use of FISH was expanded to the detection of aneuploidy and selective implantation of embryos more likely to be euploid, the rationale being to increase pregnancy rates (referral categories were typically advanced maternal age, repeated IVF failure, repeated miscarriage or severe male factor infertility). Despite initial reports of an increase in implantation rates, reduction in trisomic offspring and spontaneous abortions criticism centered around experimental design (including lack of randomization), inadequate control groups and lack of report on live births. Eleven randomized control trials (RCTs) (2004-2010) showed that preimplantation genetic screening (PGS) with FISH did not increase delivery rates with some demonstrating adverse outcomes. These RCTs, parallel improvements in culturing and cryopreservation and a shift to blastocyst biopsy essentially outdated FISH as a tool for PGS and it has now been replaced by newer technologies (array CGH, SNP arrays, qRT-PCR and NGS). Cell-by-cell follow up analysis of individual blastomeres in non-transferred embryos is however usually prohibitively expensive by these new approaches and thus FISH remains an invaluable resource for the study of mosaicism and nuclear organization. We thus developed the approach described herein for the FISH detection of chromosome copy number of all 24 human chromosomes. This approach involves 4 sequential layers of hybridization, each with 6 spectrally distinct fluorochromes and a bespoke capturing system. Here we report

  1. Detection of genetic alterations in hereditary colorectal cancer screening.

    PubMed

    Pineda, Marta; González, Sara; Lázaro, Conxi; Blanco, Ignacio; Capellá, Gabriel

    2010-11-10

    There are two major hereditary colorectal cancer syndromes: Adenomatous Polyposis, secondary to APC germline alterations (FAP, Familial Adenomatous Polyposis) or secondary to MUTYH germline alterations (MAP, MUTYH associated Polyposis), and Lynch syndrome, associated with germline mutations in mismatch repair genes (MLH1, MSH2, MSH6 and PMS2). The elucidation of their genetic basis has depicted an increasingly complex picture that has lead to the implementation of complex diagnostic algorithms that include both tumor profiling and germline analyses. A variety of techniques at the DNA, RNA and protein level are used to screen for molecular alterations both in tumor biopsies (microsatellite instability analysis, mismatch repair protein immunohistochemistry, BRAF-Val600Glu detection and MLH1 promoter hypermethylation analysis) and in the germline (point mutation screening, copy number assessment). Also functional tests are more often used to characterize variants of unknown significance. Methodological issues associated with the techniques analyzed, as well as the algorithms used, are discussed. 2009 Elsevier B.V. All rights reserved.

  2. Detection of Copy Number Alterations Using Single Cell Sequencing.

    PubMed

    Knouse, Kristin A; Wu, Jie; Hendricks, Austin

    2017-02-17

    Detection of genomic changes at single cell resolution is important for characterizing genetic heterogeneity and evolution in normal tissues, cancers, and microbial populations. Traditional methods for assessing genetic heterogeneity have been limited by low resolution, low sensitivity, and/or low specificity. Single cell sequencing has emerged as a powerful tool for detecting genetic heterogeneity with high resolution, high sensitivity and, when appropriately analyzed, high specificity. Here we provide a protocol for the isolation, whole genome amplification, sequencing, and analysis of single cells. Our approach allows for the reliable identification of megabase-scale copy number variants in single cells. However, aspects of this protocol can also be applied to investigate other types of genetic alterations in single cells.

  3. Detection of structural and numerical chromosomal abnormalities by ACM-FISH analysis in sperm of oligozoospermic infertility patients.

    PubMed

    Schmid, T E; Brinkworth, M H; Hill, F; Sloter, E; Kamischke, A; Marchetti, F; Nieschlag, E; Wyrobek, A J

    2004-06-01

    Modern reproductive technologies are enabling the treatment of infertile men with severe disturbances of spermatogenesis. The possibility of elevated frequencies of genetically and chromosomally defective sperm has become an issue of concern with the increased usage of ICSI, which can enable men with severely impaired sperm production to father children. Several papers have been published reporting aneuploidy in oligozoospermic patients, but relatively little is known about chromosome structural aberrations in the sperm of these patients. We examined sperm from infertile, oligozoospermic individuals for structural and numerical chromosomal abnormalities using a multicolour ACM fluorescence in situ hybridization (FISH) assay that utilizes DNA probes specific for three regions of chromosome 1 to detect human sperm that carry numerical chromosomal abnormalities plus two categories of structural aberrations: duplications and deletions of 1pter and 1cen, and chromosomal breaks within the 1cen-1q12 region. There was a significant increase in the average frequencies of sperm with duplications and deletions in the infertility patients compared with the healthy concurrent controls. There was also a significantly elevated level of breaks within the 1cen-1q12 region. There was no evidence for an increase in chromosome 1 disomy, or in diploidy. Our data reveal that oligozoospermia is associated with chromosomal structural abnormalities, suggesting that oligozoospermic men carry a higher burden of transmissible, chromosome damage. The findings raise the possibility of elevated levels of transmissible chromosomal defects following ICSI treatment.

  4. Efficient detection of chromosome imbalances and exome single nucleotide variants using targeted sequencing in the clinical setting.

    PubMed

    Villela, Darine; da Costa, Silvia Souza; Vianna-Morgante, Angela M; Krepischi, Ana C V; Rosenberg, Carla

    2017-09-04

    We evaluated an approach to detect copy number variants (CNVs) and single nucleotide changes (SNVs), using a clinically focused exome panel complemented with a backbone and SNP probes that allows for genome-wide copy number changes and copy-neutral absence of heterozygosity (AOH) calls; this approach potentially substitutes the use of chromosomal microarray testing and sequencing into a single test. A panel of 16 DNA samples with known alterations ranging from megabase-scale CNVs to single base modifications were used as positive controls for sequencing data analysis. The DNA panel included CNVs (n = 13) of variable sizes (23 Kb to 27 Mb), uniparental disomy (UPD; n = 1), and single point mutations (n = 2). All DNA sequence changes were identified by the current platform, showing that CNVs of at least 23 Kb can be properly detected. The estimated size of genomic imbalances detected by microarrays and next generation sequencing are virtually the same, indicating that the resolution and sensitivity of this approach are at least similar to those provided by DNA microarrays. Accordingly, our data show that the combination of a sequencing platform comprising focused exome and whole genome backbone, with appropriate algorithms, enables a cost-effective and efficient solution for the simultaneous detection of CNVs and SNVs. Copyright © 2017. Published by Elsevier Masson SAS.

  5. Evaluation of comprehensive chromosome screening platforms for the detection of mosaic segmental aneuploidy.

    PubMed

    Goodrich, David; Xing, Tongji; Tao, Xin; Lonczak, Agnieszka; Zhan, Yiping; Landis, Jessica; Zimmerman, Rebekah; Scott, Richard T; Treff, Nathan R

    2017-08-01

    A subset of preimplantation embryos identified as euploid may in fact possess both whole and sub-chromosomal mosaicism, raising concerns regarding the predictive value of current comprehensive chromosome screening (CCS) methods utilizing a single biopsy. Current CCS methods may be capable of detecting sub-chromosomal mosaicism in a trophectoderm biopsy by examining intermediate levels of segmental aneuploidy within a biopsy. This study evaluates the sensitivity and specificity of segmental aneuploidy detection by three commercially available CCS platforms utilizing a cell line mixture model of segmental mosaicism in a six-cell trophectoderm biopsy. Two cell lines with known karyotypes were obtained and mixed together at specific ratios of six total cells (0:6, 1:5, 2:4, 3:3, 4:2, 5:1, and 6:0). A female cell line containing a 16.2 Mb deletion on chromosome 5 and a male cell line containing a 25.5 Mb deletion on chromosome 4 were used to create mixtures at each level. Six replicates of each mixture were prepared, randomized, and blinded for analysis by one of the three CCS platforms (SNP-array, VeriSeq NGS, or NexCCS). Sensitivity and specificity of segmental aneuploidy at each level of mosaicism was determined and compared between each platform. Additionally, an alternative VeriSeq NGS analysis method utilizing previously published criteria was evaluated. Examination of the default settings of each platform revealed that the sensitivity was significantly different between NexCCS and SNP up to 50% mosaicism, custom VeriSeq, and SNP-array up to 66% mosaicism, and between NexCCS and custom VeriSeq up to 50% mosaicism. However, no statistical difference was observed in mixtures with >50% mosaicism with any platform. No comparison was made between default VeriSeq, as it does not report segmental imbalances. Furthermore, while the use of previously published criteria for VeriSeq NGS significantly increased sensitivity at low levels of mosaicism, a significant decrease

  6. Prenatal detection of chromosome aneuploidies in uncultured chorionic villus samples by FISH

    SciTech Connect

    Bryndorf, T.; Christensen, B.; Vad, M.; Philip, J.

    1996-10-01

    We developed a 1-d FISH assay for detection of numerical chromosome abnormalities in uncultured chorionic villus samples (CVS). Probes specific for chromosomes 13, 18, 21, X, and Y were used to determine ploidy by analysis of signal number in hybridized nuclei. Aneuploidy detection using this assay was directly compared with the results obtained by conventional cytogenetic analysis in a consecutive, clinical study of 2,709 CVS and placental samples. The FISH assay yielded discrete differences in the signal profiles between cytogenetically normal and abnormal samples. On the basis of these results, we generated FISH-assay cutoff values that discriminated between karyotypically normal and aneuploid samples. Samples with mosaicism and a single sample with possible heritable small chromosome X probe target were exceptions and showed poor agreement between FISH results and conventional cytogenetics. We conclude that the FISH assay may act as a more accurate and less labor-demanding alternative to {open_quotes}direct{close_quotes} CVS analysis. 22 refs., 1 fig., 4 tabs.

  7. [Detecting selection signatures on X chromosome in pig through high density SNPs].

    PubMed

    Ma, Yun-Long; Zhang, Qin; Ding, Xiang-Dong

    2012-10-01

    In the process of domestic pig breeding, many important economic traits were subject to strong artificial se-lection pressure. With the availability of high density single nucleotide polymorphism (SNP) markers in farm animals, selection occurring in those traits could be traced by detecting selection signatures on genome, and the genes experiencing selection can also be further mined based on selection signatures. Due to the special characteristic of X chromosome, many approaches of genetic analysis fitted for autosome are not plausible for X chromosome. Fortunately, detecting selection signature provides an effective tool to settle such situation. In this study, the Cross Population Extend Haplotype Homozygosity Test (XP-EHH) was implemented to identify selection signatures on chromosome X in three pig breeds (Landrace, Songliao, and Yorkshire) using high density SNPs, and the genes located within selection signature regions were revealed through bioinformatic analysis. In total, 29, 13, and 15 selection signature regions, with 3.59, 4.92, and 4.07 SNPs on average in each region, were identified in Landrace, Songliao, and Yorkshire, respectively. Some overlaps of selection signature regions were observed between Songliao and Landrace, and between Landrace and Yorkshire, while no overlaps between Yorkshire and Songliao were found. Bioinformatic analysis revealed that many genes in the selection signature regions were related to reproduction and immune traits, and some of them have not been reported in pigs, which might serve as important candidate genes in future study.

  8. Beyond Trisomy 21: Additional Chromosomal Anomalies Detected through Routine Aneuploidy Screening

    PubMed Central

    Metcalfe, Amy; Hippman, Catriona; Pastuck, Melanie; Johnson, Jo-Ann

    2014-01-01

    Prenatal screening is often misconstrued by patients as screening for trisomy 21 alone; however, other chromosomal anomalies are often detected. This study aimed to systematically review the literature and use diagnostic meta-analysis to derive pooled detection and false positive rates for aneuploidies other than trisomy 21 with different prenatal screening tests. Non-invasive prenatal testing had the highest detection (DR) and lowest false positive (FPR) rates for trisomy 13 (DR: 90.3%; FPR: 0.2%), trisomy 18 (DR: 98.1%; FPR: 0.2%), and 45,X (DR: 92.2%; FPR: 0.1%); however, most estimates came from high-risk samples. The first trimester combined test also had high DRs for all conditions studied (trisomy 13 DR: 83.1%; FPR: 4.4%; trisomy 18 DR: 91.9%; FPR: 3.5%; 45,X DR: 70.1%; FPR: 5.4%; triploidy DR: 100%; FPR: 6.3%). Second trimester triple screening had the lowest DRs and highest FPRs for all conditions (trisomy 13 DR: 43.9%; FPR: 8.1%; trisomy 18 DR: 70.5%; FPR: 3.3%; 45,X DR: 77.2%; FPR: 9.3%). Prenatal screening tests differ in their ability to accurately detect chromosomal anomalies. Patients should be counseled about the ability of prenatal screening to detect anomalies other than trisomy 21 prior to undergoing screening. PMID:26237381

  9. Alterations in p16 and p53 genes and chromosomal findings in patients with lung cancer: fluorescence in situ hybridization and cytogenetic studies.

    PubMed

    Demirhan, Osman; Taştemir, Deniz; Hastürk, Serap; Kuleci, Sedat; Hanta, Ismail

    2010-08-01

    Chromosomal aberrations and instability of gene(s) are two factors related to the genetic instability of cancer cells. A loss of the tumor-suppressor function of the genes p16 and p53 is the most common event leading to the development of human cancers. Carcinoma of the lung is the leading cause of cancer deaths in the world. Chromosomal abnormalities in lung cancer may provide a valuable clue to the identification of target loci and culminate in a successful search for the major genes. The aim of this study was to investigate (i) alterations of the p16 and p53 genes and (ii) chromosomal aberrations in patients with small cell and non-small cell lung cancer by fluorescence in situ hybridization (FISH) and cytogenetic studies. We carried out cytogenetic analysis by Giemsa-banding in 18 cases. FISH probes for the p16 and p53 genes were also used on interphase nuclei to screen the alterations in these genes in lung cancer (LC). We observed a high frequency of losses of the p16 - in 8/18 (44%) - and p53 - in 7/18 (39%) - genes in the cases with LC. A total of 18 patients showed predominantly numerical and structural aberrations. Among these two types, structural aberrations predominated and usually consisted of deletions, breaks, and fragilities in various chromosomes. Both structural and numerical changes were observed in almost all patients. Chromosomes 3 and 1 were found to be most frequently involved in structural abnormalities, followed by chromosomes 6, 9, and 8. Autosomal aneuploidies were also observed to be the most frequent (chromosomes 22, 19, 18, 20, 9, and 17), followed by those of the X and Y chromosomes. The expression of fragile sites was also found to be significantly higher in seven chromosomal regions: 3p14, 1q21, 1q12, 6q26, 9q13, 8q22, and 8q24. Our data confirmed that DNA damage and genomic instability may be factors contributing to the mutation profile and development of lung cancer. The patients who developed lung cancer showed a high frequency

  10. CAFE: an R package for the detection of gross chromosomal abnormalities from gene expression microarray data

    PubMed Central

    Bollen, Sander; Mah, Nancy

    2014-01-01

    Summary: The current methods available to detect chromosomal abnormalities from DNA microarray expression data are cumbersome and inflexible. CAFE has been developed to alleviate these issues. It is implemented as an R package that analyzes Affymetrix *.CEL files and comes with flexible plotting functions, easing visualization of chromosomal abnormalities. Availability and implementation: CAFE is available from https://bitbucket.org/cob87icW6z/cafe/ as both source and compiled packages for Linux and Windows. It is released under the GPL version 3 license. CAFE will also be freely available from Bioconductor. Contact: sander.h.bollen@gmail.com or nancy.mah@mdc-berlin.de Supplementary information: Supplementary data are available at Bioinformatics online. PMID:24451624

  11. Upregulation of three Drosophila homologs of human chromosome 21 genes alters synaptic function: Implications for Down syndrome

    PubMed Central

    Chang, Karen T.; Min, Kyung-Tai

    2009-01-01

    At the neuronal level of Down syndrome (DS) brains, there are evidences of altered shape, number, and density of synapses, as well as aberrant endocytosis associated with accumulation of enlarged endosomes, suggesting that proteins involved in synaptic vesicle recycling may play key roles in DS neurons. However, the exact mechanism underlying those anomalies is not well understood. We hypothesize that overexpression of three genes, dap160/itsn1, synj/synj1, and nla/dscr1, located on human chromosome 21 play important roles in DS neurons. Here, we systematically investigate the effects of multiple gene overexpression on synaptic morphology and endocytosis to identify possible dominant gene or genes. We found that overexpression of individual genes lead to abnormal synaptic morphology, but all three genes are necessary to cause impaired vesicle recycling and affect locomotor vigor. Furthermore, we report that dap160 overexpression alters the subcellular distribution of synaptojanin, and overexpression of nla regulates the phosphoinositol 5′ phosphatase activity of synaptojanin. These findings imply that restoring the level of any one of these genes may reduce endocytic defects seen in DS. PMID:19805187

  12. Upregulation of three Drosophila homologs of human chromosome 21 genes alters synaptic function: implications for Down syndrome.

    PubMed

    Chang, Karen T; Min, Kyung-Tai

    2009-10-06

    At the neuronal level of Down syndrome (DS) brains, there are evidences of altered shape, number, and density of synapses, as well as aberrant endocytosis associated with accumulation of enlarged endosomes, suggesting that proteins involved in synaptic vesicle recycling may play key roles in DS neurons. However, the exact mechanism underlying those anomalies is not well understood. We hypothesize that overexpression of three genes, dap160/itsn1, synj/synj1, and nla/dscr1, located on human chromosome 21 play important roles in DS neurons. Here, we systematically investigate the effects of multiple gene overexpression on synaptic morphology and endocytosis to identify possible dominant gene or genes. We found that overexpression of individual genes lead to abnormal synaptic morphology, but all three genes are necessary to cause impaired vesicle recycling and affect locomotor vigor. Furthermore, we report that dap160 overexpression alters the subcellular distribution of synaptojanin, and overexpression of nla regulates the phosphoinositol 5' phosphatase activity of synaptojanin. These findings imply that restoring the level of any one of these genes may reduce endocytic defects seen in DS.

  13. Unexpected rates of chromosomal instabilities and alterations of hormone levels in Namibian uranium miners

    SciTech Connect

    Zaire, R.; Notter, M.; Thiel, E.

    1997-05-01

    A common problem in determining the health consequences of radiation exposure is factoring out other carcinogenic influences. The conditions in Namibia provide a test case for distinguishing the effects of long-term low-dose exposure to uranium from the other environmental factors because of good air quality and the lack of other industries with negative health effects. Present records indicate a much higher prevalence of cancer among male workers in the open-pit uranium mine in Namibia compared with the general population. The objective of the present study was to determine whether long-term exposure to low doses of uranium increases the risk of a biological radiation damage which would lead to malignant diseases and to derive a dose-response model for these miners. To investigate this risk, we measured uranium excretion in urine, neutrophil counts and the serum level of FSH, LH and testosterone and analyzed chromosome aberrations in whole blood cells using fluorescence in situ hybridization. A representative cohort of 75 non-smoking, HIV-negative miners was compared to a control group of 31 individuals with no occupational history in mining. A sixfold increase in uranium excretion among the miners compared to the controls was recorded (P < 0.001). Furthermore, we determined a significant reduction in testosterone levels (P < 0.008) and neutrophil count (P < 0.0001). Most remarkably, cells with multiple aberrations such as {open_quotes}rogue{close_quotes} cells were observed for the first time in miners; these cells had previously been found only after short-term high-dose radiation exposure, e.g. from the Hiroshima atomic bomb or the Chernobyl accident. 19 refs., 1 fig., 3 tabs.

  14. Detection of Dental Fluorosis-Associated Quantitative Trait Loci on Mouse Chromosomes 2 and 11

    PubMed Central

    Everett, Eric T.; Yan, Dong; Weaver, Marjorie; Liu, Lixiang; Foroud, Tatiana; Martinez-Mier, E. Angeles

    2008-01-01

    Systemic exposure to greater than optimal fluoride (F) can lead to dental fluorosis (DF). Parental A/J (DF-susceptible) and 129P3/J (DF-resistant) inbred mice were used for histological studies and to generate F2 progeny. Mice were treated with 0 or 50 ppm F in their drinking water for 60 days. A clinical criterion (modified Thylstrup and Fejerskov categorical scale) was used to assess the severity of DF for each individual F2 animal. Parental strains were subjected to histological examination of maturing enamel. F treatment resulted in accumulation of amelogenins in the maturing enamel of A/J mice. Quantitative trait loci (QTL) detection was performed using phenotypic extreme F2 animals genotyped for 354 single nucleotide polymorphism-based markers distributed throughout the mouse genome followed by χ2 analysis. Significant evidence of association was observed on chromosomes 2 and 11 for a series of consecutive markers (p < 0.0001). Further analyses were performed to examine whether the phenotypic effects were found in both male and female F2 mice or whether there was evidence for gender-specific effects. Analyses performed using the markers on chromosomes 2 and 11 which were significant in the mixed-gender mice were also significant when analyses were limited to only the male or female mice. The QTL detected on chromosomes 2 and 11 which influence the variation in response to fluorosis have their effect in mice of both genders. Finally, the QTL in both chromosomes appear to have an additive effect. PMID:18701810

  15. Reverse transcription-polymerase chain reaction detection of transcribed sequences on human chromosome 21

    SciTech Connect

    Cheng, J.F.; Zhu, Y. )

    1994-03-15

    Seventy-four pairs of oligonucleotides derived from sequence-tagged sites (STSs) on the long arm of human chromosome 21, specifically from bands 21q22.1 to 21q22.3, were used in reverse transcription-polymerase chain reactions (RT-PCR) to detect the presence of expressed sequences in a fetal brain. These STSs included 69 that had not been related to transcribed sequences and 5 that had detected two known genes and three previously isolated cDNA clones. Of the 69 STSs analyzed in RT-PCR, 25 allowed amplification of specific cDNA fragments. The sizes of amplified cDNA fragments match those amplified from either human genomic DNA or somatic hybrid cells containing human chromosome 21. Of the 11 cDNA analyzed in Northern blot hybridizations, 6 hybridized to specific RNA species. The rapid screening for cDNA using previously mapped STSs has provided insight into the distribution of expressed sequences in this region of chromosome 21. Northern blot analysis of the amplified cDNA fragments has revealed interesting candidate genes in two disease loci. The marker D21S267 was previously mapped in the Down syndrome region of chromosome 21, and the marker D21S113 is closely linked to progressive myoclonus epilepsy. The cDNA fragments amplified using the primer sequences derived from D21S267 and D21S113 hybridized to 7- and 6.5-kb transcripts, respectively, which seems to express predominantly in brain. 37 refs., 3 figs., 1 tab.

  16. Buccal cell FISH and blood PCR-Y detect high rates of X chromosomal mosaicism and Y chromosomal derivatives in patients with Turner syndrome.

    PubMed

    Freriks, Kim; Timmers, Henri J L M; Netea-Maier, Romana T; Beerendonk, Catharina C M; Otten, Barto J; van Alfen-van der Velden, Janiëlle A E M; Traas, Maaike A F; Mieloo, Hanneke; van de Zande, Guillaume W H J F L; Hoefsloot, Lies H; Hermus, Ad R M M; Smeets, Dominique F C M

    2013-09-01

    Turner syndrome (TS) is the result of (partial) X chromosome monosomy. In general, the diagnosis is based on karyotyping of 30 blood lymphocytes. This technique, however, does not rule out tissue mosaicism or low grade mosaicism in the blood. Because of the associated risk of gonadoblastoma, mosaicism is especially important in case this involves a Y chromosome. We investigated different approaches to improve the detection of mosaicisms in 162 adult women with TS (mean age 29.9 ± 10.3). Standard karyotyping identified 75 patients (46.3%) with a non-mosaic monosomy 45,X. Of these 75 patients, 63 underwent additional investigations including FISH on buccal cells with X- and Y-specific probes and PCR-Y on blood. FISH analysis of buccal cells revealed a mosaicism in 19 of the 63 patients (30.2%). In five patients the additional cell lines contained a (derivative) Y chromosome. With sensitive real-time PCR we confirmed the presence of this Y chromosome in blood in three of the five cases. Although Y chromosome material was established in ovarian tissue in two patients, no gonadoblastoma was found. Our results confirm the notion that TS patients with 45,X on conventional karyotyping often have tissue specific mosaicisms, some of which include a Y chromosome. Although further investigations are needed to estimate the risk of gonadoblastoma in patients with Y chromosome material in buccal cells, we conclude that FISH or real-time PCR on buccal cells should be considered in TS patients with 45,X on standard karyotyping.

  17. A Multi-Marker Model for Detecting Chromosomal Segments Displaying Qtl Activity

    PubMed Central

    Rodolphe, F.; Lefort, M.

    1993-01-01

    A statistical method is presented for detecting quantitative trait loci (QTLs), based on the linear model. Unlike methods able to detect a few well separated QTLs and to estimate their effects and positions, this method considers the genome as a whole and enables the detection of chromosomal segments involved in the differences between two homozygous lines, and their backcross, doubled haploid, or F(2) progenies, for a quantitative trait. Genetic markers must be codominant, but missing markers are accepted, provided they are missing independently from the experiment. Asymptotic properties, which are of practical use, are developed. This method does not rely on strong genetic hypotheses, and thus does not permit any precise genetic analysis of the trait under study, but it does assess which regions of the genome are involved, whatever the complexity of the genetic determinism (number, effects and interactions among QTLs). Simultaneous use of several methods, including this one, should lead to better efficiency in QTL detection. PMID:8375662

  18. Overview of recurrent chromosomal losses in retinoblastoma detected by low coverage next generation sequencing.

    PubMed

    García-Chequer, A J; Méndez-Tenorio, A; Olguín-Ruiz, G; Sánchez-Vallejo, C; Isa, P; Arias, C F; Torres, J; Hernández-Angeles, A; Ramírez-Ortiz, M A; Lara, C; Cabrera-Muñoz, M L; Sadowinski-Pine, S; Bravo-Ortiz, J C; Ramón-García, G; Diegopérez-Ramírez, J; Ramírez-Reyes, G; Casarrubias-Islas, R; Ramírez, J; Orjuela, M A; Ponce-Castañeda, M V

    2016-03-01

    Genes are frequently lost or gained in malignant tumors and the analysis of these changes can be informative about the underlying tumor biology. Retinoblastoma is a pediatric intraocular malignancy, and since deletions in chromosome 13 have been described in this tumor, we performed genome wide sequencing with the Illumina platform to test whether recurrent losses could be detected in low coverage data from DNA pools of Rb cases. An in silico reference profile for each pool was created from the human genome sequence GRCh37p5; a chromosome integrity score and a graphics 40 Kb window analysis approach, allowed us to identify with high resolution previously reported non random recurrent losses in all chromosomes of these tumors. We also found a pattern of gains and losses associated to clear and dark cytogenetic bands respectively. We further analyze a pool of medulloblastoma and found a more stable genomic profile and previously reported losses in this tumor. This approach facilitates identification of recurrent deletions from many patients that may be biological relevant for tumor development. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Overview of recurrent chromosomal losses in retinoblastoma detected by low coverage next generation sequencing

    PubMed Central

    García-Chequer, A.J.; Méndez-Tenorio, A.; Olguín-Ruiz, G.; Sánchez-Vallejo, C.; Isa, P.; Arias, C.F.; Torres, J.; Hernández-Angeles, A.; Ramírez-Ortiz, M.A.; Lara, C.; Cabrera-Muñoz, M.L.; Sadowinski-Pine, S.; Bravo-Ortiz, J.C.; Ramón-García, G.; Diegopérez-Ramírez, J.; Ramírez-Reyes, G.; Casarrubias-Islas, R.; Ramírez, J.; Orjuela, M.A.; Ponce-Castañeda, M.V.

    2016-01-01

    Genes are frequently lost or gained in malignant tumors and the analysis of these changes can be informative about the underlying tumor biology. Retinoblastoma is a pediatric intraocular malignancy, and since deletions in chromosome 13 have been described in this tumor, we performed genome wide sequencing with the Illumina platform to test whether recurrent losses could be detected in low coverage data from DNA pools of Rb cases. An in silico reference profile for each pool was created from the human genome sequence GRCh37p5; a chromosome integrity score and a graphics 40 Kb window analysis approach, allowed us to identify with high resolution previously reported non random recurrent losses in all chromosomes of these tumors. We also found a pattern of gains and losses associated to clear and dark cytogenetic bands respectively. We further analyze a pool of medulloblastoma and found a more stable genomic profile and previously reported losses in this tumor. This approach facilitates identification of recurrent deletions from many patients that may be biological relevant for tumor development. PMID:26883451

  20. Multiscale image enhancement of chromosome banding patterns

    NASA Astrophysics Data System (ADS)

    Wu, Qiang; Castleman, Kenneth R.

    1996-10-01

    Visual examination of chromosome banding patterns is an important means of chromosome analysis. Cytogeneticists compare their patient's chromosome image against the prototype normal/abnormal human chromosome banding patterns. Automated chromosome analysis instruments facilitate this by digitally enhancing the chromosome images. Currently available systems employing traditional highpass/bandpass filtering and/or histogram equalization are approximately equivalent to photomicroscopy in their ability to support the detection of band pattern alterations. Improvements in chromosome image display quality, particularly in the detail of the banding pattern, would significantly increase the cost-effectiveness of these systems. In this paper we present our work on the use of multiscale transform and derivative filtering for image enhancement of chromosome banding patterns. A steerable pyramid representation of the chromosome image is generated by a multiscale transform. The derivative filters are designed to detect the bands of a chromosome, and the steerable pyramid transform is chosen based on its desirable properties of shift and rotation invariance. By processing the transform coefficients that correspond to the bands of the chromosome in the pyramid representation, contrast enhancement of the chromosome bands can be achieved with designed flexibility in scale, orientation and location. Compared with existing chromosome image enhancement techniques, this new approach offers the advantage of selective chromosome banding pattern enhancement that allows designated detail analysis. Experimental results indicate improved enhancement capabilities and promise more effective visual aid to comparison of chromosomes to the prototypes and to each other. This will increase the ability of automated chromosome analysis instruments to assist the evaluation of chromosome abnormalities in clinical samples.

  1. Quantitative fluorescent-PCR detection of sex chromosome aneuploidies and AZF deletions/duplications.

    PubMed

    Plaseski, Toso; Noveski, Predrag; Trivodalieva, Svetlana; Efremov, Georgi D; Plaseska-Karanfilska, Dijana

    2008-12-01

    The most common genetic causes of spermatogenic failure are sex chromosomal abnormalities (most frequently Klinefelter's syndrome) and deletions of the azoospermia factor (AZF) regions (AZFa, AZFb, and AZFc) of the Y chromosome. Several studies have proposed that partial AZFc deletions/duplications may be a risk factor for spermatogenic impairment. We describe a multiplex quantitative fluorescent-polymerase chain reaction (QF-PCR) method that allows simultaneous detection of these genetic causes and risk factors of male infertility. The 11-plex QF-PCR permitted the amplification of the amelogenin gene, four polymorphic X-specific short tandem repeat (STR) markers (XHPRT, DXS6803, DXS981, and exon 1 of the androgen receptor gene), nonpolymorphic Y-specific marker (SRY gene), polymorphic Y-specific STR marker (DYS448), and coamplification of DAZ/DAZL, MYPT2Y/MYPT2, and two CDY2/CDY1 fragments that allow for determination of the DAZ, MYPT2Y, and CDY gene copy number. A total of 357 DNA samples from infertile/subfertile men (n = 205) and fertile controls (n = 152) was studied. We detected 14 infertile males with sex chromosome aneuploidy (10 with Klinefelter's syndrome, 2 XX, and 2 XYY males). All previously detected AZF deletions, that is, AZFc (n8), AZFb (n1), AZFb + c (n1), gr/gr (n11), gr/gr with b2/b4 duplication (n3), and b2/b3 (n5), gave a specific pattern with the 11-plex QF-PCR. In addition, 32 DNA samples showed a pattern consistent with presence of gr/gr or b2/b4 and 4 with b2/b3 duplication. We conclude that multiplex QF-PCR is a rapid, simple, reliable, and inexpensive method that can be used as a first-step genetic analysis in infertile/subfertile patients.

  2. Microarray comparative genomic hybridization detection of chromosomal imbalances in uterine cervix carcinoma

    PubMed Central

    Hidalgo, Alfredo; Baudis, Michael; Petersen, Iver; Arreola, Hugo; Piña, Patricia; Vázquez-Ortiz, Guelaguetza; Hernández, Dulce; González, José; Lazos, Minerva; López, Ricardo; Pérez, Carlos; García, José; Vázquez, Karla; Alatorre, Brenda; Salcedo, Mauricio

    2005-01-01

    Background Chromosomal Comparative Genomic Hybridization (CGH) has been applied to all stages of cervical carcinoma progression, defining a specific pattern of chromosomal imbalances in this tumor. However, given its limited spatial resolution, chromosomal CGH has offered only general information regarding the possible genetic targets of DNA copy number changes. Methods In order to further define specific DNA copy number changes in cervical cancer, we analyzed 20 cervical samples (3 pre-malignant lesions, 10 invasive tumors, and 7 cell lines), using the GenoSensor microarray CGH system to define particular genetic targets that suffer copy number changes. Results The most common DNA gains detected by array CGH in the invasive samples were located at the RBP1-RBP2 (3q21-q22) genes, the sub-telomeric clone C84C11/T3 (5ptel), D5S23 (5p15.2) and the DAB2 gene (5p13) in 58.8% of the samples. The most common losses were found at the FHIT gene (3p14.2) in 47% of the samples, followed by deletions at D8S504 (8p23.3), CTDP1-SHGC- 145820 (18qtel), KIT (4q11-q12), D1S427-FAF1 (1p32.3), D9S325 (9qtel), EIF4E (eukaryotic translation initiation factor 4E, 4q24), RB1 (13q14), and DXS7132 (Xq12) present in 5/17 (29.4%) of the samples. Conclusion Our results confirm the presence of a specific pattern of chromosomal imbalances in cervical carcinoma and define specific targets that are suffering DNA copy number changes in this neoplasm. PMID:16004614

  3. Detection of Chromosomal Inversions Using Non-Repetitive Nucleic Acid Probes

    NASA Technical Reports Server (NTRS)

    Bailey, Susan M. (Inventor); Ray, F. Andrew (Inventor); Goodwin, Edwin H. (Inventor); Bedford, Joel S. (Inventor); Cornforth, Michael N. (Inventor)

    2014-01-01

    A method and a kit for the identification of chromosomal inversions are described. Single-stranded sister chromatids are generated, for example by CO-FISH. A plurality of non-repetitive, labeled probes of relatively small size are hybridized to portions of only one of a pair of single-stranded sister chromatids. If no inversion exists, all of the probes will hybridize to a first chromatid. If an inversion has occurred, these marker probes will be detected on the sister chromatid at the same location as the inversion on the first chromatid.

  4. Detection of chromosomal inversions using non-repetitive nucleic acid probes

    NASA Technical Reports Server (NTRS)

    Bailey, Susan M. (Inventor); Ray, F. Andrew (Inventor); Goodwin, Edwin H. (Inventor); Bedford, Joel S. (Inventor); Cornforth, Michael N. (Inventor)

    2012-01-01

    A method for the identification of chromosomal inversions is described. Single-stranded sister chromatids are generated, for example by CO-FISH. A plurality of non-repetitive, labeled probes of relatively small size are hybridized to portions of only one of a pair of single-stranded sister chromatids. If no inversion exists, all of the probes will hybridize to a first chromatid. If an inversion has occurred, these marker probes will be detected on the sister chromatid at the same location as the inversion on the first chromatid.

  5. Chromosomal microarray analysis as a first-line test in pregnancies with a priori low risk for the detection of submicroscopic chromosomal abnormalities

    PubMed Central

    Fiorentino, Francesco; Napoletano, Stefania; Caiazzo, Fiorina; Sessa, Mariateresa; Bono, Sara; Spizzichino, Letizia; Gordon, Anthony; Nuccitelli, Andrea; Rizzo, Giuseppe; Baldi, Marina

    2013-01-01

    In this study, we aimed to explore the utility of chromosomal microarray analysis (CMA) in groups of pregnancies with a priori low risk for detection of submicroscopic chromosome abnormalities, usually not considered an indication for testing, in order to assess whether CMA improves the detection rate of prenatal chromosomal aberrations. A total of 3000 prenatal samples were processed in parallel using both whole-genome CMA and conventional karyotyping. The indications for prenatal testing included: advanced maternal age, maternal serum screening test abnormality, abnormal ultrasound findings, known abnormal fetal karyotype, parental anxiety, family history of a genetic condition and cell culture failure. The use of CMA resulted in an increased detection rate regardless of the indication for analysis. This was evident in high risk groups (abnormal ultrasound findings and abnormal fetal karyotype), in which the percentage of detection was 5.8% (7/120), and also in low risk groups, such as advanced maternal age (6/1118, 0.5%), and parental anxiety (11/1674, 0.7%). A total of 24 (0.8%) fetal conditions would have remained undiagnosed if only a standard karyotype had been performed. Importantly, 17 (0.6%) of such findings would have otherwise been overlooked if CMA was offered only to high risk pregnancies.The results of this study suggest that more widespread CMA testing of fetuses would result in a higher detection of clinically relevant chromosome abnormalities, even in low risk pregnancies. Our findings provide substantial evidence for the introduction of CMA as a first-line diagnostic test for all pregnant women undergoing invasive prenatal testing, regardless of risk factors. PMID:23211699

  6. Detection of Inter-chromosomal Stable Aberrations by Multiple Fluorescence In Situ Hybridization (mFISH) and Spectral Karyotyping (SKY) in Irradiated Mice.

    PubMed

    Pathak, Rupak; Koturbash, Igor; Hauer-Jensen, Martin

    2017-01-11

    Ionizing radiation (IR) induces numerous stable and unstable chromosomal aberrations. Unstable aberrations, where chromosome morphology is substantially compromised, can easily be identified by conventional chromosome staining techniques. However, detection of stable aberrations, which involve exchange or translocation of genetic materials without considerable modification in the chromosome morphology, requires sophisticated chromosome painting techniques that rely on in situ hybridization of fluorescently labeled DNA probes, a chromosome painting technique popularly known as fluorescence in situ hybridization (FISH). FISH probes can be specific for whole chromosome/s or precise sub-region on chromosome/s. The method not only allows visualization of stable aberrations, but it can also allow detection of the chromosome/s or specific DNA sequence/s involved in a particular aberration formation. A variety of chromosome painting techniques are available in cytogenetics; here two highly sensitive methods, multiple fluorescence in situ hybridization (mFISH) and spectral karyotyping (SKY), are discussed to identify inter-chromosomal stable aberrations that form in the bone marrow cells of mice after exposure to total body irradiation. Although both techniques rely on fluorescent labeled DNA probes, the method of detection and the process of image acquisition of the fluorescent signals are different. These two techniques have been used in various research areas, such as radiation biology, cancer cytogenetics, retrospective radiation biodosimetry, clinical cytogenetics, evolutionary cytogenetics, and comparative cytogenetics.

  7. Double minute chromatin bodies and other chromosome alterations in human myeloid HL-60 leukemia cells susceptible or resistant to induction of differentiation by phorbol-12-myristate-13-acetate

    SciTech Connect

    Au, W.W.; Callaham, M.F.; Workman, M.L.; Huberman, E.

    1983-12-01

    An analysis of the chromosomal karyotype of the human promyelocytic HL-60 leukemia cell line and of a number of its sublines that exhibit varying degrees of resistance to induction of differentiation by phorbol-12-myristate-13-acetate was conducted. The HL-60 cell line and the derived sublines contained two consistent marker chromosomes (9p- and t(10;13)), which suggested that they have a common and possibly clonal origin. HL-60 cells that are susceptible to phorbol-12-myristate-13-acetate-induced cell differentiation contained double minute chromatine bodies. The sublines with different degrees of resistance showed a corresponding sequential reduction of double minute chromatin bodies in metaphase cells. This loss of double minute chromatin bodies was not associated with an appearance of homogeneously staining chromosomal regions. Resistant and susceptible HL-60 cell differed also in a number of other chromosomal alteration, including gains or losses involving chromosomes 5, 8, 11, 13, 16, and 17. Thus, it is suggested that acquisition of resistance to phorbol-12-myristate-13-acetate-induced cell differentiation in the HL-60 cells may involve one or more of the above chromosomal changes.

  8. Y chromosome in Turner syndrome: detection of hidden mosaicism and the report of a rare X;Y translocation case.

    PubMed

    Bispo, Adriana Valéria Sales; Burégio-Frota, Pollyanna; Oliveira dos Santos, Luana; Leal, Gabriela Ferraz; Duarte, Andrea Rezende; Araújo, Jacqueline; Cavalcante da Silva, Vanessa; Muniz, Maria Tereza Cartaxo; Liehr, Thomas; Santos, Neide

    2014-10-01

    Turner syndrome (TS) is a common genetic disorder in females associated with the absence of complete or parts of a second sex chromosome. In 5-12% of patients, mosaicism for a cell line with a normal or structurally abnormal Y chromosome is identified. The presence of Y-chromosome material is of medical importance because it results in an increased risk of developing gonadal tumours and virilisation. Molecular study and fluorescence in situ hybridisation approaches were used to study 74 Brazilian TS patients in order to determine the frequency of hidden Y-chromosome mosaicism, and to infer the potential risk of developing malignancies. Additionally, we describe one TS girl with a very uncommon karyotype 46,X,der(X)t(X;Y)(p22.3?2;q11.23) comprising a partial monosomy of Xp22.3?2 together with a partial monosomy of Yq11.23. The presence of cryptic Y-chromosome-specific sequences was detected in 2.7% of the cases. All patients with Y-chromosome-positive sequences showed normal female genitalia with no signs of virilisation. Indeed, the clinical data from Y-chromosome-positive patients was very similar to those with Y-negative results. Therefore, we recommend that the search for hidden Y-chromosome mosaicism should be carried out in all TS cases and not be limited to virilised patients or carriers of a specific karyotype.

  9. Array-Based Comparative Genomic Hybridization for the Genomewide Detection of Submicroscopic Chromosomal Abnormalities

    PubMed Central

    Vissers, Lisenka E. L. M. ; de Vries, Bert B. A. ; Osoegawa, Kazutoyo ; Janssen, Irene M. ; Feuth, Ton ; Choy, Chik On ; Straatman, Huub ; van der Vliet, Walter ; Huys, Erik H. L. P. G. ; van Rijk, Anke ; Smeets, Dominique ; van Ravenswaaij-Arts, Conny M. A. ; Knoers, Nine V. ; van der Burgt, Ineke ; de Jong, Pieter J. ; Brunner, Han G. ; van Kessel, Ad Geurts ; Schoenmakers, Eric F. P. M. ; Veltman, Joris A. 

    2003-01-01

    Microdeletions and microduplications, not visible by routine chromosome analysis, are a major cause of human malformation and mental retardation. Novel high-resolution, whole-genome technologies can improve the diagnostic detection rate of these small chromosomal abnormalities. Array-based comparative genomic hybridization allows such a high-resolution screening by hybridizing differentially labeled test and reference DNAs to arrays consisting of thousands of genomic clones. In this study, we tested the diagnostic capacity of this technology using ∼3,500 flourescent in situ hybridization–verified clones selected to cover the genome with an average of 1 clone per megabase (Mb). The sensitivity and specificity of the technology were tested in normal-versus-normal control experiments and through the screening of patients with known microdeletion syndromes. Subsequently, a series of 20 cytogenetically normal patients with mental retardation and dysmorphisms suggestive of a chromosomal abnormality were analyzed. In this series, three microdeletions and two microduplications were identified and validated. Two of these genomic changes were identified also in one of the parents, indicating that these are large-scale genomic polymorphisms. Deletions and duplications as small as 1 Mb could be reliably detected by our approach. The percentage of false-positive results was reduced to a minimum by use of a dye-swap-replicate analysis, all but eliminating the need for laborious validation experiments and facilitating implementation in a routine diagnostic setting. This high-resolution assay will facilitate the identification of novel genes involved in human mental retardation and/or malformation syndromes and will provide insight into the flexibility and plasticity of the human genome. PMID:14628292

  10. A simplified method to detect epididymal sperm aneuploidy (ESA) in mice using three-chromosome fish

    SciTech Connect

    Lowe, X.; O`Hogan, S.; Wyrobek, A.

    1995-11-01

    We developed a new method (ESA) to detect aneuploidy and polyploidy in epididymal sperm of mice using three-chromosome FISH. In comparison to a previous method (TSA-testicular spermatid aneuploidy), which required late-step spermatids, the ESA method utilizes epididymal sperm, which are easier to collect than testicular cells. The ESA method also provides a homogenous population of cells, which significantly speeds up the scoring procedure. A total of 6 mice were investigated by the ESA method and results compared with those obtained by the TSA method: 2 mice each of Robertsonian (8.14) heterozygotes, Rb(8.14) homozygotes and B6C3F1. About 10,000 sperm were scored per mouse. For the ESA method, epididimides were cut into small pieces and filtered. Sperm were prepared for hybridization by sonication and a modification of the DTT/LIS method previously described. Sperm aneuploidy was detected by multi-color FISH using three DNA probes specific for mouse chromosomes X, Y and 8. The sex ratio of X8(49.7%) and Y8(49.6%) did not differ from the expected 1:1. The efficiency of ESA was very high; -0.3% of the cells showed no hybridization domain. Hyperhaploidy frequencies for chromosomes X, Y and 8 compared well between the ESA and TSA methods for Rb(8.14) heterozygous (p=0.79) and B6C3F1 mice (p>0.05). The data obtained from Rb(8.14) homozygotes were similar to those from B6C3F1, as predicted (p=0.3). This highly efficient ESA assay is therefore, recommended for future studies of the mechanism of induction of aneuploidy in male germ cells. It also lays a solid foundation for automated scoring.

  11. Detection of Chromosome Aneuploidies in Chorionic Villus Samples by Multiplex Ligation-Dependent Probe Amplification

    PubMed Central

    Kooper, Angelique J.A.; Faas, Brigitte H.W.; Feuth, Ton; Creemers, Johan W.T.; Zondervan, Hans H.; Boekkooi, Peter F.; Quartero, Rik W.P.; Rijnders, Robbert J.P.; van der Burgt, Ineke; van Kessel, Ad Geurts; Smits, Arie P.T.

    2009-01-01

    The objective of this study was to examine the suitability of multiplex ligation-dependent probe amplification (MLPA) in chorionic villus samples as a replacement for traditional karyotyping for the detection of (an)euploidies of chromosomes 21, 18, 13, X, and Y. Chorionic villus samples were diagnosed by traditional karyotyping using short-term cultures (STC) and long-term cultures (LTC), and by MLPA using kit P095. DNA was extracted after digestion of whole villi with proteinase K and/or trypsin and collagenase. Different cell-dissociation procedures were tested to obtain MLPA results representative of the cytotrophoblast layer and the mesenchymal core. Over 95% of the MLPA results were in concordance with the traditional karyotyping of STC and LTC. Traditional karyotyping revealed seven mosaics. After digestion of whole villi with proteinase K, only abnormal cell lines confined to the STC gave rise to abnormal MLPA results. In one sample, the complete discrepancy between STC and LTC was resolved after enzymatic dissociation of cells from the cytotrophoblast layer and the mesenchymal core. MLPA in chorionic villus samples was found to be a reliable test for the detection of (an)euploidies of chromosomes 21, 18, 13, X, and Y. Whole villi digestion with proteinase K resulted in the over-representation of cytotrophoblasts in the DNA pool. To obtain MLPA results representative for STC and LTC, enzymatic dissociation of cells from the cytotrophoblast layer and mesenchymal core is required. PMID:19074591

  12. Detection of imprinting effects for qualitative traits on X chromosome based on nuclear families.

    PubMed

    Zhou, Ji-Yuan; You, Xiao-Ping; Yang, Ran; Fung, Wing Kam

    2016-01-01

    Methods for detecting imprinting effects have been developed primarily for autosomal markers. However, no method is available in the literature to test for imprinting effects on X chromosome. Therefore, it is necessary to suggest methods for detecting such imprinting effects. In this article, the parental-asymmetry test on X chromosome (XPAT) is first developed to test for imprinting for qualitative traits in the presence of association, based on family trios each with both parents and their affected daughter. Then, we propose 1-XPAT to deal with parent-daughter pairs, each with one parent and his/her affected daughter. By simultaneously considering family trios and parent-daughter pairs, C-XPAT (the combined test statistic of XPAT and 1-XPAT) is constructed to test for imprinting. Further, we extend the proposed methods to accommodate complete (with both parents) and incomplete (with one parent) nuclear families having multiple daughters of which at least one is affected. Simulation results demonstrate that the proposed methods control the size well, irrespective of the inbreeding coefficient in females being zero or non-zero. By incorporating incomplete nuclear families, C-XPAT is more powerful than XPAT using only complete nuclear families. For practical use, these proposed methods are applied to analyse the rheumatoid arthritis data and Turner's syndrome data.

  13. Baseline characteristics, chromosomal alterations, and treatment affecting prognosis of deletion 17p in newly diagnosed myeloma.

    PubMed

    Merz, Maximilian; Hielscher, Thomas; Seckinger, Anja; Hose, Dirk; Mai, Elias K; Raab, Marc S; Goldschmidt, Hartmut; Jauch, Anna; Hillengass, Jens

    2016-11-01

    Deletion 17p13, del(17p), is associated with poor outcome in myeloma but some patients show long-term survival. With the current study we intended to identify factors impacting outcome of such high risk patients. We analyzed 110 newly diagnosed, symptomatic patients with del(17p) detected by fluorescence in situ hybridization (FISH) in CD138-purified myeloma cells to identify prognostic factors for survival. Age >65 years, ISS III, and elevated LDH negatively impacted survival. Patients with subclonal (10-60% of plasma cells) del(17p) had longer progression-free survival (PFS) than patients with del(17p) in >60% of plasma cells (26 vs. 19 months, P = 0.03). Additional gain of 1q21 was associated with shorter PFS (17 vs. 25 months, P = 0.01). Hyperdiploidy did not ameliorate impact of del(17p), but gain 19q13 predicted longer PFS (30 vs. 18 months, P = 0.01) and overall survival (50 vs. 29 months, P = 0.01). Multivariate analysis in transplant eligible patients (≤65 years) revealed better survival for patients treated with upfront autologous transplantation (hazard ratio, [95% confidence interval]: 0.15 [0.04, 0.58], P = 0.006). Application of maintenance therapy was associated with better survival in transplant-eligible patients (0.30 [0.09, 0.99], P = 0.05). We demonstrate heterogeneous outcome of patients with del(17p) according to baseline characteristics and treatment. 19q13 should be included in routine FISH panel, since gains were associated with better survival. Am. J. Hematol. 91:E473-E477, 2016. © 2016 Wiley Periodicals, Inc.

  14. Angelman syndrome: Validation of molecular cytogenetic analysis of chromosome 15q11-q13 for deletion detection

    SciTech Connect

    White, L.; Knoll, J.H.M.

    1995-03-13

    In a series of 18 individuals comprising parents of Angelman syndrome (AS) patients and AS patients with large deletions, microdeletions, and no deletions, we utilized fluorescence in situ hybridization (FISH) with genomic phage clones for loci D15S63 and GABRB3 for deletion detection of chromosome 15q11-q13. Utilization of probes at these loci allows detection of common large deletions and permits discrimination of less common small deletions. In all individuals the molecular cytogenetic data were concordant with the DNA deletion analyses. FISH provides an accurate method of deletion detection for chromosome 15q11-q13. 23 refs., 2 figs., 1 tab.

  15. Somatic Instability of a Drosophila Chromosome

    PubMed Central

    Wines, D. R.; Henikoff, S.

    1992-01-01

    A mitotically unstable chromosome, detectable because of mosaic expression of marker genes, was generated by X-ray mutagenesis in Drosophila. Nondisjunction of this chromosome is evident in mitotic chromosome preparations, and premature sister chromatid separation is frequent. The mosaic phenotype is modified by genetic elements that are thought to alter chromatin structure. We hypothesize that the mitotic defects result from a breakpoint deep in the pericentric heterochromatin, within or very near to the DNA sequences essential for centromere function. This unique chromosome may provide a tool for the genetic and molecular dissection of a higher eukaryotic centromere. PMID:1628811

  16. Switching the centromeres on and off: epigenetic chromatin alterations provide plasticity in centromere activity stabilizing aberrant dicentric chromosomes.

    PubMed

    Sato, Hiroshi; Saitoh, Shigeaki

    2013-12-01

    The kinetochore, which forms on a specific chromosomal locus called the centromere, mediates interactions between the chromosome and the spindle during mitosis and meiosis. Abnormal chromosome rearrangements and/or neocentromere formation can cause the presence of multiple centromeres on a single chromosome, which results in chromosome breakage or cell cycle arrest. Analyses of artificial dicentric chromosomes suggested that the activity of the centromere is regulated epigenetically; on some stably maintained dicentric chromosomes, one of the centromeres no longer functions as a platform for kinetochore formation, although the DNA sequence remains intact. Such epigenetic centromere inactivation occurs in cells of various eukaryotes harbouring 'regional centromeres', such as those of maize, fission yeast and humans, suggesting that the position of the active centromere is determined by epigenetic markers on a chromosome rather than the nucleotide sequence. Our recent findings in fission yeast revealed that epigenetic centromere inactivation consists of two steps: disassembly of the kinetochore initiates inactivation and subsequent heterochromatinization prevents revival of the inactivated centromere. Kinetochore disassembly followed by heterochromatinization is also observed in normal senescent human cells. Thus epigenetic centromere inactivation may not only stabilize abnormally generated dicentric chromosomes, but also be part of an intrinsic mechanism regulating cell proliferation.

  17. A DNA fragment from the human X chromosome short arm which detects a partially homologous sequence on the Y chromosomes long arm.

    PubMed Central

    Koenig, M; Camerino, G; Heilig, R; Mandel, J L

    1984-01-01

    An X linked human DNA fragment (named DXS31 ) which detects partially homologous sequences on the Y chromosome has been isolated. Regional localisation of the two sex linked sequences was determined using a panel of rodent-human somatic cell hybrids. The X specific sequence is located at the tip of the short arm ( Xp22 .3-pter), i.e. within or close to the region which pairs with the Y chromosome short arm at meiosis. However the Y specific sequence is located in the heterochromatic region of the long arm ( Yq11 -qter) and lies outside from the pairing region. DNAs from several XX male subjects were probed with DXS31 and in all cases a double dose of the X linked fragment was found, and the Y specific fragment was absent. DXS31 detects in chimpanzee a male-female differential pattern identical to that found in man. However results obtained in a more distantly related species, the brown lemur, suggest that the sequences detected by DXS31 in this species might be autosomally coded. The features observed with these X-Y related sequences do not fit with that expected from current hypotheses of homology between the pairing regions of the two sex chromosomes, nor with the pattern observed with other X-Y homologous sequences recently characterized. Our results suggest also that the rule of conservation of X linkage in mammals might not apply to sequences present on the tip of the X chromosome short arm, in bearing with the controversial issue of steroid sulfatase localisation in mouse. Images PMID:6728677

  18. Automated identification of abnormal metaphase chromosome cells for the detection of chronic myeloid leukemia using microscopic images

    NASA Astrophysics Data System (ADS)

    Wang, Xingwei; Zheng, Bin; Li, Shibo; Mulvihill, John J.; Chen, Xiaodong; Liu, Hong

    2010-07-01

    Karyotyping is an important process to classify chromosomes into standard classes and the results are routinely used by the clinicians to diagnose cancers and genetic diseases. However, visual karyotyping using microscopic images is time-consuming and tedious, which reduces the diagnostic efficiency and accuracy. Although many efforts have been made to develop computerized schemes for automated karyotyping, no schemes can get be performed without substantial human intervention. Instead of developing a method to classify all chromosome classes, we develop an automatic scheme to detect abnormal metaphase cells by identifying a specific class of chromosomes (class 22) and prescreen for suspicious chronic myeloid leukemia (CML). The scheme includes three steps: (1) iteratively segment randomly distributed individual chromosomes, (2) process segmented chromosomes and compute image features to identify the candidates, and (3) apply an adaptive matching template to identify chromosomes of class 22. An image data set of 451 metaphase cells extracted from bone marrow specimens of 30 positive and 30 negative cases for CML is selected to test the scheme's performance. The overall case-based classification accuracy is 93.3% (100% sensitivity and 86.7% specificity). The results demonstrate the feasibility of applying an automated scheme to detect or prescreen the suspicious cancer cases.

  19. Next-generation sequencing strategies enable routine detection of balanced chromosome rearrangements for clinical diagnostics and genetic research.

    PubMed

    Talkowski, Michael E; Ernst, Carl; Heilbut, Adrian; Chiang, Colby; Hanscom, Carrie; Lindgren, Amelia; Kirby, Andrew; Liu, Shangtao; Muddukrishna, Bhavana; Ohsumi, Toshiro K; Shen, Yiping; Borowsky, Mark; Daly, Mark J; Morton, Cynthia C; Gusella, James F

    2011-04-08

    The contribution of balanced chromosomal rearrangements to complex disorders remains unclear because they are not detected routinely by genome-wide microarrays and clinical localization is imprecise. Failure to consider these events bypasses a potentially powerful complement to single nucleotide polymorphism and copy-number association approaches to complex disorders, where much of the heritability remains unexplained. To capitalize on this genetic resource, we have applied optimized sequencing and analysis strategies to test whether these potentially high-impact variants can be mapped at reasonable cost and throughput. By using a whole-genome multiplexing strategy, rearrangement breakpoints could be delineated at a fraction of the cost of standard sequencing. For rearrangements already mapped regionally by karyotyping and fluorescence in situ hybridization, a targeted approach enabled capture and sequencing of multiple breakpoints simultaneously. Importantly, this strategy permitted capture and unique alignment of up to 97% of repeat-masked sequences in the targeted regions. Genome-wide analyses estimate that only 3.7% of bases should be routinely omitted from genomic DNA capture experiments. Illustrating the power of these approaches, the rearrangement breakpoints were rapidly defined to base pair resolution and revealed unexpected sequence complexity, such as co-occurrence of inversion and translocation as an underlying feature of karyotypically balanced alterations. These findings have implications ranging from genome annotation to de novo assemblies and could enable sequencing screens for structural variations at a cost comparable to that of microarrays in standard clinical practice.

  20. Activation of the c-abl oncogene by viral transduction or chromosomal translocation generates altered c-abl proteins with similar in vitro kinase properties.

    PubMed Central

    Davis, R L; Konopka, J B; Witte, O N

    1985-01-01

    The v-abl protein of Abelson murine leukemia virus is a tyrosine-specific kinase. Its normal cellular homolog, murine c-abl, does not possess detectable tyrosine kinase activity in vitro. Previously, we have detected tyrosine kinase activity in vitro for an altered c-abl gene product (c-abl P210) in the K562 human chronic myelogenous leukemia cell line. The expression of this variant c-abl gene product correlates with chromosomal translocation and amplification of the c-abl gene in K562 cells. Like v-abl, c-abl P210 is a fusion protein containing non-abl sequences near the amino terminus of c-abl. We compared the in vitro tyrosine kinase activity of c-abl P210 with that of wild-type murine v-abl. The remarkable similarities of these two proteins with respect to cis-acting autophosphorylation, trans-acting phosphorylation of exogenous substrates, and kinase inhibition, using site-directed abl-specific antisera, suggested that c-abl P210 could function similarly to v-abl in vivo. In addition, c-abl P210 possessed an associated serine kinase activity in immunoprecipitates. The serine kinase activity was not inhibited by site-directed, abl-specific antisera that inhibit the tyrosine kinase activity, suggesting that the serine kinase activity is not an intrinsic property of c-abl P210. Thus, the activation of the c-abl gene in a human leukemia cell line may have functional consequences analogous to activation of the c-abl gene in Abelson murine leukemia virus. Images PMID:4039028

  1. Detection of a subtle rearrangement of chromosome 22 using molecular techniques

    SciTech Connect

    Biesecker, L.G.; Rosenberg, M.; Dziadzio, L.

    1995-09-25

    Conventional cytogenetics is a useful clinical tool that has a lower limit of sensitivity of 2-5 Mb for detection of duplications or deletions. Because the threshold of clinically significant aneusomy is below this range, there is a need for approaches to improve the sensitivity of the detection of aneusomy. We have implemented a system of screening for subtle unbalanced translocations in children with multiple congenital anomalies of unknown cause. Our approach uses subtelomeric microsatellite markers to detect small areas of segmental aneusomy due to unbalanced translocations. Herein we report a patient with severe multiple congenital anomalies and a normal karyotype who was diagnosed by this approach. Microsatellite markers from 41 telomeres were analyzed and were normal with the exception of those on distal chromosome 22. Further analysis with additional microsatellites and fluorescent in situ hybridization confirmed duplication of 22q13.2-qter. We conclude that microsatellite screening can detect subtle unbalanced translocations in children with severe anomalies. 15 refs., 3 figs., 1 tab.

  2. Microsatellite DNA markers detects 95% of chromosome 22q11 deletions.

    PubMed

    Bonnet, D; Cormier-Daire, V; Kachaner, J; Szezepanski, I; Souillard, P; Sidi, D; Munnich, A; Lyonnet, S

    1997-01-20

    Cono-truncal cardiac malformations account for some 50% of congenital heart defects in newborn infants. Recently, hemizygosity for chromosome 22q11.2 was reported in patients with the DiGeorge/Velo-cardio-facial syndromes (DGS/VCFS) and causally related disorders. We have explored the potential use of microsatellite DNA markers for rapid detection of 22q11 deletions in 19 newborn infants referred for cono-truncal heart malformations with associated DGS/VCFS anomalies. A failure of parental inheritance was documented in 84.2% of cases (16/19). PCR-based genotyping using microsatellite DNA markers located within the commonly deleted region allowed us either to confirm or reject a 22q11 microdeletion in 94.3% of cases (18/19) within 24 hours. This test is now currently performed in the infants referred to us for a cono-truncal heart malformation as a first intention screening for 22q11 microdeletion.

  3. Rapid detection of chromosome 18 copy number in buccal smears using DNA probes and FISH

    SciTech Connect

    Harris, C.; Nunez, M.; Giraldez, R.

    1994-09-01

    Rapid diagnosis of trisomy 18 in newborns is often critical to clinical management decisions that must be made in a minimum of time. DNA probes combined with FISH can be used to accurately to determine the copy number of chromosome 18 in interphase cells. We have used the D18Z1 alpha satellite DNA probe to determine signal frequency in normal, previously karyotyped subjects, 12 females and 6 males. We also present one clinical case of trisomy 18, confirmed by karyotype, for comparison to the results obtained from normal subjects. Buccal smears, unlike cytogenetic preparations from peripheral blood, are quite resistant to penetration of probes and detection reagents resulting in higher levels of false monosomy. We have studied 19 individuals and have obtained consistent FISH results, ranging from 64 to 90% disomy. False monosomy rates ranged from 10 to 36%, while false trisomy or tetrasomy was less than 1% in all samples. High rates of false monosomy make this test questionable for detection of low order mosaicism for monosomy, but the extremely low false hyperploidy rate suggests that this is a dependable procedure for detection of trisomy 18, enabling the use of buccal epithelium which can be collected easily from even premature and tiny infants.

  4. A chromosomally based luminescent bioassay for mercury detection in red soil of China.

    PubMed

    Wei, He; Cheng, Han; Ting, Mao; Wen-Hui, Zhong; Xian-Gui, Lin

    2010-07-01

    A luminescent reporter gene system was constructed by fusing the mercury-inducible promoter, P ( merT ), and its regulatory gene, merR, with a promoterless reporter gene EGFP. A stable and nonantibiotic whole-cell reporter (BMB-ME) was created by introducing the system cassette into the chromosome of Pseudomonas putida strain and then applied it for mercury detection in the red soil of China. Spiked with 10 and 100 microg g(-1) Hg(2+) and after 15 and 30 days incubation, soil samples were extracted and evaluated water soluble, bioavailable, organic matter bound, and residual fractions of mercury by both BMB-ME and chemical way. The expression of EGFP was confirmed in soil extraction, and fluorescence intensity was quantified by luminescence spectrometer. The sensor strain BMB-ME appeared to have a detection range similar to that of reversed-phase high-performance liquid chromatography method. The optimal temperature for EGFP expression was 35 degrees C and the lowest detectable concentration of Hg(2+) 200 nM. Cu(2+), Fe(2+), Mn(2+), Sn(2+), Zn(2+), Co(2+), Ag(+), Ba(2+), Mg(2+), and Pb(2+) ions at nanomolar level did not interfere with the measurement. These results showed that the BMB-ME constitute an adaptable system for easy sensing of small amounts of mercury in the red soil of China.

  5. Array CGH detection of a novel cryptic deletion at 3q13 in a complex chromosome rearrangement.

    PubMed

    López-Expósito, Isabel; Ballesta-Martinez, María Juliana; Bafalliu, Juan Antonio; Vera-Carbonell, Ascensión; Domingo-Jiménez, Rosario; López-González, Vanesa; Fernández, Asunción; Guillén-Navarro, Encarna

    2014-04-01

    Complex chromosome rearrangements (CCRs) are extremely rare in humans. About 20% of the apparently balanced CCRs have an abnormal phenotype and the degree of severity correlates with a higher number of breakpoints. Several studies using FISH and microarray technologies have shown that deletions in the breakpoints are common although duplications, insertions and inversions have also been detected. We report a patient with two simultaneous reciprocal translocations, t(3;4) and t(2;14;18), involving five chromosomes and six breakpoints. He showed dysmorphic features, preaxial polydactyly in the left hand, brachydactyly, postnatal growth retardation and developmental delay. The rearrangement was characterized by FISH analysis which detected an interstitial segment from chromosome 14 inserted in the derivative chromosome 2, and by whole genome array which revealed an interstitial deletion of approximately 4.5 Mb at the breakpoint site on chromosome 3. To our knowledge this microdeletion has not been previously reported and includes ~12 genes. The haploinsufficiency of one or several of these genes is likely to have contributed to the clinical phenotype of the patient. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Chromosome 15q11-13 duplication syndrome brain reveals epigenetic alterations in gene expression not predicted from copy number

    PubMed Central

    Hogart, Amber; Leung, Karen N.; Wang, Nicholas J.; Wu, David J.; Driscoll, Jennette; Vallero, Roxanne O.; Schanen, N. Carolyn

    2008-01-01

    Background Chromosome 15q11-13 contains a cluster of imprinted genes essential for normal mammalian neurodevelopment. Deficiencies in paternal or maternal 15q11-13 alleles result in Prader-Willi or Angelman syndromes, respectively, and maternal duplications lead to a distinct condition that often includes autism. Overexpression of maternally expressed imprinted genes is predicted to cause 15q11-13-associated autism, but a link between gene dosage and expression has not been experimentally determined in brain. Methods Post-mortem brain tissue was obtained from a male with 15q11-13 hexasomy and a female with 15q11-13 tetrasomy. Quantitative RT-PCR was used to measure ten 15q11-13 transcripts in maternal 15q11-13 duplication, Prader-Willi syndrome, and control brain samples. Southern blot, bisulfite sequencing and fluorescence in situ hybridization were used to investigate epigenetic mechanisms of gene regulation. Results Gene expression and DNA methylation correlated with parental gene dosage in the male 15q11-13 duplication sample with severe cognitive impairment and seizures. Strikingly, the female with autism and milder Prader-Willi-like characteristics demonstrated unexpected deficiencies in the paternally expressed transcripts SNRPN, NDN, HBII85, and HBII52 and unchanged levels of maternally expressed UBE3A compared to controls. Paternal expression abnormalities in the female duplication sample were consistent with elevated DNA methylation of the 15q11-13 imprinting control region (ICR). Expression of nonimprinted 15q11-13 GABA receptor subunit genes was significantly reduced specifically in the female 15q11-13 duplication brain without detectable GABRB3 methylation differences. Conclusion Our findings suggest that genetic copy number changes combined with additional genetic or environmental influences on epigenetic mechanisms impact outcome and clinical heterogeneity of 15q11-13 duplication syndromes. PMID:18835857

  7. Chromosomal aneusomy in sputum, as detected by Fluorescence In Situ Hybridization (FISH) predicts lung cancer incidence

    PubMed Central

    Varella-Garcia, Marileila; Schulte, Aline P.; Wolf, Holly J.; Feser, William J; Zeng, Chan; Braudrick, Sarah; Yin, Xiang; Hirsch, Fred R.; Kennedy, Timothy C.; Keith, Robert L.; Barón, Anna E.; Belinsky, Steven A.; Miller, York E.; Byers, Tim; Franklin, Wilbur A.

    2009-01-01

    Lung cancer is usually disseminated at diagnosis making prognosis poor. Smokers are at high risk for lung cancer and are targets for prevention and early detection strategies. Sputum is a potential source for lung cancer biomarkers, but no test is currently available with sufficient sensitivity and specificity for clinical screening utility. Chromosomal aneusomy (CA) was measured in sputum samples collected prospectively from 100 incident lung cancer cases and 96 controls matched on age, gender, and date of collection. The CA-FISH assay was performed using a four-target DNA FISH probe including EGFR, MYC, 5p15 and CEP6. Sensitivity for a positive CA-FISH assay (abnormal for ≥ 2 of the 4 markers) was substantially higher for samples collected within 18 months (76%) than >18 months before lung cancer diagnosis (31%). Specificity for a positive FISH by this same definition was 85%. Among subjects providing sputum sample within 18 months before diagnosis, sensitivity was higher for squamous cell cancers (94%) than for other histologic types (69%). The adjusted odds ratios for specimens collected within 18 months of cancer diagnosis were higher using the CA-FISH assay (OR=27.2, 95% CI 7.8 to 94.1) than previous studies assessing cytologic atypia (OR=2.3, CI 0.8 to 6.4) or gene promoter methylation (OR=6.5; CI 1.2 to 35.5). In conclusion, chromosomal aneusomy in sputum is a promising biomarker for prediction of lung cancer risk. Evaluation of the 4-DNA targets was more effective than any single marker and had highest sensitivity for samples collected ≤ 18 months to lung cancer diagnosis and patients diagnosed with squamous cell carcinoma. PMID:20332298

  8. Leukemia-related chromosomal loss detected in hematopoietic progenitor cells of benzene-exposed workers

    PubMed Central

    Zhang, Luoping; Lan, Qing; Ji, Zhiying; Li, Guilan; Shen, Min; Vermeulen, Roel; Guo, Weihong; Hubbard, Alan E.; McHale, Cliona M.; Rappaport, Stephen M.; Hayes, Richard B.; Linet, Martha S.; Yin, Songnian; Smith, Martyn T.; Rothman, Nathaniel

    2012-01-01

    Benzene exposure causes acute myeloid leukemia, and hematotoxicity, shown as suppression of mature blood and myeloid progenitor cell numbers. As the leukemia-related aneuploidies monosomy 7 and trisomy 8 previously had been detected in the mature peripheral blood cells of exposed workers, we hypothesized that benzene could cause leukemia through the induction of these aneuploidies in hematopoietic stem and progenitor cells. We measured loss and gain of chromosomes 7 and 8 by fluorescence in situ hybridization in interphase colony-forming unit-granulocyte-macrophage (CFU-GM) cells cultured from otherwise healthy benzene-exposed (n=28) and unexposed (n=14) workers. CFU-GM monosomy 7 and 8 levels (but not trisomy) were significantly increased in subjects exposed to benzene overall, compared to levels in the control subjects (p=0.0055 and p=0.0034, respectively). Levels of monosomy 7 and 8 were significantly increased in subjects exposed to <10 ppm (20%, p=0.0419 and 28%, p=0.0056, respectively) and ≥10 ppm (48%, p=0.0045 and 32%, p=0.0354) benzene, compared with controls, and significant exposure-response trends were detected (ptrend=0.0033 and 0.0057). These data show that monosomies 7 and 8 are produced in a dose-dependent fashion in the blood progenitor cells of workers exposed to benzene and may be mechanistically relevant biomarkers of early effect for benzene and other leukemogens. PMID:22643707

  9. Detection of Selection Signatures on the X Chromosome in Three Sheep Breeds.

    PubMed

    Zhu, Caiye; Fan, Hongying; Yuan, Zehu; Hu, Shijin; Zhang, Li; Wei, Caihong; Zhang, Qin; Zhao, Fuping; Du, Lixin

    2015-08-28

    Artificial selection has played a critical role in animal breeding. Detection of artificial selection footprints in genomic regions can provide insights for understanding the function of specific phenotypic traits and better guide animal breeding. To more fully understand the relationship between genomic composition and phenotypic diversity arising from breed development, a genome-wide scan was conducted using an OvineSNP50 BeadChip and integrated haplotype score and fixation index analyses to detect selection signatures on the X chromosome in three sheep breeds. We identified 49, 34, and 55 candidate selection regions with lengths of 27.49, 16.47, and 25.42 Mb in German Mutton, Dorper, and Sunit sheep, respectively. Bioinformatics analysis showed that some of the genes in these regions with selection signatures, such as BMP15, were relevant to reproduction. We also identified some selection regions harboring genes that had human orthologs, including BKT, CENPI, GUCY2F, MSN, PCDH11X, PLP1, VSIG4, PAK3, WAS, PCDH19, PDHA1, and SRPX2. The VSIG4 and PCDH11X genes are associated with the immune system and disease, PDHA1 is associated with biosynthetic related pathways, and PCDH19 is expressed in the nervous system and skin. These genes may be useful as candidate genes for molecular breeding.

  10. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH)

    PubMed Central

    Cortés-Gutiérrez, Elva I.; Ortíz-Hernández, Brenda L.; Dávila-Rodríguez, Martha I.; Cerda-Flores, Ricardo M; Fernández, José Luis; López-Fernández, Carmen; Gosálvez, Jaime

    2013-01-01

    We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH). A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL), 10 with high-grade SIL (HG-SIL), and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1. PMID:23429197

  11. 5-bp Classical Satellite DNA Loci from Chromosome-1 Instability in Cervical Neoplasia Detected by DNA Breakage Detection/Fluorescence in Situ Hybridization (DBD-FISH).

    PubMed

    Cortés-Gutiérrez, Elva I; Ortíz-Hernández, Brenda L; Dávila-Rodríguez, Martha I; Cerda-Flores, Ricardo M; Fernández, José Luis; López-Fernández, Carmen; Gosálvez, Jaime

    2013-02-19

    We aimed to evaluate the association between the progressive stages of cervical neoplasia and DNA damage in 5-bp classical satellite DNA sequences from chromosome-1 in cervical epithelium and in peripheral blood lymphocytes using DNA breakage detection/fluorescence in situ hybridization (DBD-FISH). A hospital-based unmatched case-control study was conducted in 2011 with a sample of 30 women grouped according to disease stage and selected according to histological diagnosis; 10 with low-grade squamous intraepithelial lesions (LG-SIL), 10 with high-grade SIL (HG-SIL), and 10 with no cervical lesions, from the Unidad Medica de Alta Especialidad of The Mexican Social Security Institute, IMSS, Mexico. Specific chromosome damage levels in 5-bp classical satellite DNA sequences from chromosome-1 were evaluated in cervical epithelium and peripheral blood lymphocytes using the DBD-FISH technique. Whole-genome DNA hybridization was used as a reference for the level of damage. Results of Kruskal-Wallis test showed a significant increase according to neoplastic development in both tissues. The instability of 5-bp classical satellite DNA sequences from chromosome-1 was evidenced using chromosome-orientation FISH. In conclusion, we suggest that the progression to malignant transformation involves an increase in the instability of 5-bp classical satellite DNA sequences from chromosome-1.

  12. Chromosome 17 aneusomy detected by fluorescence in situ hybridization in vulvar squamous cell carcinomas and synchronous vulvar skin.

    PubMed

    Carlson, J A; Healy, K; Tran, T A; Malfetano, J; Wilson, V L; Rohwedder, A; Ross, J S

    2000-09-01

    samples and was significantly associated with women with SCC in situ (HPV-related). Both DNA content and Ki-67 labeling positively and significantly correlated with mean chromosome 17 copy number (r = 0.1, P: = 0.007). A high degree of genetic instability (aneuploidy) occurs in the skin surrounding vulvar carcinomas. As these events could be detected in histologically normal skin and inflammatory lesions (lichen sclerosus), chromosomal abnormalities may be a driving force in the early stages of carcinogenesis. Differences in chromosomal patterns (loss or gain) support the concept of at least two pathways in vulvar carcinogenesis.

  13. Live birth after PGD with confirmation by a comprehensive approach (karyomapping) for simultaneous detection of monogenic and chromosomal disorders.

    PubMed

    Natesan, Senthilkumar A; Handyside, Alan H; Thornhill, Alan R; Ottolini, Christian S; Sage, Karen; Summers, Michael C; Konstantinidis, Michalis; Wells, Dagan; Griffin, Darren K

    2014-11-01

    Preimplantation genetic diagnosis (PGD) for monogenic disorders has the drawback of time and cost associated with tailoring a specific test for each couple, disorder, or both. The inability of any single assay to detect the monogenic disorder in question and simultaneously the chromosomal complement of the embryo also limits its application as separate tests may need to be carried out on the amplified material. The first clinical use of a novel approach ('karyomapping') was designed to circumvent this problem. In this example, karyomapping was used to confirm the results of an existing PGD case detecting both chromosomal abnormalities and a monogenic disorder (Smith-Lemli-Opitz [SLO] syndrome) simultaneously. The family underwent IVF, ICSI and PGD, and both polar body and cleavage stage biopsy were carried out. Following whole genome amplification, array comparative genomic hybridisation of the polar bodies and minisequencing and STR analysis of single blastomeres were used to diagnose maternal aneuploidies and SLO status, respectively. This was confirmed, by karyomapping. Unlike standard PGD, karyomapping required no a-priori test development. A singleton pregnancy and live birth, unaffected with SLO syndrome and with no chromosome abnormality, ensued. Karyomapping is potentially capable of detecting a wide spectrum of monogenic and chromosome disorders and, in this context, can be considered a comprehensive approach to PGD.

  14. Detecting Spatial Chromatin Organization by Chromosome Conformation Capture II: Genome-Wide Profiling by Hi-C.

    PubMed

    Vietri Rudan, Matteo; Hadjur, Suzana; Sexton, Tom

    2017-01-01

    The chromosome conformation capture (3C) method has been invaluable in studying chromatin interactions in a population of cells at a resolution surpassing that of light microscopy, for example in the detection of functional contacts between enhancers and promoters. Recent developments in sequencing-based chromosomal contact mapping (Hi-C, 5C and 4C-Seq) have allowed researchers to interrogate pairwise chromatin interactions on a wider scale, shedding light on the three-dimensional organization of chromosomes. These methods present significant technical and bioinformatic challenges to consider at the start of the project. Here, we describe two alternative methods for Hi-C, depending on the size of the genome, and discuss the major computational approaches to convert the raw sequencing data into meaningful models of how genomes are organized.

  15. Complex structural rearrangement of chromosomes 13, 19 and 20 detected cytogenetically and by fluorescence in situ hybridization (FISH)

    SciTech Connect

    Al-Nassar, K.E.; Murthy, S.K.; Verghese, L.

    1994-09-01

    Complex chromosomal rearrangements (CCRs) are rare. To date, 72 CCRs have been reported. We report here a case of CCR in a 10 year old boy and his mother involving chromosomes 13, 19 and 20, detected by G-,C- and Ag-NOR banding and by fluorescence in situ hybridization (FISH) technique. Father and other sibs were found to be chromosomally normal. The patient presented with clinical features having obesity, micropenis, slow learning and IQ=70. Mother was clinically normal. Karyotype of the patient and mother showed apparently balanced chromosomal rearrangements involving chromosomes 13, 19 and 20. The karyotypes were interpreted as: 45,XY,-19,der(13),der(20),t(13;19)(13p11.2)::(19q13.2{r_arrow}19pter),t(19;20)(19q13.3{r_arrow}19qter::20qter) in the patient and 45,XX,-19,t(13;19),t(19;20) involving the same breakpoints in the mother. C-banding showed dicentric der(13). FISH using alpha-satellite DNA probes for 13/21 showed the presence of centromeric region of 13 in the der(13). Deletion of 13p11.2{r_arrow}pter was confirmed by negative Ag-NOR staining in der(13).

  16. Prenatal detection and characterization of supernumerary marker chromosomes by array-CGH

    USDA-ARS?s Scientific Manuscript database

    Small supernumerary marker chromosomes (sSMC) occur in about 0.043% of newborns and in 0.076% of prenatal diagnoses. The phenotypes associated with sSMC vary substantially depending on size, gene content, and chromosome origin, which cannot easily be determined by karyotype or FISH analysis. There...

  17. Chromosomal imbalances exclusively detected in invasive front area are associated with poor outcome in laryngeal carcinomas from different anatomical sites.

    PubMed

    Ambrosio, Eliane Papa; Silveira, Cássia Gisele Terrassani; Drigo, Sandra Aparecida; Sacomano, Vivian de Souza; Molck, Miriam Coelho; Rocha, Rafael Malagoli; Domingues, Maria Aparecida Custódio; Soares, Fernando Augusto; Kowalski, Luiz Paulo; Rogatto, Silvia Regina

    2013-10-01

    Laryngeal squamous cell carcinoma (LSCC) is a malignant neoplasm exhibiting aggressive phenotype, high recurrence rate, and risk of developing second primary tumors. Current evidence suggests that cells in the invasive front of carcinomas have different molecular profiles compared to those in superficial areas. This study aimed to identify candidate genes in the invasive front and superficial cells from laryngeal carcinomas that would be useful as molecular markers. Invasive front and tumor surface cells of 32 LSCC were evaluated by high-resolution comparative genomic hybridization. Both CCND1 copy number gains and cyclin D1 protein expression were evaluated to confirm gains of 11q13.3. Losses of 3q26.2-q29 and 18q23 were confirmed by loss of heterozygosity analysis. The most frequent chromosomal alterations observed only in invasive front cells involved gains of 1p, 4q, and 9p and losses of 3p, 11p, 12p, 13q, 17q, 18p, 19q, 20q, 21q, and Xp. Gains of 11q13 were detected in both components from glottis and supraglottis but only in invasive front cells from transglottic tumors. Fluorescence in situ hybridization confirmed gains of CCND1/CPE11 in a subset of cases. In supraglottic tumors, cyclin D1 positivity was associated with distant metastasis (P = 0.0018) and with decreased disease-free survival (P = 0.042). Loss of heterozygosity at 3q26.2 and 18q23 were associated with lymph node involvement (P = 0.055) and worsened prognosis, respectively. In conclusion, this study revealed regions that could be targeted in the search for molecular markers in LSCC. Cyclin D1 may be useful as a prognostic marker in supraglottic tumors.

  18. Detection of alterations in human sperm using magnetic orientation techniques

    NASA Astrophysics Data System (ADS)

    Sakhnini, Lama; Dairi, Maheen; Manaa, Hacene

    2007-09-01

    In this study we report on magnetic orientation of human sperms. Samples were taken from 17 donors. Normal human sperms became oriented with their long axis perpendicular to the magnetic field ( 1 Tesla maximum). Total orientation was achieved with magnetic field at about one Tesla, while for abnormal sperms the magnetic behavior was different. The dependence of the measured degree of orientation on the intensity of the magnetic field was in good agreement with the theoretical equation for the magnetic orientation of diamagnetic substances. As a result for a numerical analysis based on the equation, the anisotropic diamagnetic susceptibility of normal sperm was found to be ▵ χ= 8×10 -20 J/T2. The degree of orientation was influenced by the alterations in the shape of the head, body or the tail. It has been suggested that the DNA in the sperm head retain the strong magnetic anisotropy to counter balance the magnetic anisotropy retained by flagellum microtubules. Recent studies demonstrated a well-defined nuclear architecture in human sperm nucleus, where the head morphology has significant correlation with sperm chromatin structure assay SCSA. Then as the methods to evaluate SCSA can be difficult and expensive our simple magnetic orientation technique can be an alternative to diagnose alteration in DNA.

  19. Association between occupational exposure to benzene and chromosomal alterations in lymphocytes of Brazilian petrochemical workers removed from exposure.

    PubMed

    Gonçalves, Rozana Oliveira; de Almeida Melo, Neli; Rêgo, Marco Antônio Vasconcelos

    2016-06-01

    We aimed to investigate the association between chronic exposure to benzene and genotoxicity in the lymphocytes of workers removed from exposure. The study included 20 workers with hematological disorders who had previously worked in the petrochemical industry of Salvador, Bahia, Brazil; 16 workers without occupational exposure to benzene served as the control group. Chromosomal analysis was performed on lymphocytes from peripheral blood, to assess chromosomal breaks and gaps and to identify aneuploidy. The Kruskal-Wallis test was used to compare the mean values between two groups, and Student's t test for comparison of two independent means. The frequency of gaps was statistically higher in and the exposed group than in the controls (2.13 ± 2.86 vs. 0.97 ± 1.27, p = 0.001). The frequency of chromosomal breaks was significantly higher among cases (0.21 ± 0.58) than among controls (0.12 ± 0.4) (p = 0.0002). An association was observed between chromosomal gaps and breaks and occupational exposure to benzene. Our study showed that even when removed from exposure for several years, workers still demonstrated genotoxic damage. Studies are still needed to clarify the long-term genotoxic potential of benzene after removal from exposure.

  20. Rapid screening for Native American mitochondrial and Y-chromosome haplogroups detection in routine DNA analysis.

    PubMed

    Zuccarelli, Gala; Alechine, Evguenia; Caputo, Mariela; Bobillo, Cecilia; Corach, Daniel; Sala, Andrea

    2011-03-01

    Aiming to detect individuals of Native American maternal or paternal ancestry a rapid screening approach has been developed. Its strategy was based on SNP typing by Real Time PCR (rt-PCR) followed by High Resolution Melting analysis (HRM). After extraction, DNA was quantitated by rt-PCR using commercial kits; samples were then submitted to two multiplex reactions in order to determine the major Native American mtDNA and Y-chromosome haplogroups by HRM. One cocktail included primers flanking nucleotide substitutions that define mtDNA haplogroup C and sub-haplogroups A2, B2, and D1. The other included primers flanking Y-SNPs M3, M269 and U179 that allowed discriminating Q and non-Q haplogroups. In all cases amplicons were <125 nucleotides long in order to increase the peak resolution. The accuracy of the results obtained was established by means of sequencing analysis of the amplicons. The new working-flow here proposed facilitates and speeds-up the screening process that may preclude a detailed sequencing analysis of particular samples, or for further molecular epidemiological investigations in which continental origin influences might be relevant.

  1. Detecting a hierarchical genetic population structure via Multi-InDel markers on the X chromosome

    PubMed Central

    Fan, Guang Yao; Ye, Yi; Hou, Yi Ping

    2016-01-01

    Detecting population structure and estimating individual biogeographical ancestry are very important in population genetics studies, biomedical research and forensics. Single-nucleotide polymorphism (SNP) has long been considered to be a primary ancestry-informative marker (AIM), but it is constrained by complex and time-consuming genotyping protocols. Following up on our previous study, we propose that a multi-insertion-deletion polymorphism (Multi-InDel) with multiple haplotypes can be useful in ancestry inference and hierarchical genetic population structures. A validation study for the X chromosome Multi-InDel marker (X-Multi-InDel) as a novel AIM was conducted. Genetic polymorphisms and genetic distances among three Chinese populations and 14 worldwide populations obtained from the 1000 Genomes database were analyzed. A Bayesian clustering method (STRUCTURE) was used to discern the continental origins of Europe, East Asia, and Africa. A minimal panel of ten X-Multi-InDels was verified to be sufficient to distinguish human ancestries from three major continental regions with nearly the same efficiency of the earlier panel with 21 insertion-deletion AIMs. Along with the development of more X-Multi-InDels, an approach using this novel marker has the potential for broad applicability as a cost-effective tool toward more accurate determinations of individual biogeographical ancestry and population stratification. PMID:27535707

  2. Microsatellite DNA markers detects 95% of chromosome 22q11 deletions

    SciTech Connect

    Bonnet, D.; Cormier-Daire, V.; Munnich, A.; Lyonnet, S.

    1997-01-20

    Cono-truncal cardiac malformations account for some 50% of congenital heart defects in newborn infants. Recently, hemizygosity for chromosome 22q11.2 was reported in patients with the DiGeorge/Velo-cardio-facial syndromes (DGS/VCFS) and causally related disorders. We have explored the potential use of microsatellite DNA markers for rapid detection of 22q11 deletions in 19 newborn infants referred for cono-truncal heart malformations with associated DGS/VCFS anomalies. A failure of parental inheritance was documented in 84.2% of cases (16/19). PCR-based genotyping using microsatellite DNA markers located within the commonly deleted region allowed us either to confirm or reject a 22q11 microdeletion in 94.3% of cases (18/19) within 24 hours. This test is now currently performed in the infants referred to us for a cono-truncal heart malformation as a first intention screening for 22q11 microdeletion. 10 refs., 1 fig., 1 tab.

  3. An Optimized Method for Accurate Fetal Sex Prediction and Sex Chromosome Aneuploidy Detection in Non-Invasive Prenatal Testing

    PubMed Central

    Li, Haibo; Ding, Jie; Wen, Ping; Zhang, Qin; Xiang, Jingjing; Li, Qiong; Xuan, Liming; Kong, Lingyin; Mao, Yan; Zhu, Yijun; Shen, Jingjing; Liang, Bo; Li, Hong

    2016-01-01

    Massively parallel sequencing (MPS) combined with bioinformatic analysis has been widely applied to detect fetal chromosomal aneuploidies such as trisomy 21, 18, 13 and sex chromosome aneuploidies (SCAs) by sequencing cell-free fetal DNA (cffDNA) from maternal plasma, so-called non-invasive prenatal testing (NIPT). However, many technical challenges, such as dependency on correct fetal sex prediction, large variations of chromosome Y measurement and high sensitivity to random reads mapping, may result in higher false negative rate (FNR) and false positive rate (FPR) in fetal sex prediction as well as in SCAs detection. Here, we developed an optimized method to improve the accuracy of the current method by filtering out randomly mapped reads in six specific regions of the Y chromosome. The method reduces the FNR and FPR of fetal sex prediction from nearly 1% to 0.01% and 0.06%, respectively and works robustly under conditions of low fetal DNA concentration (1%) in testing and simulation of 92 samples. The optimized method was further confirmed by large scale testing (1590 samples), suggesting that it is reliable and robust enough for clinical testing. PMID:27441628

  4. Chromosomal lesions and uniparental disomy detected by SNP arrays in MDS, MDS/MPD, and MDS-derived AML

    PubMed Central

    Gondek, Lukasz P.; Tiu, Ramon; O'Keefe, Christine L.; Sekeres, Mikkael A.; Theil, Karl S.

    2008-01-01

    Using metaphase cytogenetics (MC), chromosomal abnormalities are found in only a proportion of patients with myelodysplastic syndrome (MDS). We hypothesized that with new precise methods more cryptic karyotypic lesions can be uncovered that may show important clinical implications. We have applied 250K single nucleotide polymorphisms (SNP) arrays (SNP-A) to study chromosomal lesions in samples from 174 patients (94 MDS, 33 secondary acute myeloid leukemia [sAML], and 47 myelodysplastic/myeloproliferative disease [MDS/MPD]) and 76 controls. Using SNP-A, aberrations were found in around three-fourths of MDS, MDS/MPD, and sAML (vs 59%, 37%, 53% by MC; in 8% of patients MC was unsuccessful). Previously unrecognized lesions were detected in patients with normal MC and in those with known lesions. Moreover, segmental uniparental disomy (UPD) was found in 20% of MDS, 23% of sAML, and 35% of MDS/MPD patients, a lesion resulting in copy-neutral loss of heterozygosity undetectable by MC. The potential clinical significance of abnormalities detected by SNP-A, but not seen on MC, was demonstrated by their impact on overall survival. UPD involving chromosomes frequently affected by deletions may have prognostic implications similar to the deletions visible by MC. SNP-A–based karyotyping shows superior resolution for chromosomal defects, including UPD. This technique further complements MC to improve clinical prognosis and targeted therapies. PMID:17954704

  5. In silico and in vitro evaluation of PCR-based assays for the detection of Bacillus anthracis chromosomal signature sequences

    PubMed Central

    Ågren, Joakim; Hamidjaja, Raditijo A; Hansen, Trine; Ruuls, Robin; Thierry, Simon; Vigre, Håkan; Janse, Ingmar; Sundström, Anders; Segerman, Bo; Koene, Miriam; Löfström, Charlotta; Van Rotterdam, Bart; Derzelle, Sylviane

    2013-01-01

    Bacillus anthracis, the causative agent of anthrax, is a zoonotic pathogen that is relatively common throughout the world and may cause life threatening diseases in animals and humans. There are many PCR-based assays in use for the detection of B. anthracis. While most of the developed assays rely on unique markers present on virulence plasmids pXO1 and pXO2, relatively few assays incorporate chromosomal DNA markers due to the close relatedness of B. anthracis to the B. cereus group strains. For the detection of chromosomal DNA, different genes have been used, such as BA813, rpoB, gyrA, plcR, S-layer, and prophage-lambda. Following a review of the literature, an in silico analysis of all signature sequences reported for identification of B. anthracis was conducted. Published primer and probe sequences were compared for specificity against 134 available Bacillus spp. genomes. Although many of the chromosomal targets evaluated are claimed to be specific to B. anthracis, cross-reactions with closely related B. cereus and B. thuringiensis strains were often observed. Of the 35 investigated PCR assays, only 4 were 100% specific for the B. anthracis chromosome. An interlaboratory ring trial among five European laboratories was then performed to evaluate six assays, including the WHO recommended procedures, using a collection of 90 Bacillus strains. Three assays performed adequately, yielding no false positive or negative results. All three assays target chromosomal markers located within the lambdaBa03 prophage region (PL3, BA5345, and BA5357). Detection limit was further assessed for one of these highly specific assays. PMID:24005110

  6. In silico and in vitro evaluation of PCR-based assays for the detection of Bacillus anthracis chromosomal signature sequences.

    PubMed

    Ågren, Joakim; Hamidjaja, Raditijo A; Hansen, Trine; Ruuls, Robin; Thierry, Simon; Vigre, Håkan; Janse, Ingmar; Sundström, Anders; Segerman, Bo; Koene, Miriam; Löfström, Charlotta; Van Rotterdam, Bart; Derzelle, Sylviane

    2013-11-15

    Bacillus anthracis, the causative agent of anthrax, is a zoonotic pathogen that is relatively common throughout the world and may cause life threatening diseases in animals and humans. There are many PCR-based assays in use for the detection of B. anthracis. While most of the developed assays rely on unique markers present on virulence plasmids pXO1 and pXO2, relatively few assays incorporate chromosomal DNA markers due to the close relatedness of B. anthracis to the B. cereus group strains. For the detection of chromosomal DNA, different genes have been used, such as BA813, rpoB, gyrA, plcR, S-layer, and prophage-lambda. Following a review of the literature, an in silico analysis of all signature sequences reported for identification of B. anthracis was conducted. Published primer and probe sequences were compared for specificity against 134 available Bacillus spp. genomes. Although many of the chromosomal targets evaluated are claimed to be specific to B. anthracis, cross-reactions with closely related B. cereus and B. thuringiensis strains were often observed. Of the 35 investigated PCR assays, only 4 were 100% specific for the B. anthracis chromosome. An interlaboratory ring trial among five European laboratories was then performed to evaluate six assays, including the WHO recommended procedures, using a collection of 90 Bacillus strains. Three assays performed adequately, yielding no false positive or negative results. All three assays target chromosomal markers located within the lambdaBa03 prophage region (PL3, BA5345, and BA5357). Detection limit was further assessed for one of these highly specific assays.

  7. Visualizing Chromosome Mosaicism and Detecting Ethnic Outliers by the Method of “Rare” Heterozygotes and Homozygotes (RHH)

    PubMed Central

    McGinnis, Ralph E.; Deloukas, Panos; McLaren, William M.; Inouye, Michael

    2010-01-01

    We describe a novel approach for evaluating SNP genotypes of a genome-wide association scan to identify “ethnic outlier” subjects whose ethnicity is different or admixed compared to most other subjects in the genotyped sample set. Each ethnic outlier is detected by counting a genomic excess of “rare” heterozygotes and/or homozygotes whose frequencies are low (<1%) within genotypes of the sample set being evaluated. This method also enables simple and striking visualization of non-Caucasian chromosomal DNA segments interspersed within the chromosomes of ethnically admixed individuals. We show that this visualization of the mosaic structure of admixed human chromosomes gives results similar to another visualization method (SABER) but with much less computational time and burden. We also show that other methods for detecting ethnic outliers are enhanced by evaluating only genomic regions of visualized admixture rather than diluting outlier ancestry by evaluating the entire genome considered in aggregate. We have validated our method in the Wellcome Trust Case Control Consortium (WTCCC) study of 17,000 subjects as well as in HapMap subjects and simulated outliers of known ethnicity and admixture. The method's ability to precisely delineate chromosomal segments of non-Caucasian ethnicity has enabled us to demonstrate previously unreported non-Caucasian admixture in two HapMap Caucasian parents and in a number of WTCCC subjects. Its sensitive detection of ethnic outliers and simple visual discrimination of discrete chromosomal segments of different ethnicity implies that this method of rare heterozygotes and homozygotes (RHH) is likely to have diverse and important applications in humans and other species. PMID:20211853

  8. Detection of chromosomal abnormalities, congenital abnormalities and transfusion syndrome in twins.

    PubMed

    Sperling, L; Kiil, C; Larsen, L U; Brocks, V; Wojdemann, K R; Qvist, I; Schwartz, M; Jørgensen, C; Espersen, G; Skajaa, K; Bang, J; Tabor, A

    2007-05-01

    To evaluate the outcome of screening for structural malformations in twins and the outcome of screening for twin-twin transfusion syndrome (TTTS) among monochorionic twins through a number of ultrasound scans from 12 weeks' gestation. Enrolled into this prospective multicenter observational study were women with twin pregnancies diagnosed before 14 + 6 gestational weeks. The monochorionic pregnancies were scanned every second week until 23 weeks in order to rule out early TTTS. All pregnancies had an anomaly scan in week 19 and fetal echocardiography in week 21 that was performed by specialists in fetal echocardiography. Zygosity was determined by DNA analysis in all twin pairs with the same sex. Among the 495 pregnancies the prenatal detection rate for severe structural abnormalities including chromosomal aneuploidies was 83% by the combination of a first-trimester nuchal translucency scan and the anomaly scan in week 19. The incidence of severe structural abnormalities was 2.6% and two-thirds of these anomalies were cardiac. There was no significant difference between the incidence in monozygotic and dizygotic twins, nor between twins conceived naturally or those conceived by assisted reproduction. The incidence of TTTS was 23% from 12 weeks until delivery, and all those monochorionic twin pregnancies that miscarried had signs of TTTS. Twin pregnancies have an increased risk of congenital malformations and one out of four monochorionic pregnancies develops TTTS. Ultrasound screening to assess chorionicity and follow-up of monochorionic pregnancies to detect signs of TTTS, as well as malformation screening, are therefore essential in the antenatal care of twin pregnancies. Copyright (c) 2007 ISUOG.

  9. Sex chromosome trisomies in Europe: prevalence, prenatal detection and outcome of pregnancy

    PubMed Central

    Boyd, Patricia Anne; Loane, Maria; Garne, Ester; Khoshnood, Babak; Dolk, Helen

    2011-01-01

    This study aims to assess prevalence and pregnancy outcome for sex chromosome trisomies (SCTs) diagnosed prenatally or in the first year of life. Data held by the European Surveillance of Congenital Anomalies (EUROCAT) database on SCT cases delivered 2000–2005 from 19 population-based registries in 11 European countries covering 2.5 million births were analysed. Cases included were livebirths diagnosed to 1 year of age, fetal deaths from 20 weeks gestation and terminations of pregnancy for fetal anomaly (TOPFA). In all, 465 cases of SCT were diagnosed between 2000 and 2005, a prevalence of 1.88 per 10,000 births (95% CI 1.71–2.06). Prevalence of XXX, XXY and XYY were 0.54 (95% CI 0.46–0.64), 1.04 (95% CI 0.92–1.17) and 0.30 (95% CI 0.24–0.38), respectively. In all, 415 (89%) were prenatally diagnosed and 151 (36%) of these resulted in TOPFA. There was wide country variation in prevalence (0.19–5.36 per 1000), proportion prenatally diagnosed (50–100%) and proportion of prenatally diagnosed resulting in TOPFA (13–67%). Prevalence of prenatally diagnosed cases was higher in countries with high prenatal detection rates of Down syndrome. The EUROCAT prevalence rate for SCTs diagnosed prenatally or up to 1 year of age represents 12% of the prevalence expected from cytogenetic studies of newborn babies, as the majority of cases are never diagnosed or are diagnosed later in life. There is a wide variation between European countries in prevalence, prenatal detection and TOPFA proportions, related to differences in screening policies as well as organizational and cultural factors. PMID:20736977

  10. Utility of Linearly Amplified RNA for RT-PCR Detection of Chromosomal Translocations

    PubMed Central

    Schumacher, Jonathan A.; Jenson, Stephen D.; Elenitoba-Johnson, Kojo S. J.; Lim, Megan S.

    2004-01-01

    The requirement for sufficient quantities of starting RNA has limited the ability to evaluate multiple transcripts using reverse transcriptase-polymerase chain reaction (RT-PCR). In this study, we demonstrate the utility of linear RNA amplification for RT-PCR analysis of multiple gene transcripts including a chromosomal translocation, using the t(2;5)(p23;q35) as a model. RNA from the t(2;5)-positive cell line, SU-DHL-1, and the t(2;5)-negative cell line, HUT-78, was extracted and exposed to two rounds of linear amplification. RT-PCR using cDNA from the resultant amplified (a) RNA and total RNA resulted in the 177 bp NPM-ALK fusion gene product from the SU-DHL-1 cell line, but not from aRNA or total RNA from the HUT-78 cell line. DNA sequencing of the RT-PCR products from total and aRNA of SU-DHL-1 cells demonstrated identical sequences corresponding to the NPM-ALK fusion gene. Evaluation of 25 snap-frozen tissue samples, including eight NPM-ALK-positive ALCLs demonstrated 100% concordance of t(2;5) detection between cDNA from total RNA and that from aRNA. Our results show that linear amplification of RNA can enhance starting RNA greater than 200-fold and can be used for rapid and specific detection of multiplex gene expression from a variety of sources. This method can generate a renewable archive of representative cDNA, which can be used for retrospective screening of stored samples as well as positive controls for the clinical molecular diagnostic laboratory. PMID:14736822

  11. Detection of low level sex chromosome mosaicism in Ullrich-Turner syndrome patients.

    PubMed

    Wiktor, Anne E; Van Dyke, Daniel L

    2005-10-15

    Ullrich-Turner syndrome (UTS) is most commonly due to a 45,X chromosome defect, but is also seen in patients with a variety of X-chromosome abnormalities or 45,X/46,XY mosaicism. The phenotype of UTS patients is highly variable, and depends largely on the karyotype. Patients are at an increased risk of gonadoblastoma when a Y-derived chromosome or chromosome fragment is present. Since constitutional mosaicism is present in approximately 50% of UTS patients, the identification of minor cell populations is clinically important and a challenge to laboratories. We identified 50 females with a 45,X karyotype as the sole abnormality or as part of a more complex karyotype. Twenty two (44%) had a 45,X karyotype; mosaicism for a second normal or structurally abnormal X was observed in 24 (48%) samples, and mosaicism for Y chromosomal material in 4 (8%) cases. To further investigate the possibility of mosaicism in the 22 patients with an apparently non-mosaic 45,X karyotype, we performed FISH using centromere probes for the X and Y chromosomes. A minor XX cell line was identified in 3 patients, and the 45,X result was confirmed in 19 samples. No samples with XY mosaicism were identified. We describe our validation process for a FISH assay to be used in clinical practice to identify XX or XY mosaicism. FISH as an adjunct to karyotype analysis provides a sensitive and cost-effective technique to identify sex chromosome mosaicism in UTS patients.

  12. Detected microsatellite polymorphisms in genetically altered inbred mouse strains.

    PubMed

    Du, Xiaoyan; Cui, Jing; Wang, Chao; Huo, Xueyun; Lu, Jing; Li, Yichen; Chen, Zhenwen

    2013-08-01

    Microsatellites are 50-200 repetitive DNA sequences composed of 1- to 6-base-pair-long reiterative motifs within the genome. They are vulnerable to DNA modifications, such as recombination and/or integration, and are recognized as "sentinel" DNA. Our previous report indicated that the genotypes of the microsatellite loci could change from mono- to poly-morphisms (CMP) in gene knockout (KO) mice, implying that genetic modification induces microsatellite mutation. However, it is still unclear whether the random insertion of DNA fragments into mice genomes produced via transgene (Tg) or N-ethyl-N-nitrosourea (ENU) would also result in microsatellite mutations or microsatellite loci genotypes changes. This study was designed to find possible clues to answer this question. In brief, 198 microsatellite loci that were distributed among almost all of the chromosomes (except for the Y) were examined through polymerase chain reaction to screen possible CMPs in six Tg strains. First, for each strain, the microsatellite sequences of all loci were compared between Tg and the corresponding background strain to exclude genetic interference. Simultaneously, to exclude spontaneous mutation-related CMPs that might exist in the examined six strains, mice from five spontaneously mutated inbred strains were used as the negative controls. Additionally, the sequences of all loci in these spontaneous mutated mice were compared to corresponding genetic background controls. The results showed that 40 of the 198 (20.2%) loci were identified as having CMPs in the examined Tg mice strains. The CMP genotypes were either homozygous or heterozygous compared to the background controls. Next, we applied the 40 CMP positive loci in ENU-mutated mice and their corresponding background controls. After that, a general comparison of CMPs that exist among Tg, ENU-treated and KO mouse strains was performed. The results indicated that four (D11mit258, D13mit3, D14mit102 and DXmit172) of the 40 (10%) CMP

  13. Structural brain alterations can be detected early in HIV infection

    PubMed Central

    Ochs, Renee; Wu, Ying; Sammet, Christina L.; Shoukry, Alfred; Epstein, Leon G.

    2012-01-01

    Objective: Brain changes occurring early in HIV infection are not well characterized. The Chicago Early HIV Infection Study aimed to evaluate the presence and extent of structural brain alterations using quantitative MRI. Methods: Forty-three HIV and 21 control subjects were enrolled. Mean length of infection was estimated as less than 1 year based on assay results. High-resolution neuroanatomical images were acquired. Automated image analysis was used to derive measurements for total brain, ventricular volume, and for tissue classes (total and cortical gray matter, white matter, and CSF). A separate image analysis algorithm was used to calculate measurements for individual brain regions. Cognitive function was assessed by neuropsychological evaluation. Results: Reductions were quantified in total (p = 0.0547) and cortical (p = 0.0109) gray matter in the HIV group. Analysis of individual brain regions with a separate image analysis algorithm revealed consistent findings of reductions in cerebral cortex (p = 0.042) and expansion of third ventricle (p = 0.046). The early HIV group also demonstrated weaker performance on several neuropsychological tests, with the most pronounced difference in psychomotor speed (p = 0.001). Conclusions: This cross-sectional brain volumetric study indicates structural alterations early in HIV infection. The findings challenge the prevailing assumption that the brain is spared in this period. Revisiting the question of the brain's vulnerability to processes unfolding in the initial virus-host interaction and the early natural history may yield new insights into neurologic injury in HIV infection and inform neuroprotection strategies. PMID:23197750

  14. Identification of Stmm3 locus Conferring Resistance to Late-stage Chemically Induced Skin Papillomas on Mouse Chromosome 4 by Congenic Mappingand Allele-specific Alteration Analysis

    PubMed Central

    Saito, Megumi; Okumura, Kazuhiro; Miura, Ikuo; Wakana, Shigeharu; Kominami, Ryo; Wakabayashi, Yuichi

    2014-01-01

    Genome-wide association studies have revealed that many low-penetrance cancer susceptibility loci are located throughout the genome; however, a very limited number of genes have been identified so far. Using a forward genetics approach to map such loci in a mouse skin cancer model, we previously identified strong genetic loci conferring resistance to chemically induced skin papillomas on chromosome 4 and 7 with a large number of [(FVB/N × MSM/Ms) F1 × FVB/N] backcross mice. In this report, we describe a combination of congenic mapping and allele-specific alteration analysis of the loci on chromosome 4. We used linkage analysis and a congenic mouse strain, FVB.MSM-Stmm3 to refine the location of Stmm3 (Skin tumor modifier of MSM 3) locus within a physical interval of about 34 Mb on distal chromosome 4. In addition, we used patterns of allele-specific imbalances in tumors from N2 and N10 congenic mice to narrow down further the region of Stmm3 locus to a physical distance of about 25 Mb. Furthermore, immunohistochemical analysis showed papillomas from congenic mice had less proliferative activity. These results suggest that Stmm3 responsible genes may have an influence on papilloma formation in the two-stage skin carcinogenesis by regulating papilloma growth rather than development. PMID:25077764

  15. V(D)J recombinase mediated inter-chromosomal HPRT alterations at cryptic recombination signal sequences in peripheral human T cells.

    PubMed

    Messier, Terri L; O'Neill, J Patrick; Finette, Barry A

    2006-08-01

    The V(D)J recombinase enzyme complex is responsible for the development of a diverse immune system by catalyzing intra-molecular rearrangements of immunoglobulin (Ig) and T cell receptor (TCR) genes at specific recombination signal sequences (RSSs). This enzyme complex has also been implicated in mediating pathologic and non-pathologic intra- and inter-molecular genomic rearrangements at cryptic (Psi) RSSs outside the immune system loci in lymphoid cells. We describe here two V(D)J recombinase mediated genomic rearrangements resulting in alterations at the HPRT locus in human T-cells. These are inter-chromosomal insertions in which DNA fragments are inserted at breakpoints generated by V(D)J recombinase cleavage at Psi RSS sites in the HPRT locus at Xq26. In the first, a TCR signal ended segment from chromosome 14q11 is inserted at a Psi RSS in intron 1 of the HPRT locus. In the second, a DNA fragment from 9q22 is integrated between the coding ends generated by a V(D)J recombinase mediated HPRT deletion. Identification of these in vivo V(D)J mediated inter-chromosomal insertions at Psi RSSs in the HPRT gene supports the accumulating evidence that V(D)J recombinase can mediate mutagenic rearrangements in humans with potential pathologic consequences.

  16. Spectra of Chromosomal Aberrations in 325 Leukemia Patients and Implications for the Development of New Molecular Detection Systems

    PubMed Central

    Choi, Hyun-Jung; Kim, Hye-Ran; Kook, Hoon; Kim, Hyeoung-Joon; Shin, Jong-Hee; Suh, Soon-Pal; Ryang, Dong-Wook

    2011-01-01

    This study investigated the spectrum of chromosomal abnormalities in 325 leukemia patients and developed optimal profiles of leukemic fusion genes for multiplex RT-PCR. We prospectively analyzed blood and bone marrow specimens of patients with acute leukemia. Twenty types of chromosomal abnormalities were detected in 42% from all patients by commercially available multiplex RT-PCR for detecting 28 fusion genes and in 35% by cytogenetic analysis including FISH analysis. The most common cytogenetic aberrations in acute myeloid leukemia patients was PML/PARA, followed by AML1/MGT8 and MLL1, and in acute lymphoid leukemia patients was BCR/ABL, followed by TEL/AML1 and MLL1 gene rearrangement. Among the negative results for multiplex RT-PCR, clinically significant t(3;3)(q21;q26.2), t(8;14)(q24;q32) and i(17)(q10) were detected by conventional cytogenetics. The spectrum and frequency of chromosomal abnormalities in our leukemia patients are differed from previous studies, and may offer optimal profiles of leukemic fusion genes for the development of new molecular detection systems. PMID:21738341

  17. Detection of a large RIII-derived chromosomal segment on chromosome 10 in the H-2 congenic strain B10.RIII(71NS)/Sn

    SciTech Connect

    Dong, P.; Hood, L.; McIndoe, R.A.

    1996-01-15

    This report describes the results of a study of the chromosomal localization of certain loci related to the susceptibility of specific mouse strains to collagen-induced arthritis, the biological model for rheumatoid arthritis. There were surprising results concerning the chromosomal mapping of mouse chromosome 10 and 17 and the backcrosses of mice involved. 7 refs., 1 fig., 2 tabs.

  18. Rapid detection of chromosome aneuploidies by quantitative fluorescence PCR: first application on 247 chorionic villus samples

    PubMed Central

    Pertl, B.; Kopp, S.; Kroisel, P.; Tului, L.; Brambati, B.; Adinolfi, M.

    1999-01-01

    We report the results of the first major study of applying quantitative fluorescence polymerase chain reaction (QF-PCR) assays for the detection of major chromosome numerical disorders. The QF-PCR tests were performed on a total of 247 chorionic villus samples, which were analysed blind, without any knowledge of the results obtained using conventional cytogenetic analysis.
The aims of this investigation were to evaluate the detection power and accuracy of this approach by testing a large number of fetal samples and to assess the diagnostic value of each of the chromosome specific small tandem repeat (STR) markers used. In addition, we introduced three more markers specific for chromosomes 13, 18, and X to allow an accurate analysis of samples homozygous for a particular STR. Fluorescent labelled primers were used to amplify 12 STRs specific for chromosomes 21, 18, 13, X, and the amylogenin-like DNA sequence AMXY, expressed on the X and Y chromosomes.
In this blind study of 247 fetal samples, 222 were correctly diagnosed by QF-PCR as normal for each of the five chromosomes investigated; 20 were diagnosed by QF-PCR as trisomic for chromosomes 21, 18, or 13, in agreement with the cytogenetic tests. Only one false negative result was observed, probably owing to the mishandling of the sample, which had been transferred through three laboratories before being analysed by QF-PCR. The 247 samples also included four cases of mosaicism or translocation; one case of mosaic trisomy 21 was detected by QF-PCR and the other cases were not identified by QF-PCR.
The results of this investigation provide clear evidence that the QF-PCR assays are powerful adjuncts to conventional cytogenetic techniques and can be applied for the rapid and accurate prenatal diagnosis of the most frequent aneuploidies.


Keywords: prenatal diagnosis; aneuploidies; quantitative fluorescence PCR PMID:10227397

  19. A non-syndromic intellectual disability associated with a de novo microdeletion at 7q and 18p, microduplication at Xp, and 18q partial trisomy detected using chromosomal microarray analysis approach

    PubMed Central

    2014-01-01

    Background Chromosome abnormalities that segregate with a disease phenotype can facilitate the identification of disease loci and genes. The relationship between chromosome 18 anomalies with severe intellectual disability has attracted the attention of cytogeneticists worldwide. Duplications of the X chromosome can cause intellectual disability in females with variable phenotypic effects, due in part to variations in X-inactivation patterns. Additionally, deletions of the 7qter region are associated with a range of phenotypes. Results We report the first case of de novo microdeletion at 7q and 18p, 18q partial trisomy, microduplication at Xp associated to intellectual disability in a Brazilian child, presenting a normal karyotype. Karyotyping showed any chromosome alteration. Chromosomal microarray analysis detected a de novo microdeletion at 18p11.32 and 18q partial trisomy, an inherited microdeletion at 7q31.1 and a de novo microduplication at Xp22.33p21.3. Conclusions Our report illustrates a case that presents complex genomic imbalances which may contribute to a severe clinical phenotypes. The rare and complex phenotypes have to be investigated to define the subsets and allow the phenotypes classification. PMID:25028595

  20. A non-syndromic intellectual disability associated with a de novo microdeletion at 7q and 18p, microduplication at Xp, and 18q partial trisomy detected using chromosomal microarray analysis approach.

    PubMed

    Pinto, Irene Plaza; Minasi, Lysa Bernardes; da Cruz, Alex Silva; de Melo, Aldaires Vieira; da Cruz E Cunha, Damiana Míriam; Pereira, Rodrigo Roncato; Ribeiro, Cristiano Luiz; da Silva, Claudio Carlos; de Melo E Silva, Daniela; da Cruz, Aparecido Divino

    2014-01-01

    Chromosome abnormalities that segregate with a disease phenotype can facilitate the identification of disease loci and genes. The relationship between chromosome 18 anomalies with severe intellectual disability has attracted the attention of cytogeneticists worldwide. Duplications of the X chromosome can cause intellectual disability in females with variable phenotypic effects, due in part to variations in X-inactivation patterns. Additionally, deletions of the 7qter region are associated with a range of phenotypes. We report the first case of de novo microdeletion at 7q and 18p, 18q partial trisomy, microduplication at Xp associated to intellectual disability in a Brazilian child, presenting a normal karyotype. Karyotyping showed any chromosome alteration. Chromosomal microarray analysis detected a de novo microdeletion at 18p11.32 and 18q partial trisomy, an inherited microdeletion at 7q31.1 and a de novo microduplication at Xp22.33p21.3. Our report illustrates a case that presents complex genomic imbalances which may contribute to a severe clinical phenotypes. The rare and complex phenotypes have to be investigated to define the subsets and allow the phenotypes classification.

  1. Detection of cryptic chromosomal abnormalities in unexplained mental retardation: A general strategy using hypervariable subtelomeric DNA polymorphisms

    SciTech Connect

    Wilkie, A.O.M.

    1993-09-01

    Given the availability of DNA from both parents, unusual segregation of hypervariable DNA polymorphisms (HVPs) in the offspring may be attributable to deletion, unbalanced chromosomal translocation, or uniparental disomy. The telomeric regions of chromosomes are rich in both genes and hypervariable minisatellite sequences and may also be particularly prone to cryptic breakage events. Here the author describes and analyzes a general approach to the detection of subtelomeric abnormalities and uniparental disomy in patients with unexplained mental retardation. With 29 available polymorphic systems, [approximately]50%-70% of these abnormalities could currently be detected. Development of subtelomeric HVPs physically localized with respect to their telomers should provide a valuable resource in routine diagnostics. 73 refs., 4 figs., 4 tabs.

  2. Detection of genetic alterations in advanced prostate cancer by comparative genomic hybridization.

    PubMed

    Kasahara, Kotaro; Taguchi, Takahiro; Yamasaki, Ichiro; Kamada, Masayuki; Yuri, Kazunari; Shuin, Taro

    2002-08-01

    In this study, we examined nine cases of advanced Japanese prostate cancer by comparative genomic hybridization (CGH) to detect chromosomal imbalances across the entire genome and to identify several new regions likely to contain genes important to the development and progression of this disease. These cases had been previously examined for numerical chromosomal aberrations by fluorescence in situ hybridization (FISH). By CGH, the following regions were found to be over-represented (gains), with fluorescence ratio values higher than the threshold: 4p, 6p, 8q, 11q, 12q, 15q, 16p, 17q, 20, and 21 (>4 cases); underrepresentation (losses) involved: 1q, 4q, 5q, 6q, 13q, 14q, and 22 (>4 cases). The shortest regions of overlap (SRO) of gains were noted at 8q24.1 through q24.3, 12q23, and 17q23 through q24 (>5 cases). The SRO of losses were seen at 5q14 through q21, 6q16.1 through q21, 13q21.3 through q22, and 14q21 (>5 cases). Notably, the gain of chromosomes 8 and 12 by CGH was in agreement with the FISH data, suggesting that the gain of chromosomes 8 and 12 may play an important role in prostate carcinogenesis. The genes on the SRO regions were also discussed in relation to oncogenes and bone metastases.

  3. Genetic Alterations in Pesticide Exposed Bolivian Farmers: An evaluation by analysis of chromosomal aberrations and the comet assay.

    PubMed

    Jørs, Erik; Gonzáles, Ana Rosa; Ascarrunz, Maria Eugenia; Tirado, Noemi; Takahashi, Catharina; Lafuente, Erika; Dos Santos, Raquel A; Bailon, Natalia; Cervantes, Rafael; O, Huici; Bælum, Jesper; Lander, Flemming

    2007-11-12

    Pesticides are of concern in Bolivia because of increasing use. Frequent intoxications have been demonstrated due to use of very toxic pesticides, insufficient control of distribution and sale and little knowledge among farmers of protective measures and hygienic procedures. Questionnaires were applied and blood tests taken from 81 volunteers from La Paz County, of whom 48 were pesticide exposed farmers and 33 non-exposed controls. Sixty males and 21 females participated with a mean age of 37.3 years (range 17-76). Data of exposure and possible genetic damage were collected and evaluated by well known statistical methods, controlling for relevant confounders. To measure genetic damage chromosomal aberrations and the comet assay analysis were performed. Pesticide exposed farmers had a higher degree of genetic damage compared to the control group. The number of chromosomal aberrations increased with the intensity of pesticide exposure. Females had a lower number of chromosomal aberrations than males, and people living at altitudes above 2500 metres seemed to exhibit more DNA damage measured by the comet assay. Bolivian farmers showed signs of genotoxic damage, probably related to exposure to pesticides. Due to the potentially negative long term health effects of genetic damage on reproduction and the development of cancer, preventive measures are recommended. Effective control with imports and sales, banning of the most toxic pesticides, education and information are possible measures, which could help preventing the negative effects of pesticides on human health and the environment.

  4. The rad52-Y66A allele alters the choice of donor template during spontaneous chromosomal recombination.

    PubMed

    de Mayolo, Adriana Antúnez; Sunjevaric, Ivana; Reid, Robert; Mortensen, Uffe H; Rothstein, Rodney; Lisby, Michael

    2010-01-02

    Spontaneous mitotic recombination is a potential source of genetic changes such as loss of heterozygosity and chromosome translocations, which may lead to genetic disease. In this study we have used a rad52 hyper-recombination mutant, rad52-Y66A, to investigate the process of spontaneous heteroallelic recombination in the yeast Saccharomyces cerevisiae. We find that spontaneous recombination has different genetic requirements, depending on whether the recombination event occurs between chromosomes or between chromosome and plasmid sequences. The hyper-recombination phenotype of the rad52-Y66A mutation is epistatic with deletion of MRE11, which is required for establishment of DNA damage-induced cohesion. Moreover, single-cell analysis of strains expressing YFP-tagged Rad52-Y66A reveals a close to wild-type frequency of focus formation, but with foci lasting 6 times longer. This result suggests that spontaneous DNA lesions that require recombinational repair occur at the same frequency in wild-type and rad52-Y66A cells, but that the recombination process is slow in rad52-Y66A cells. Taken together, we propose that the slow recombinational DNA repair in the rad52-Y66A mutant leads to a by-pass of the window-of-opportunity for sister chromatid recombination normally promoted by MRE11-dependent damage-induced cohesion thereby causing a shift towards interchromosomal recombination.

  5. Segmental chromosomal alterations lead to a higher risk of relapse in infants with MYCN-non-amplified localised unresectable/disseminated neuroblastoma (a SIOPEN collaborative study)

    PubMed Central

    Schleiermacher, G; Michon, J; Ribeiro, A; Pierron, G; Mosseri, V; Rubie, H; Munzer, C; Bénard, J; Auger, N; Combaret, V; Janoueix-Lerosey, I; Pearson, A; Tweddle, D A; Bown, N; Gerrard, M; Wheeler, K; Noguera, R; Villamon, E; Cañete, A; Castel, V; Marques, B; de Lacerda, A; Tonini, G P; Mazzocco, K; Defferrari, R; de Bernardi, B; di Cataldo, A; van Roy, N; Brichard, B; Ladenstein, R; Ambros, I; Ambros, P; Beiske, K; Delattre, O; Couturier, J

    2011-01-01

    Background: In neuroblastoma (NB), the presence of segmental chromosome alterations (SCAs) is associated with a higher risk of relapse. Methods: In order to analyse the role of SCAs in infants with localised unresectable/disseminated NB without MYCN amplification, we have performed an array CGH analysis of tumours from infants enroled in the prospective European INES trials. Results: Tumour samples from 218 out of 300 enroled patients could be analysed. Segmental chromosome alterations were observed in 11%, 20% and 59% of infants enroled in trials INES99.1 (localised unresectable NB), INES99.2 (stage 4s) and INES99.3 (stage 4) (P<0.0001). Progression-free survival was poorer in patients whose tumours harboured SCA, in the whole population and in trials INES99.1 and INES99.2, in the absence of clinical symptoms (log-rank test, P=0.0001, P=0.04 and P=0.0003, respectively). In multivariate analysis, a SCA genomic profile was the strongest predictor of poorer progression-free survival. Conclusion: In infants with stage 4s MYCN-non-amplified NB, a SCA genomic profile identifies patients who will require upfront treatment even in the absence of other clinical indication for therapy, whereas in infants with localised unresectable NB, a genomic profile characterised by the absence of SCA identifies patients in whom treatment reduction might be possible. These findings will be implemented in a future international trial. PMID:22146831

  6. Patterns of Chromosomal Aberrations in Solid Tumors

    PubMed Central

    Grade, Marian; Difilippantonio, Michael J.

    2016-01-01

    Chromosomal abnormalities are a defining feature of solid tumors. Such cytogenetic alterations are mainly classified into structural chromosomal aberrations and copy number alterations, giving rise to aneuploid karyotypes. The increasing detection of these genetic changes allowed the description of specific tumor entities and the associated patterns of gene expression. In fact, tumor-specific landscapes of gross genomic copy number changes, including aneuploidies of entire chromosome arms and chromosomes result in a global deregulation of the transcriptome of cancer cells. Furthermore, the molecular characterization of cytogenetic abnormalities has provided insights into the mechanisms of tumorigenesis and has, in a few instances, led to the clinical implementation of effective diagnostic and prognostic tools, as well as treatment strategies that target a specific genetic abnormality. PMID:26376875

  7. Rapid detection of genomic imbalances using micro-arrays consisting of pooled BACs covering all human chromosome arms.

    PubMed

    Knijnenburg, Jeroen; van der Burg, Marja; Nilsson, Philomeen; Ploos van Amstel, Hans Kristian; Tanke, Hans; Szuhai, Károly

    2005-10-12

    A strategy is presented to select, pool and spot human BAC clones on an array in such a way that each spot contains five well performing BAC clones, covering one chromosome arm. A mini-array of 240 spots was prepared representing all human chromosome arms in a 5-fold as well as some controls, and used for comparative genomic hybridization (CGH) of 10 cell lines with aneusomies frequently found in clinical cytogenetics and oncology. Spot-to-spot variation within five replicates was below 6% and all expected abnormalities were detected 100% correctly. Sensitivity was such that replacing one BAC clone in a given spot of five by a BAC clone from another chromosome, thus resulting in a change in ratio of 20%, was reproducibly detected. Incubation time of the mini-array was varied and the fluorescently labelled target DNA was diluted. Typically, aneusomies could be detected using 30 ng of non-amplified random primed labelled DNA amounts in a 4 h hybridization reaction. Potential application of these mini-arrays for genomic profiling of disseminated tumour cells or of blastomeres for preimplantation genetic diagnosis, using specially designed DNA amplification methods, are discussed.

  8. Induction of chromosome instability and stomach cancer by altering the expression pattern of mitotic checkpoint genes in mice exposed to areca-nut

    PubMed Central

    2013-01-01

    Background There are strong indications for a causal association between areca-nut consumption and cancers. In Meghalaya, India, the variety of areca-nut is used as raw and unprocessed form whose chemical composition and pharmacological actions have been reported. Yet we know little on the initial pathway involved in areca-nut associated carcinogenesis since it is difficult to assess its effects on genetic alterations without interference of other compounding factors. Therefore, present study was undertaken in mice to verify the ability of raw areca-nut (RAN) to induce cancer and to monitor the expression of certain genes involved in carcinogenesis. This study was not intended to isolate any active ingredients from the RAN and to look its action. Methods Three groups of mice (n = 25 in each) were taken and used at different time-points for different experimental analysis. The other three groups of mice (n = 15 in each) were considered for tumor induction studies. In each set, two groups were administered RAN-extract ad libitum in drinking water with or without lime. The expression of certain genes was assessed by conventional RT-PCR and immunoblotting. The mice were given the whole RAN-extract with and without lime in order to mimic the human consumption style of RAN. Results Histological preparation of stomach tissue revealed that RAN induced stomach cancer. A gradual increase in the frequency of precocious anaphase and aneuploid cells was observed in the bone marrow cells with a greater increment following RAN + lime administeration. Levels of p53, Bax, Securin and p65 in esophageal and stomach cells were elevated during early days of RAN exposure while those of different mitotic checkpoint proteins were downregulated. Apoptotic cell death was detected in non-cancerous stomach cells but not in tumor cells which showed overexpression of Bax and absence of PARP. Conclusion Present study suggested (a) RAN induces stomach cancer, however, presence of lime

  9. Induction of chromosome instability and stomach cancer by altering the expression pattern of mitotic checkpoint genes in mice exposed to areca-nut.

    PubMed

    Kurkalang, Sillarine; Banerjee, Atanu; Ghoshal, Nitin; Dkhar, Hughbert; Chatterjee, Anupam

    2013-06-28

    There are strong indications for a causal association between areca-nut consumption and cancers. In Meghalaya, India, the variety of areca-nut is used as raw and unprocessed form whose chemical composition and pharmacological actions have been reported. Yet we know little on the initial pathway involved in areca-nut associated carcinogenesis since it is difficult to assess its effects on genetic alterations without interference of other compounding factors. Therefore, present study was undertaken in mice to verify the ability of raw areca-nut (RAN) to induce cancer and to monitor the expression of certain genes involved in carcinogenesis. This study was not intended to isolate any active ingredients from the RAN and to look its action. Three groups of mice (n = 25 in each) were taken and used at different time-points for different experimental analysis. The other three groups of mice (n = 15 in each) were considered for tumor induction studies. In each set, two groups were administered RAN-extract ad libitum in drinking water with or without lime. The expression of certain genes was assessed by conventional RT-PCR and immunoblotting. The mice were given the whole RAN-extract with and without lime in order to mimic the human consumption style of RAN. Histological preparation of stomach tissue revealed that RAN induced stomach cancer. A gradual increase in the frequency of precocious anaphase and aneuploid cells was observed in the bone marrow cells with a greater increment following RAN + lime administeration. Levels of p53, Bax, Securin and p65 in esophageal and stomach cells were elevated during early days of RAN exposure while those of different mitotic checkpoint proteins were downregulated. Apoptotic cell death was detected in non-cancerous stomach cells but not in tumor cells which showed overexpression of Bax and absence of PARP. Present study suggested (a) RAN induces stomach cancer, however, presence of lime promoted higher cell transformation

  10. Molecular detection of chromosomal abnormalities in germ and somatic cells of aged male mice

    SciTech Connect

    Lowe, X.; Baulch, J.; Quintana, L.; Ramsey, M.; Breneman, J.; Tucker, J.; Wyrobek, A.; Collins, B.; Allen, J.; Holland, N.

    1994-12-31

    Three cytogenetic methods were applied to eight B6C3F1 male mice aged 22.5 - 30.5mo to determine if advanced age was associated with an elevated risk of producing chromosomally defective germinal and somatic cells; sperm aneuploidy analysis by multi-color fluorescence in situ hybridization for three chromosomes, spermatid micronucleus analysis with anti-kinetochore antibodies, and translocation analysis of somatic metaphases by {open_quotes}painting{close_quotes} for two chromosomes. Eight mice aged 2.4mo served as controls. Sperm aneuploidy was measured by multi-color fluorescence in situ co-hybridization with DNA probes specific for chromosomes X, Y and 8, scoring 10,000 cells per animal. The aged group showed significant 1.5 - 2.0 fold increases in the hyperhaploidy phenotypes X-X-8, Y-Y-8, 8-8-Y, and 8-8-X with the greater effects appearing in animals aged >29mo. The aged group also showed significantly increased frequencies of micronucleated spermatids (2.0 vs 0.4 per 1000; all were kinetochore negative). Analysis of metaphase chromosomes from blood by {open_quotes}painting{close_quotes} of chromosomes 2 and 8 yielded 4 translocation per 858 cell-equivalents in the aged group which was a non-significant elevation over 0/202 in controls. Although interpretation must be cautious due to the small number of animals analyzed, these findings suggest that advanced paternal age may be a risk factor for chromosomal abnormalities of reproductive and somatic importance.

  11. Assessment of molecular genetic detection of chromosome translocations in the differential diagnosis of pediatric sarcomas.

    PubMed

    Dockhorn-Dworniczak, B; Schäfer, K L; Blasius, S; Christiansen, H; Koscielniak, E; Ritter, J; Winkelmann, W; Jürgens, H; Böcker, W

    1997-01-01

    Recent studies have shown that many types of soft-tissue sarcomas are characterized by specific chromosomal translocations, which are likely to be of etiologic significance. In order to evaluate their diagnostic impact, a panel of 129 sarcomas comprising 78 Ewing's tumors (ET), 19 rhabdomyosarcomas (RMS), 20 neuroblastomas (NB), 9 synovialsarcomas, 2 esthesioneuroblastomas, and 1 desmoplastic small-round-cell tumor (DSRCT) were analysed for the occurrence of the major recurrent translocations, such as t(11;22)(q24;q12), t(21;22)(q22;q12), t(11;22)(p13;q12), t(2;13)(q35;q14), t(1;13)(p36;q14), and t(X;18)(p11;q11). Nitrogen-frozen tissue material was analysed by means of Reverse Transcription followed by PCR (Polymerase-Chain Reaction) and nested PCR (RT-PCR). Specificity of the PCR products obtained was confirmed by non-isotopic Southern-Blot analysis with gene-specific probes and/or automated direct sequence analysis. 75 ETs have been shown to carry either a t(11;22) or t(21;22) translocation by identification of chimeric EWS-FLI-1 or EWS-ERG gene-fusion transcripts respectively. 3 ETs were lacking EWS/FLI-1 or EWS-ERG fusion products. 2 of these tumors were shown on review to have unusual morphological features for ETs. 8/19 RMS were initially diagnosed as alveolar RMS. These tumours were shown to carry either a t(2;13) translocation exhibiting chimeric PAX3-FKHR fusion transcripts or a t(1;13) translocation with PAX7-FKHR chimeric gene products. One RMS of the embryonal group also carried a t(1;13) translocation. Reevaluation demonstrated a partly alveolar morphology. In 8/9 synovial sarcomas a t(X;18) translocation was identified. Expression of a EWS-WTI gene-fusion product associated with a t(11;22) translocation was found in the DSRCT. None of these rearrangements were detected in the NBs and 2 esthesioneuroblastomas. Our results support the concept that the major recurrent translocations are histogenetically specific for a subset of sarcomas. Thus, the

  12. Reversal of DNA methylation with 5-azacytidine alters chromosome replication patterns in human lymphocyte and fibroblast cultures.

    PubMed

    Shafer, D A; Priest, J H

    1984-05-01

    Prior studies demonstrated that developmental or induced methylation of DNA can inactivate associated gene loci. Such DNA methylation can be reversed and specific genes reactivated by treatment with 5-azacytidine (5- azaC ). The present cytogenetic studies using replication banding methods show that 5- azaC treatment also results in an increase or decrease in replication staining at one or more band locations in human lymphocyte and fibroblast chromosomes. New replication band locations are not formed. These changes in replication staining, which reflect changes in timing of replication, are different between these two tissues. However, in both tissues, the delayed onset of replication in the heterocyclic, inactive X is shortened by 5- azaC . A correlation is thus suggested between the induced temporal change to earlier DNA replication, and induced hypomethylation and gene activation. The temporal effect on chromosome replication in 5- azaC -treated cells depends on the portion of the S-period studied. Toward the beginning of S, early-replication patterns are increased in both lymphocytes and fibroblasts. Toward the end of S, late-replication patterns are increased only in lymphocytes, suggesting a differential effect of 5- azaC in: (1) early-vs. late-S, and (2) lymphocytes vs. fibroblasts. Generally, 5- azaC has its greatest effect on the inactive chromosome regions that are typically late-replicating prior to 5- azaC treatment. These observed changes in replication band staining suggest that DNA methylation may modify regional groups of genes in concert.

  13. Aneuploidy in spermatozoa detected by FISH. Comparison with sperm chromosome data obtained via hamster system

    SciTech Connect

    Estop, A.M.; Van Kirk, V.; Cieply, K.

    1994-09-01

    Fluorescence in-situ hybridization (FISH) with two-color and cocktail DNA probes was used to assess the rates of aneuploidy for the X,Y and 18 chromosomes in 3 male donors. (Experiment 1). These individuals had previously been studied with the hamster system and published. Experiment 2 was designed in order to compare aneuploidy rates for chromosome 18 in donor 2 in conjunction with chromosome 6 and 12 as an internal control. (1) Aneuploidy for the sex chromosomes in the hamster system was 0.5 for Donor 1 and 0.7 (3) which was very similar to 0.49 (1) and 0.41 (3) found in this experiment. However, Donor 2 showed a lower rate of sex non-disjunction with this system: 0.18 vs. 0.7 with the hamster system. (2) Diploidy rates are in the same ranges in experiments 1 and 2. (3) If autosome aneuploidy rates are extrapolated to 22 chromosomes, the following values are found: Donor 1:2.42 (vs. 2.0 in the hamster system); donor 3:2.2 (vs. 1.34 with the hamster system) and donor 2:1.32 which is lower than 4.32 found with the hamster system. More data needs to be collected on the use of FISH for this study of aneuploides in sperm cells and attention needs to be paid to the different types of probes used for validation of results.

  14. Chromosomal and Multifactorial Genetic Disorders with Oral Manifestations

    PubMed Central

    Patil, Shankargouda; Rao, Roopa S; Majumdar, Barnali

    2014-01-01

    The chromosomal disorders are individually rare, but collectively they are common whereas the multifactorial disorders are the most common form of genetic disorders. The chromosomal anomalies typically arise from alterations in the DNA containing chromosomal regions and can be reliably detected by karyotype analysis, whereas the multifactorial disorders demonstrate multi-gene as well as environmental interactions. Both the chromosomal and multifactorial disorders may manifest signs and symptoms such as a combination of birth defects, physical disabilities, challenging behavior and certain craniofacial defects as well, the knowledge of which can aid in a better patient management in everyday practice of dentistry. PMID:25395808

  15. Rapid and simultaneous detection of linear chromosome and large plasmids in Proteobacteria.

    PubMed

    Ramírez-Bahena, Martha Helena; Nesme, Xavier; Muller, Daniel

    2012-12-01

    Bacterial genomic architectures are complex and play important roles in genome evolution. While most bacterial lineage genomes contain a single chromosome often accompanied by plasmids, more and more genomes are described as harboring a linear or a second chromosome. Thus, the development of bacterial genomics leads to the study of bacterial genome architectures. In order to avoid laborious techniques combining several methods, we developed an original plug-free pulsed field gel electrophoresis procedure, that enabled us to co-characterize reproducibly the presence of linear chromosomes and plasmids ranging between 30 kb and 2000 kb in various proteobacterial lineages. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Nonrandom distribution of methotrexate-induced aberrations on human chromosomes. Detection of further folic acid sensitive fragile sites.

    PubMed

    Barbi, G; Steinbach, P; Vogel, W

    1984-01-01

    Eleven folic acid sensitive fragile sites (3p14, 7p13, 7q31.1, 7q32, 9q32, 11p13, 14q23, 15q22, 16q23, Xp22.2, Xq22) were detected in one individual, eight of them previously unknown. These sites seem to bear each its specific sensitivity to folic acid deficiency. Six of the sites were observed simultaneously on both homologous chromosomes in at least one cell. Each of these 11 sites was also found in at least one among 12 individuals further examined. Some of these individuals showed six of these 11 sites. The fragile site 3p14 was detected in all individuals examined. The homologous sites 3p14 of one individual differed from each other in their frequency of lesions induced by methotrexate as well as fluorodeoxyuridine. This observation suggests that folic acid sensitivity is a property inherent in the chromatin of an individual chromosome at the site involved in fragility. This property seems to be responsible for the nonrandom fragility at that site and also for the individual sensitivity of each chromosomal site.

  17. The gene-reduction effect of chromosomal losses detected in gastric cancers

    PubMed Central

    2010-01-01

    Background The level of loss of heterozygosity (LOH) that reduces a gene dose and exerts a cell-adverse effect is known to be a parameter for the genetic staging of gastric cancers. This study investigated if the cell-adverse effect induced with the gene reduction was a rate-limiting factor for the LOH events in two distinct histologic types of gastric cancers, the diffuse- and intestinal-types. Methods The pathologic specimens obtained from 145 gastric cancer patients were examined for the level of LOH using 40 microsatellite markers on eight cancer-associated chromosomes (3p, 4p, 5q, 8p, 9p, 13q, 17p and 18q). Results Most of the cancer-associated chromosomes were found to belong to the gene-poor chromosomes and to contain a few stomach-specific genes that were highly expressed. A baseline-level LOH involving one or no chromosome was frequent in diffuse-type gastric cancers. The chromosome 17 containing a relatively high density of genes was commonly lost in intestinal-type cancers but not in diffuse-type cancers. A high-level LOH involving four or more chromosomes tended to be frequent in the gastric cancers with intestinal and mixed differentiation. Disease relapse was common for gastric cancers with high-level LOH through both the hematogenous (38%) and non-hematogenous (36%) routes, and for the baseline-level LOH cases through the non-hematogenous route (67%). Conclusions The cell-adverse effect of gene reduction is more tolerated in intestinal-type gastric cancers than in diffuse-type cancers, and the loss of high-dose genes is associated with hematogenous metastasis. PMID:21092121

  18. Spectral Karyotyping Detects Chromosome Damage in Bronchial Cells of Smokers and Patients with Cancer

    PubMed Central

    Varella-Garcia, Marileila; Chen, Lin; Powell, Roger L.; Hirsch, Fred R.; Kennedy, Timothy C.; Keith, Robert; Miller, York E.; Mitchell, John D.; Franklin, Wilbur A.

    2007-01-01

    Rationale: Lung cancer is a multistep process that is preceded and often accompanied by molecular cytogenetic lesions in benign bronchial epithelium, the precise character, extent and timing of which are not well defined. Objectives: In this study we comprehensively defined molecular cytogenetic changes in bronchial cells that may precede lung carcinoma using spectral karyotyping (SKY). Methods: SKY was applied to cultured benign bronchial cells from 43 high-risk smokers without carcinoma, 14 patients with concurrent lung carcinoma, and 14 never-smoker healthy volunteers. Measurements and Main Results: The proportion of cells displaying numeric or structural anomalies/total number of metaphase cells was calculated for each case and was referred to as the chromosomal abnormality index. Mean chromosomal abnormality indices were 15.8, 10.1, and 0.7% for patients with cancer, high-risk smokers, and never-smokers, respectively. Clonal abnormalities were found in 17 (40%) of the high-risk smokers without carcinoma and 7 (50%) of the patients with carcinoma, but in none of 14 (0%) never-smokers. Chromosomal gains observed by SKY were confirmed in interphase cultured cells or paraffin sections of biopsy specimens by fluorescence in situ hybridization in 11 of 13 cases for which appropriate probes were available. In 6 of 57 high-risk patients or those with carcinoma, identical clonal abnormalities were dispersed at multiple bronchial sites and were admixed with nonclonal cells. Conclusions: Clonal and single-cell chromosomal abnormalities are frequent in benign bronchial epithelium during lung carcinogenesis, indicating that chromosomal missegregation and other chromosomal rearrangements occur before overt malignancy. PMID:17600274

  19. [Combined G-banded karyotyping and multiplex ligation-dependent probe amplification for the detection of chromosomal abnormalities in fetuses with congenital heart defects].

    PubMed

    Liu, Yang; Xie, Jiansheng; Geng, Qian; Xu, Zhiyong; Wu, Weiqin; Luo, Fuwei; Li, Suli; Wang, Qin; Chen, Wubin; Tan, Hongxi; Zhang, Hu

    2017-02-10

    To assess the value of G-banded karyotyping in combination with multiplex ligation-dependent probe amplification (MLPA) as a tool for the detection of chromosomal abnormalities in fetuses with congenital heart defects. The combined method was used to analyze 104 fetuses with heart malformations identified by ultrasonography. Abnormal findings were confirmed with chromosomal microarray analysis (CMA). Nineteen (18%) fetuses were found to harbor chromosomal aberrations by G-banded karyotyping and MLPA. For 93 cases, CMA has detected abnormalities in 14 cases including 10 pathogenic copy number variations (CNVs) and 4 CNVs of uncertain significance (VOUS). MLPA was able to detect all of the pathogenic CNVs and 1 VOUS CNV. Combined use of G-banded karyotyping and MLPA is a rapid, low-cost and effective method to detect chromosomal abnormalities in fetuses with various heart malformations.

  20. Hi-C as a tool for precise detection and characterisation of chromosomal rearrangements and copy number variation in human tumours.

    PubMed

    Harewood, Louise; Kishore, Kamal; Eldridge, Matthew D; Wingett, Steven; Pearson, Danita; Schoenfelder, Stefan; Collins, V Peter; Fraser, Peter

    2017-06-27

    Chromosomal rearrangements occur constitutionally in the general population and somatically in the majority of cancers. Detection of balanced rearrangements, such as reciprocal translocations and inversions, is troublesome, which is particularly detrimental in oncology where rearrangements play diagnostic and prognostic roles. Here we describe the use of Hi-C as a tool for detection of both balanced and unbalanced chromosomal rearrangements in primary human tumour samples, with the potential to define chromosome breakpoints to bp resolution. In addition, we show copy number profiles can also be obtained from the same data, all at a significantly lower cost than standard sequencing approaches.

  1. Evaluation of chromosomal alteration in electrical workers occupationally exposed to low frequency of electro magnetic field (EMFs) in Coimbatore population, India.

    PubMed

    Balamuralikrishnan, Balasubramanian; Balachandar, Vellingiri; Kumar, Shanmugam Suresh; Stalin, Nattan; Varsha, Prakash; Devi, Subramaniam Mohana; Arun, Meyyazhagan; Manikantan, Pappuswamy; Venkatesan, Chinnakulandhai; Sasikala, Keshavarao; Dharwadkar, Shahnaz N

    2012-01-01

    Extremely low frequency electro magnetic fields (EMFs) have been classified as possibly carcinogenic to humans by the International Agency for Research on Cancer. An increased number of chromosomal alterations in peripheral lymphocytes are correlated with elevated incidence of cancer. The aim of the present study was to assess occupationally induced chromosomal damage in EMF workers exposed to low levels of radiation. We used conventional metaphase chromosome aberration (CA) analysis and the micronucleus (MN) assay as biological indicators of non ionizing radiation exposure. In the present study totally 70 subjects were selected including 50 exposed and 20 controls. Informed written consent was obtained from all participants and the study was performed in accordance with the Declaration of Helsinki and the approval of the local ethical committee. A higher degree of CA and MN was observed in exposed subjects compared to controls, the frequency of CA being significantly enhanced with long years of exposure (P<0.05). Moreover increase in CA and MN with age was noted in both exposed subjects and controls, but was significantly greater in the former. The results of this study demonstrated that a significant induction of cytogenetic damage in peripheral lymphocytes of workers occupationally exposed to EMFs in electric transformer and distribution stations. In conclusion, our findings suggest that EMFs possess genotoxic capability, as measured by CA and MN assays; CA analysis appeared more sensitive than other cytogenetic end-points. It can be concluded that chronic occupational exposure to EMFs may lead to an increased risk of genetic damage among electrical workers.

  2. Analysis of chromosome 17p13 (p53 locus) alterations in gastric carcinoma cells by dual-color fluorescence in situ hybridization.

    PubMed

    Kobayashi, M; Kawashima, A; Mai, M; Ooi, A

    1996-11-01

    Chromosome 17 and p53 gene locus alterations were determined on 67 gastric carcinomas by dual-color fluorescence in situ hybridization, using probes for centromere 17 and the 17p13.1 (p53 locus). The results were compared with loss of heterozygosity (LOH) at 17p13.3, direct sequencing of exons 5 to 9 of p53, and nuclear overexpression of p53 protein. Deletion of p53 was found in 26 of 67 tumors (39%). All 26 also showed LOH at 17p13.3, frequently overexpressed p53 protein, and had polysomy 17. The functional loss of p53 gene in these tumors, 85% of which were of intestinal type, appears to be caused by both deletion of 17p13.1 and missense mutation of the remaining allele. There were 9 tumors that had neither deletion nor LOH but had a large proportion of cancer cells that overexpressed p53 election. Despite evidence of LOH, there was no p53 deletion in 11 tumors. Finally, 21 tumors, mostly of diffuse type, showed neither deletions, LOH, nor p53 overexpression. Our data suggest that in gastric cancer, deletion of 17p is principally responsible for the allelic loss at the p53 gene and that analysis of deletions by the dual-color fluorescence in situ hybridization is a sensitive and useful approach to clarify chromosomal aberrations.

  3. Y chromosome and HIV DNA Detection in Vaginal Swabs as Biomarkers of Semen and HIV Exposure in Women

    PubMed Central

    Penrose, Kerri J.; Richardson, Barbra A.; Besson, Guillaume; Dezzutti, Charlene S.; Herold, Betsy C.; Abdool Karim, Salim S.; Mellors, John; Parikh, Urvi M.

    2014-01-01

    Background The inability to quantify sexual exposure to HIV limits the power of HIV prevention trials of vaccines, microbicides and pre-exposure prophylaxis (PrEP) in women. We investigated detection of HIV-1 and Y chromosomal (Yc) DNA in vaginal swabs from 83 participants in the HPTN 035 microbicide trial as biomarkers of HIV exposure and unprotected sexual activity. Methods 143 vaginal swabs from 85 women were evaluated for the presence of Y chromosomal DNA (Quantifiler Duo DNA quantification kit, Applied Biosystems) and total HIV-1 DNA (single copy in-house qPCR assay). Y DNA detection was paired with self-reported behavioral data with regard to recent coitus (≤ 1 week prior to collection) and condom usage (100% vs. <100% compliance). Results Yc DNA was detected in 62/143 (43%) swabs. For the 126 visits at which both behavioral data and swabs were collected, Yc DNA was significantly more frequent in women reporting <100% condom usage (OR 10.69; 95% confidence interval: 2.27 – 50.32; p=0.003). Notably, 27 of 83 (33%) swabs from women reporting 100% condom usage were positive for Yc DNA. HIV DNA was only detected in swabs collected post-seroconversion. Conclusions The use of Yc DNA in HIV prevention trials could reliably identify sub-groups of women who have unprotected sexual activity and could provide valuable exposure-based estimates of efficacy. PMID:25299415

  4. Characterizing patients with multiple chromosomal aberrations detected by FISH in chronic lymphocytic leukemia.

    PubMed

    González-Gascón Y Marín, Isabel; Hernández-Sanchez, María; Rodríguez-Vicente, Ana Eugenia; Puiggros, Anna; Collado, Rosa; Luño, Elisa; González, Teresa; Ruiz-Xivillé, Neus; Ortega, Margarita; Gimeno, Eva; Muñoz, Carolina; Infante, Maria Stefania; Delgado, Julio; Vargas, María Teresa; González, Marcos; Bosch, Francesc; Espinet, Blanca; Hernández-Rivas, Jesús María; Hernández, José Ángel

    2017-07-21

    We analyzed the features of chronic lymphocytic leukemia (CLL) with multiple abnormalities (MA) detected by FISH. A local database including 2095 CLL cases was used and 323 with MA (15.4%) were selected. MA was defined by the presence of two or more alterations (deletions of 13q14 (13q-), 11q22 (11q-), 17p13 (17p-) or trisomy 12 (+12)). The combination of 13q- with 11q- and 13q- with 17p-, accounted for 58.2% of the series, in contrast to 11q- with 17p- (3.7%). Patients carrying MA since diagnosis presented a short time to first therapy(TTFT) (27 months) and overall survival (OS) (76 months). The combinations including 17p- had a shorter OS (58 months) than the ones without 17p- (not reached, p = .002). Patients with a complex-FISH were the ones with worse OS (34 months). MA imply poor prognosis when they emerge at diagnosis, probably due to the high incidence of bad prognosis markers, which may be a reflection of a more complex karyotype.

  5. Detection and chromosomal assignment of SV40-DNA integration in Chinese hamster cell lines by chromosome sorting and dot blot hybridization.

    PubMed

    Hutter, K J; Klefenz, H; Goerttler, K

    1990-01-01

    A combination of cytometric (chromosome sorting), molecular (dot blot hybridization using radio-active and/or biotinylated DNA probes) and cytogenetic (G-banding) evaluation is described which allows the rapid identification of single copy and repetitive viral integrates and their assignment to chromosome groups or even individual chromosomes. In the case of Chinese hamster cell line CO 631 it could be demonstrated that SV40 DNA was solely integrated into a submetacentric marker chromosome. Such a cytometric/molecular/cytogenetic "identogram" may prove to be a useful tool in many areas of cell and tumor biology. Furthermore, amounts of chromosomes sufficient for analysis as well as subsequent cloning experiments can be accumulated.

  6. Using a combination of MLPA kits to detect chromosomal imbalances in patients with multiple congenital anomalies and mental retardation is a valuable choice for developing countries.

    PubMed

    Jehee, Fernanda Sarquis; Takamori, Jean Tetsuo; Medeiros, Paula F Vasconcelos; Pordeus, Ana Carolina B; Latini, Flavia Roche M; Bertola, Débora Romeo; Kim, Chong Ae; Passos-Bueno, Maria Rita

    2011-01-01

    Conventional karyotyping detects anomalies in 3-15% of patients with multiple congenital anomalies and mental retardation (MCA/MR). Whole-genome array screening (WGAS) has been consistently suggested as the first choice diagnostic test for this group of patients, but it is very costly for large-scale use in developing countries. We evaluated the use of a combination of Multiplex Ligation-dependent Probe Amplification (MLPA) kits to increase the detection rate of chromosomal abnormalities in MCA/MR patients. We screened 261 MCA/MR patients with two subtelomeric and one microdeletion kits. This would theoretically detect up to 70% of all submicroscopic abnormalities. Additionally we scored the de Vries score for 209 patients in an effort to find a suitable cut-off for MLPA screening. Our results reveal that chromosomal abnormalities were present in 87 (33.3%) patients, but only 57 (21.8%) were considered causative. Karyotyping detected 15 abnormalities (6.9%), while MLPA identified 54 (20.7%). Our combined MLPA screening raised the total detection number of pathogenic imbalances more than three times when compared to conventional karyotyping. We also show that using the de Vries score as a cut-off for this screening would only be suitable under financial restrictions. A decision analytic model was constructed with three possible strategies: karyotype, karyotype + MLPA and karyotype + WGAS. Karyotype + MLPA strategy detected anomalies in 19.8% of cases which account for 76.45% of the expected yield for karyotype + WGAS. Incremental Cost Effectiveness Ratio (ICER) of MLPA is three times lower than that of WGAS, which means that, for the same costs, we have three additional diagnoses with MLPA but only one with WGAS. We list all causative alterations found, including rare findings, such as reciprocal duplications of regions deleted in Sotos and Williams-Beuren syndromes. We also describe imbalances that were considered polymorphisms or rare variants, such as the new SNP

  7. A first look at women's perspectives on noninvasive prenatal testing to detect sex chromosome aneuploidies and microdeletion syndromes.

    PubMed

    Agatisa, Patricia K; Mercer, Mary Beth; Leek, Angela C; Smith, Marissa B; Philipson, Elliot; Farrell, Ruth M

    2015-07-01

    The aim of this study is to explore women's opinions about the use of noninvasive prenatal testing (NIPT) to assess the risk of sex chromosome aneuploidies and microdeletion syndromes. Focus groups were conducted with women who were currently pregnant or had recently delivered. Qualitative analysis using interpretive description was used to generate study findings. Thirty-one women (mean age 32.4 years) participated in the focus groups. Participants were unfamiliar with sex chromosome aneuploidies but expressed support for the use of NIPT to detect these conditions. Participants were uncertain about the utility and actionability of receiving information about microdeletion syndromes with variable or unknown phenotypic expression. Participants voiced their desire to be informed of all conditions assessed by NIPT prior to testing. They considered clinicians to be the key provider of such information, although stated that patients have a responsibility to be knowledgeable prior to testing in order to support informed decision making. The use of NIPT to identify sex chromosome aneuploidies and microdeletion syndromes will introduce new challenges for clinicians to ensure pregnant women have the information and resources to make informed choices about NIPT when used for these conditions. © 2015 John Wiley & Sons, Ltd.

  8. Detection of hypomethylation syndrome among patients with epigenetic alterations at the GNAS locus.

    PubMed

    Perez-Nanclares, Gustavo; Romanelli, Valeria; Mayo, Sonia; Garin, Intza; Zazo, Celia; Fernandez-Rebollo, Eduardo; Martínez, Francisco; Lapunzina, Pablo; de Nanclares, Guiomar Pérez

    2012-06-01

    Genomic imprinting is the modification of the genome so that genes from only one (rather than two) of the parental alleles are expressed. The mechanism underlying imprinting is epigenetic, occurring via changes in DNA methylation and histone modifications rather than through alterations in the DNA sequence. To date, nine different imprinting disorders have been clinically and genetically identified and a considerable research effort has been focused on determining the cause of the corresponding methylation defects. Our objective was to identify multilocus imprinting defects and characterize any mutations in trans-acting genes in patients with pseudohypoparathyroidism (PHP) caused by epigenetic alterations at GNAS locus. We have investigated multilocus imprinting defects in 22 PHP patients with aberrant methylation at the GNAS locus not due to previously described deletions or to paternal uniparental disomy (UPD) of chromosome 20. We found that, in contrast to what has been described in growth disorders, multilocus hypomethylation is an uncommon event in PHP patients. We were also unable to identify any genetic alteration causative of the epigenetic defects in the currently known methylation regulatory genes. Our work suggests that a trans-acting gene regulating the establishment or maintenance of imprinting at GNAS locus, if it exists, should be specific to PHP cases caused by epigenetic defects at GNAS.

  9. A role for maternal serum screening in detecting chromosomal abnormalities in fetuses with isolated choroid plexus cysts: a prospective multicentre study.

    PubMed

    Brown, T; Kliewer, M A; Hertzberg, B S; Ruiz, C; Stamper, T H; Rosnes, J; Lucas, A; Wright, L N; Chescheir, N C; Farmer, L; Jordan, S; Kay, H H

    1999-05-01

    A prospective multicentre study was performed to identify patients with fetal choroid plexus cysts and examine the association between choroid plexus cysts and chromosome abnormalities in the context of variables such as maternal age, serum triple-screen results, race, other prenatally-identified fetal anomalies and cyst characteristics. A total of 18 437 scans were performed in 5 centres and 257 fetuses were identified with choroid plexus cysts. Outcome was available on 250 patients, and of these, chromosomal abnormalities were detected in a total of 13 (5.2 per cent) fetuses. 26 patients in the group had additional ultrasound abnormalities, and 8 of these had fetal chromosome abnormalities. Among the 224 patients with isolated choroid plexus cysts, 5 (2.2 per cent) were found to have chromosomal abnormalities. All cases with identified chromosomal abnormalities were associated with an additional risk factor, such as other ultrasound findings, advanced maternal age or abnormal maternal serum triple-screen results.

  10. The Williams syndrome chromosome 7q11.23 hemideletion confers hypersocial, anxious personality coupled with altered insula structure and function.

    PubMed

    Jabbi, Mbemba; Kippenhan, J Shane; Kohn, Philip; Marenco, Stefano; Mervis, Carolyn B; Morris, Colleen A; Meyer-Lindenberg, Andreas; Berman, Karen Faith

    2012-04-03

    Although it is widely accepted that genes can influence complex behavioral traits such as human temperament, the underlying neurogenetic mechanisms remain unclear. Williams syndrome (WS), a rare disorder caused by a hemizygous deletion on chromosome 7q11.23, including genes important for neuronal migration and maturation (LIMK1 and CLIP2), is typified by a remarkable hypersocial but anxious personality and offers a unique opportunity to investigate this open issue. Based on the documented role of the insula in mediating emotional response tendencies and personality, we used multimodal imaging to characterize this region in WS and found convergent anomalies: an overall decrease in dorsal anterior insula (AI) gray-matter volume along with locally increased volume in the right ventral AI; compromised white-matter integrity of the uncinate fasciculus connecting the insula with the amygdala and orbitofrontal cortex; altered regional cerebral blood flow in a pattern reminiscent of the observed gray-matter alterations (i.e., widespread reductions in dorsal AI accompanied by locally increased regional cerebral blood flow in the right ventral AI); and disturbed neurofunctional interactions between the AI and limbic regions. Moreover, these genetically determined alterations of AI structure and function predicted the degree to which the atypical WS personality profile was expressed in participants with the syndrome. The AI's rich anatomical connectivity, its transmodal properties, and its involvement in the behaviors affected in WS make the observed genetically determined insular circuitry perturbations and their association with WS personality a striking demonstration of the means by which neural systems can serve as the interface between genetic variability and alterations in complex behavioral traits.

  11. Progression from colorectal adenoma to carcinoma is associated with non-random chromosomal gains as detected by comparative genomic hybridisation.

    PubMed Central

    Meijer, G A; Hermsen, M A; Baak, J P; van Diest, P J; Meuwissen, S G; Beliën, J A; Hoovers, J M; Joenje, H; Snijders, P J; Walboomers, J M

    1998-01-01

    AIMS: Chromosomal gains and losses were surveyed by comparative genomic hybridisation (CGH) in a series of colorectal adenomas and carcinomas, in search of high risk genomic changes involved in colorectal carcinogenesis. METHODS: Nine colorectal adenomas and 14 carcinomas were analysed by CGH, and DNA ploidy was assessed with both flow and image cytometry. RESULTS: In the nine adenomas analysed, an average of 6.6 (range 1 to 11) chromosomal aberrations were identified. In the 14 carcinomas an average of 11.9 (range 5 to 17) events were found per tumour. In the adenomas the number of gains and losses was in balance (3.6 v 3.0) while in carcinomas gains occurred more often than losses (8.2 v 3.7). Frequent gains involved 13q, 7p, 8q, and 20q, whereas losses most often occurred at 18q, 4q, and 8p. Gains of 13q, 8q, and 20q, and loss of 18q occurred more often in carcinomas than in adenomas (p = 0.005, p = 0.05, p = 0.05, and p = 0.02, respectively). Aneuploid tumours showed more gains than losses (mean 9.3 v 4.9, p = 0.02), in contrast to diploid tumours where gains and losses were nearly balanced (mean 3.1 v 4.1, p = 0.5). CONCLUSIONS: The most striking difference between chromosomal aberrations in colorectal adenomas and carcinomas, as detected by CGH, is an increased number of chromosomal gains that show a nonrandom distribution. Gains of 13q and also of 20q and 8q seem especially to be involved in the progression of adenomas to carcinomas, possibly owing to low level overexpression of oncogenes at these loci. Images PMID:10070331

  12. Ordered-subsets linkage analysis detects novel Alzheimer disease loci on chromosomes 2q34 and 15q22.

    PubMed

    Scott, William K; Hauser, Elizabeth R; Schmechel, Donald E; Welsh-Bohmer, Kathleen A; Small, Gary W; Roses, Allen D; Saunders, Ann M; Gilbert, John R; Vance, Jeffery M; Haines, Jonathan L; Pericak-Vance, Margaret A

    2003-11-01

    Alzheimer disease (AD) is a complex disorder characterized by a wide range, within and between families, of ages at onset of symptoms. Consideration of age at onset as a covariate in genetic-linkage studies may reduce genetic heterogeneity and increase statistical power. Ordered-subsets analysis includes continuous covariates in linkage analysis by rank ordering families by a covariate and summing LOD scores to find a subset giving a significantly increased LOD score relative to the overall sample. We have analyzed data from 336 markers in 437 multiplex (>/=2 sampled individuals with AD) families included in a recent genomic screen for AD loci. To identify genetic heterogeneity by age at onset, families were ordered by increasing and decreasing mean and minimum ages at onset. Chromosomewide significance of increases in the LOD score in subsets relative to the overall sample was assessed by permutation. A statistically significant increase in the nonparametric multipoint LOD score was observed on chromosome 2q34, with a peak LOD score of 3.2 at D2S2944 (P=.008) in 31 families with a minimum age at onset between 50 and 60 years. The LOD score in the chromosome 9p region previously linked to AD increased to 4.6 at D9S741 (P=.01) in 334 families with minimum age at onset between 60 and 75 years. LOD scores were also significantly increased on chromosome 15q22: a peak LOD score of 2.8 (P=.0004) was detected at D15S1507 (60 cM) in 38 families with minimum age at onset >/=79 years, and a peak LOD score of 3.1 (P=.0006) was obtained at D15S153 (62 cM) in 43 families with mean age at onset >80 years. Thirty-one families were contained in both 15q22 subsets, indicating that these results are likely detecting the same locus. There is little overlap in these subsets, underscoring the utility of age at onset as a marker of genetic heterogeneity. These results indicate that linkage to chromosome 9p is strongest in late-onset AD and that regions on chromosome 2q34 and 15q22 are

  13. Solar activity cycle and the incidence of foetal chromosome abnormalities detected at prenatal diagnosis

    NASA Astrophysics Data System (ADS)

    Halpern, Gabrielle J.; Stoupel, Eliahu G.; Barkai, Gad; Chaki, Rina; Legum, Cyril; Fejgin, Moshe D.; Shohat, Mordechai

    1995-06-01

    We studied 2001 foetuses during the period of minimal solar activity of solar cycle 21 and 2265 foetuses during the period of maximal solar activity of solar cycle 22, in all women aged 37 years and over who underwent free prenatal diagnosis in four hospitals in the greater Tel Aviv area. There were no significant differences in the total incidence of chromosomal abnormalities or of trisomy between the two periods (2.15% and 1.8% versus 2.34% and 2.12%, respectively). However, the trend of excessive incidence of chromosomal abnormalities in the period of maximal solar activity suggests that a prospective study in a large population would be required to rule out any possible effect of extreme solar activity.

  14. Frequencies of complex chromosome exchange aberrations induced by 238Pu alpha-particles and detected by fluorescence in situ hybridization using single chromosome-specific probes.

    PubMed

    Griffin, C S; Marsden, S J; Stevens, D L; Simpson, P; Savage, J R

    1995-04-01

    We undertook an analysis of chromosome-type exchange aberrations induced by alpha-particles using fluorescence in situ hybridization (FISH) with whole chromosome-specific probes for human chromosomes 1 or 4, together with a pan-centromeric probe. Contact-inhibited primary human fibroblasts (in G1) were irradiated with 0.41-1.00 Gy 238Pu alpha-particles and aberrations were analysed at the next mitosis following a single chromosome paint. Exchange and aberration painting patterns were classified according to Savage and Simpson (1994a). Of exchange aberrations, 38-47% were found to be complex derived, i.e. resulting from three or more breaks in two or more chromosomes, and the variation with dose was minimal. The class of complex aberrations most frequently observed were insertions, derived from a minimum of three breaks in two chromosomes. There was also an elevated frequency of rings. The high level of complex aberrations observed after alpha-particle irradiation indicates that, when chromosome domains are traversed by high linear energy transfer alpha-particle tracks, there is an enhanced probability of production of multiple localized double-strand breaks leading to more complicated interactions.

  15. Identification of FISH biomarkers to detect chromosome abnormalities associated with prostate adenocarcinoma in tumour and field effect environment

    PubMed Central

    2014-01-01

    Background To reduce sampling error associated with cancer detection in prostate needle biopsies, we explored the possibility of using fluorescence in situ hybridisation (FISH) to detect chromosomal abnormalities in the histologically benign prostate tissue from patients with adenocarcinoma of prostate. Methods Tumour specimens from 33 radical prostatectomy (RP) cases, histologically benign tissue from 17 of the 33 RP cases, and 26 benign prostatic hyperplasia (BPH) control cases were evaluated with Locus Specific Identifier (LSI) probes MYC (8q24), LPL (8p21.22), and PTEN (10q23), as well as with centromere enumerator probes CEP8, CEP10, and CEP7. A distribution of FISH signals in the tumour and histologically benign adjacent tissue was compared to that in BPH specimens using receiver operating characteristic curve analysis. Results The combination of MYC gain, CEP8 Abnormal, PTEN loss or chromosome 7 aneusomy was positive in the tumour area of all of the 33 specimens from patients with adenocarcinomas, and in 88% of adjacent histologically benign regions (15 out of 17) but in only 15% (4 out of 26) of the benign prostatic hyperplasia control specimens. Conclusions A panel of FISH markers may allow detection of genomic abnormalities that associate with adenocarcinoma in the field adjacent to and surrounding the tumour, and thus could potentially indicate the presence of cancer in the specimen even if the cancer focus itself was missed by biopsy and histology review. PMID:24568597

  16. Noninvasive prenatal detection of sex chromosomal aneuploidies by sequencing circulating cell-free DNA from maternal plasma.

    PubMed

    Mazloom, Amin R; Džakula, Željko; Oeth, Paul; Wang, Huiquan; Jensen, Taylor; Tynan, John; McCullough, Ron; Saldivar, Juan-Sebastian; Ehrich, Mathias; van den Boom, Dirk; Bombard, Allan T; Maeder, Margo; McLennan, Graham; Meschino, Wendy; Palomaki, Glenn E; Canick, Jacob A; Deciu, Cosmin

    2013-06-01

    Whole-genome sequencing of circulating cell free (ccf) DNA from maternal plasma has enabled noninvasive prenatal testing for common autosomal aneuploidies. The purpose of this study was to extend the detection to include common sex chromosome aneuploidies (SCAs): [47,XXX], [45,X], [47,XXY], and [47,XYY] syndromes. Massively parallel sequencing was performed on ccf DNA isolated from the plasma of 1564 pregnant women with known fetal karyotype. A classification algorithm for SCA detection was constructed and trained on this cohort. Another study of 411 maternal samples from women with blinded-to-laboratory fetal karyotypes was then performed to determine the accuracy of the classification algorithm. In the training cohort, the new algorithm had a detection rate (DR) of 100% (95%CI: 82.3%, 100%), a false positive rate (FPR) of 0.1% (95%CI: 0%, 0.3%), and nonreportable rate of 6% (95%CI: 4.9%, 7.4%) for SCA determination. The blinded validation yielded similar results: DR of 96.2% (95%CI: 78.4%, 99.8%), FPR of 0.3% (95%CI: 0%, 1.8%), and nonreportable rate of 5% (95%CI: 3.2%, 7.7%) for SCA determination Noninvasive prenatal identification of the most common sex chromosome aneuploidies is possible using ccf DNA and massively parallel sequencing with a high DR and a low FPR. © 2013 John Wiley & Sons, Ltd.

  17. Detection of single copy sequences using BAC-FISH and C-PRINS techniques in sunflower chromosomes.

    PubMed

    Talia, Paola; Greizerstein, Eduardo J; Hopp, H Esteban; Paniego, Norma; Poggio, Lidia; Heinz, Ruth A

    2011-04-01

    Bacterial artificial chromosome-fluorescence in situ hybridization (BAC-FISH) and cycling-primed in situ labeling (C-PRINS) techniques were evaluated for integration of physical and genetic maps of sunflower (Helianthus annuus L.). Single-site SSR markers were selected from three linkage groups of a high-density sunflower genetic map. This selection was based on previously identified QTL associated to S. sclerotiorum. These markers were used to select BACs contaning single copy sequences for BAC-FISH aplication. Blocking of highly dispersed repetitive sunflower sequences reduced unspecific hybridization, and allowed the detection of specific signals for BACs containing SSR markers HA4222 and HA2600, anchored to LG 16 and LG 10, respectively. Single-site FISH signal detection was optimized by adjusting the relative quantity and quality of unlabelled repetitive sequences present in the blocking DNA. The SSR marker ORS1247 anchored to the LG 17 was detected by C-PRINS, which yielded fluorescence signals that were specific and intense. This progress in localizing single-copy sequences using BAC-FISH and indirect C-PRINS strategies in sunflower will facilitate the integration of genetic and physical maps, allowing the identification of chromosomes containing key genes and/or QTL associated to agronomic important traits in sunflower.

  18. Comparison of longitudinal variance components and regression-based approaches for linkage detection on chromosome 17 for systolic blood pressure

    PubMed Central

    Andrade, Mariza de; Olswold, Curtis

    2003-01-01

    We compare two methods to detect genetic linkage by using serial observations of systolic blood pressure in pedigree data from the Framingham Heart Study focusing on chromosome 17. The first method is a variance components (VC) approach that incorporates longitudinal pedigree data, and the second method is a regression-based approach that summarizes all longitudinal measures in one single measure. No evidence of linkage was found either using the VC longitudinal approach or the regression-based approach, except when all time points were used from Cohorts 1 and 2 and only subjects aged 25 and 75 years were included. PMID:14975085

  19. Chromatid Paints: A New Method for Detecting Tumor-Specific Chromosomal Inversions

    DTIC Science & Technology

    1999-10-01

    a neomycin resistance marker (NEO) by homologous recombination in the yeast Saccha- have been established and previously described [14]. romyces...irslSF-C7 mutant knockout in DT40 chicken lymphoblastoid cells, line. This implies that, while XRCC2 and XRCC3 shutdown of RAD51 expression resulted in...Morita, H. Yamada, M. Oshimura, Construction of mouse A9 clones containing a single human chromosome tagged with neomycin -resistance gene via mi- [1

  20. Detection and analysis of chromosomal arsenic resistance in Pseudomonas fluorescens strain MSP3.

    PubMed

    Prithivirajsingh, S; Mishra, S K; Mahadevan, A

    2001-02-09

    Pseudomonas fluorescens MSP3 isolated from sea water was resistant to arsenate. This bacterium harbored no plasmids, indicating that arsenic resistance was chromosomally encoded. The chromosomal DNA from MSP3 when transformed onto Escherichia coli DH5alpha using pBluescript exhibited resistance to sodium arsenate and sodium arsenite. Three clones MSA1, MSA2, and MSI3 containing the ars genes were obtained and further subcloning resulted in three fragments of size 2.2, 2.6, and 2.1 kb for pMSA11, pMSA12, and pMSI13, respectively, which contained the genes arsRBC of the arsenic operon. An efflux mechanism of detoxification was observed which was ATP dependent. The resistance mechanism was encoded from a single operon which consisted of an arsenite inducible repressor that regulates the expression of arsenate reductase (ars C) and inner membrane associated arsenite export system encoded by ars B. The chromosomal operon was cloned, sequenced, and found to consist of three cistrons, named as ars R, ars B, and ars C. Southern hybridization and mating experiments confirmed the functioning of the ars genes in the operon, thereby conferring increased resistance to sodium arsenate and sodium arsenite.

  1. 3D organization of synthetic and scrambled chromosomes.

    PubMed

    Mercy, Guillaume; Mozziconacci, Julien; Scolari, Vittore F; Yang, Kun; Zhao, Guanghou; Thierry, Agnès; Luo, Yisha; Mitchell, Leslie A; Shen, Michael; Shen, Yue; Walker, Roy; Zhang, Weimin; Wu, Yi; Xie, Ze-Xiong; Luo, Zhouqing; Cai, Yizhi; Dai, Junbiao; Yang, Huanming; Yuan, Ying-Jin; Boeke, Jef D; Bader, Joel S; Muller, Héloïse; Koszul, Romain

    2017-03-10

    Although the design of the synthetic yeast genome Sc2.0 is highly conservative with respect to gene content, the deletion of several classes of repeated sequences and the introduction of thousands of designer changes may affect genome organization and potentially alter cellular functions. We report here the Hi-C-determined three-dimensional (3D) conformations of Sc2.0 chromosomes. The absence of repeats leads to a smoother contact pattern and more precisely tractable chromosome conformations, and the large-scale genomic organization is globally unaffected by the presence of synthetic chromosome(s). Two exceptions are synIII, which lacks the silent mating-type cassettes, and synXII, specifically when the ribosomal DNA is moved to another chromosome. We also exploit the contact maps to detect rearrangements induced in SCRaMbLE (synthetic chromosome rearrangement and modification by loxP-mediated evolution) strains.

  2. ABILITY OF THE MALE RAT PUBERTAL ASSAY TO DETECT ENVIRONMENTAL CHEMICALS THAT ALTER THYROID HORMONE HOMEOSTASIS

    EPA Science Inventory

    ABILITY OF THE MALE RAT PUBERTAL ASSAY TO DETECT ENVIRONMENTAL CHEMICALS THAT ALTER THYROID HORMONE HOMEOSTASIS

    Stoker, Tammy E.1; Laws, Susan C.1; Ferrell, Janet M.1; Cooper, Ralph L.1.

    Endocrinology Branch, RTD, NHEERL, ORD, U.S. EPA, RTP, NC, 27711.

    The...

  3. ABILITY OF THE MALE RAT PUBERTAL ASSAY TO DETECT ENVIRONMENTAL CHEMICALS THAT ALTER THYROID HORMONE HOMEOSTASIS

    EPA Science Inventory

    ABILITY OF THE MALE RAT PUBERTAL ASSAY TO DETECT ENVIRONMENTAL CHEMICALS THAT ALTER THYROID HORMONE HOMEOSTASIS

    Stoker, Tammy E.1; Laws, Susan C.1; Ferrell, Janet M.1; Cooper, Ralph L.1.

    Endocrinology Branch, RTD, NHEERL, ORD, U.S. EPA, RTP, NC, 27711.

    The...

  4. ORIENTATION REQUIREMENT TO DETECT MAGNETIC FIELD-INDUCTED ALTERATION OF GAP JUNCTION COMMUNICATION IN EPITHELIAL CELLS

    EPA Science Inventory

    ORIENTATION REQUIREMENT TO DETECT MAGNETIC FIELD-INDUCED ALTERATION OF GAP JUNCTION COMMUNICATION IN EPITHELIAL CELLS.
    OBJECTIVE: We have shown that functional gap junction communication as measured by Lucifer yellow dye transfer (DT) in Clone-9 rat liver epithelial cells, c...

  5. 46 CFR 109.425 - Repairs and alterations: Fire detecting and extinguishing equipment.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 46 Shipping 4 2010-10-01 2010-10-01 false Repairs and alterations: Fire detecting and extinguishing equipment. 109.425 Section 109.425 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) A-MOBILE OFFSHORE DRILLING UNITS OPERATIONS Reports, Notifications, and Records Reports and...

  6. ORIENTATION REQUIREMENT TO DETECT MAGNETIC FIELD-INDUCTED ALTERATION OF GAP JUNCTION COMMUNICATION IN EPITHELIAL CELLS

    EPA Science Inventory

    ORIENTATION REQUIREMENT TO DETECT MAGNETIC FIELD-INDUCED ALTERATION OF GAP JUNCTION COMMUNICATION IN EPITHELIAL CELLS.
    OBJECTIVE: We have shown that functional gap junction communication as measured by Lucifer yellow dye transfer (DT) in Clone-9 rat liver epithelial cells, c...

  7. Y chromosome detection of three-dimensional tissue-engineered skeletal muscle constructs in a syngeneic rat animal model.

    PubMed

    Beier, J P; Kneser, U; Stern-Sträter, J; Stark, G B; Bach, A D

    2004-01-01

    Surgical reconstruction of muscle tissue lost by trauma or tumor ablation is limited by the lack of availability of functional native tissue substitution. Moreover, so far most inherited or acquired muscle diseases are lacking sufficient treatment, because only few alternatives exist to provide functional restoration of lost muscle tissues. Engineering those tissues and transplantation into sites of dysfunction may be an alternative approach and may allow replacement of such damaged or failing skeletal muscle tissues. Techniques attempting reconstruction of some human tissues and organs (tissue engineering) have been introduced into clinical practice recently. One major problem that previous transplantation studies were facing is the ability of detection of transplanted cells after integration. Using the Y chromosome in situ hybridization technique in a syngeneic rat model allows transplantation of cell constructs orthotopically, without manipulation of the cells, with no rejection or immunosuppression being implied, but providing a nondilutable genetic marker to identify transplanted cells. The purpose of our study was to create functional skeletal muscle tissue in vivo using the transplantation of primary myoblasts precultivated within a three-dimensional (3D) fibrin matrix and to determine the fate of the transplanted cells using the Y chromosome detection technique. 3D myoblast cultures were established derived from male donor rats and after 7 days of cultivation we performed an orthotopic transplantation of 3D cell constructs into a created muscle defect within the gracilis muscle of syngeneic female rats. Anti-desmin immunostaining and Y chromosome in situ hybridization indicated the survival and integration of transplanted male myoblasts into the female recipient animal, thus demonstrating the feasibility of this approach in tissue engineering and the research of cell transplantation in general.

  8. Aneuploidy detection for chromosomes 1, X and Y by fluorescence in situ hybridization in human sperm from oligoasthenoteratozoospermic patients

    SciTech Connect

    Pang, M.G.; Zackowski, J.L.; Acosta, A.A.

    1994-09-01

    Oligoasthenoteratozoospermic males (n=15) were investigated for infertility as compared with proven fertile donors. The oligoasthenoteratozoospermic population showed a mean sperm concentration of 9.7 x 10{sup 6}/ml (Range 4.2-19.7), mean motility of 38.5% (Range 10.6-76.8) and morphology (measured by the percentage of normal forms evaluated by strict criteria) with a mean of 3.49% (Range 1.5-5.0). Fluorescence in situ hybridization (FISH) using satellite DNA probes specific for chromosomes 1 (puc 1.77), X (alpha satellite), and Y (satellite-III at Yqh) was performed on human interphase sperm nuclei. DNA probes were either directly labelled with rhodamine-dUTP, FITC-dUTP, or biotinylated by nick translation. Hybridization and signal detection were done by routine laboratory protocols. Microscopic analysis was performed using a cooled CCD camera attached to an epi-fluorescent microscope. After hybridization, fertile donors yielded a frequency of 0.96% (n=12) nullisomic, 98.5% (n=1231) monosomic and 0.96% (n=12) disomic for chromosome 1, whereas oligoasthenoteratozoospermic males yielded a frequency of 16% (n=600) nullisomic, 74.5% (n=2792) monosomic and 9.9% (n=370) disomic. In addition, fertile donors yielded a frequency of 45.7% (n=633) monosomic and 0.7% (n=11) disomic for chromosome X, whereas oligoasthenoteratozoospermic males yielded a frequency of 38.7% (n=760) monosomic and 0.8% (n=13) disomic. Chromosome Y frequencies for fertile donors showed 44.6% (n=614) monosomic and 0.6% (n=2) disomic, whereas oligoasthenoteratozoospermic males yielded a frequency of 33.2% (n=701) monosomic and 0.8% (n=15) disomic. This suggests that the frequency of nullisomy for chromosome 1 is significantly higher (p<0.001) in sperm from oligoasthenoteratozoospermic makes versus sperm from our fertile donors. We conclude that FISH is a powerful tool to determine the frequency of aneuploidy in sperm from oligoasthenoteratozoospermic patients.

  9. Recombinations in Staphylococcal Cassette Chromosome mec Elements Compromise the Molecular Detection of Methicillin Resistance in Staphylococcus aureus

    PubMed Central

    Hill-Cawthorne, Grant A.; Hudson, Lyndsey O.; El Ghany, Moataz Fouad Abd; Piepenburg, Olaf; Nair, Mridul; Dodgson, Andrew; Forrest, Matthew S.

    2014-01-01

    Clinical laboratories are increasingly using molecular tests for methicillin-resistant Staphylococcus aureus (MRSA) screening. However, primers have to be targeted to a variable chromosomal region, the staphylococcal cassette chromosome mec (SCCmec). We initially screened 726 MRSA isolates from a single UK hospital trust by recombinase polymerase amplification (RPA), a novel, isothermal alternative to PCR. Undetected isolates were further characterised using multilocus sequence, spa typing and whole genome sequencing. 96% of our tested phenotypically MRSA isolates contained one of the six orfX-SCCmec junctions our RPA test and commercially available molecular tests target. However 30 isolates could not be detected. Sequencing of 24 of these isolates demonstrated recombinations within the SCCmec element with novel insertions that interfered with the RPA, preventing identification as MRSA. This result suggests that clinical laboratories cannot rely solely upon molecular assays to reliably detect all methicillin-resistance. The presence of significant recombinations in the SCCmec element, where the majority of assays target their primers, suggests that there will continue to be isolates that escape identification. We caution that dependence on amplification-based molecular assays will continue to result in failure to diagnose a small proportion (∼4%) of MRSA isolates, unless the true level of SCCmec natural diversity is determined by whole genome sequencing of a large collection of MRSA isolates. PMID:24972080

  10. Comparative study of microsatellite and cytogenetic markers for detecting the origin of the nondisjoined chromosome 21 in down syndrome

    SciTech Connect

    Petersen, M.B.; Frantzen, M.; Lund, C.; Olsen, B.; Poulsen, H.; Sand, A.; Tommerup, N.; Mikkelsen, M. ); Antonarakis, S.E.; Warren, A.C. ); Van Broeckhoven, C. ); Chakravarti, A.; Cox, T.K. )

    1992-09-01

    Nondisjunction in trisomy 21 has traditionally been studied by cytogenetic heteromorphisms. Those studies assumed no crossing-over on the short arm of chromosome 21. Recently, increased accuracy of detection of the origin of nondisjunction has been demonstrated by DNA polymorphism analysis. The authors describe a comparative study of cytogenetic heteromorphisms and seven PCR-based DNA polymorphism analysis. They describe a comparative study of cytogenetic heteromorphisms and seven PCR-based DNA polymorphisms for detecting the origin of the additional chromosome 21 in 68 cases of Down syndrome. The polymorphisms studied were the highly informative microsatellites at loci D21S120, D21S192, IFNAR, D21S156, HMG14, and D21S171. The meiotic stage of nondisjunction was assigned on the basis of the pericentromeric markers D21S215, D21S120, and D21S192. Only unequivocal cytogenetic results were compared with the results of the DNA analysis. The parental and meiotic division origin could be determined in 51% of the cases by using the cytogenetic markers and in 88% of the cases by using the DNA markers. Although there were no discrepancies between the two scoring systems regarding parental origin, there were eight discrepancies regarding meiotic stage of nondisjunction. The results raise the possibility of recombination between the two marker systems, particularly on the short arm. 46 refs., 2 figs., 3 tabs.

  11. Recombinations in staphylococcal cassette chromosome mec elements compromise the molecular detection of methicillin resistance in Staphylococcus aureus.

    PubMed

    Hill-Cawthorne, Grant A; Hudson, Lyndsey O; El Ghany, Moataz Fouad Abd; Piepenburg, Olaf; Nair, Mridul; Dodgson, Andrew; Forrest, Matthew S; Clark, Taane G; Pain, Arnab

    2014-01-01

    Clinical laboratories are increasingly using molecular tests for methicillin-resistant Staphylococcus aureus (MRSA) screening. However, primers have to be targeted to a variable chromosomal region, the staphylococcal cassette chromosome mec (SCCmec). We initially screened 726 MRSA isolates from a single UK hospital trust by recombinase polymerase amplification (RPA), a novel, isothermal alternative to PCR. Undetected isolates were further characterised using multilocus sequence, spa typing and whole genome sequencing. 96% of our tested phenotypically MRSA isolates contained one of the six orfX-SCCmec junctions our RPA test and commercially available molecular tests target. However 30 isolates could not be detected. Sequencing of 24 of these isolates demonstrated recombinations within the SCCmec element with novel insertions that interfered with the RPA, preventing identification as MRSA. This result suggests that clinical laboratories cannot rely solely upon molecular assays to reliably detect all methicillin-resistance. The presence of significant recombinations in the SCCmec element, where the majority of assays target their primers, suggests that there will continue to be isolates that escape identification. We caution that dependence on amplification-based molecular assays will continue to result in failure to diagnose a small proportion (∼4%) of MRSA isolates, unless the true level of SCCmec natural diversity is determined by whole genome sequencing of a large collection of MRSA isolates.

  12. Development and evaluation of automated systems for detection and classification of banded chromosomes: current status and future perspectives

    NASA Astrophysics Data System (ADS)

    Wang, Xingwei; Zheng, Bin; Wood, Marc; Li, Shibo; Chen, Wei; Liu, Hong

    2005-08-01

    Automated detection and classification of banded chromosomes may help clinicians diagnose cancers and other genetic disorders at an early stage more efficiently and accurately. However, developing such an automated system (including both a high-speed microscopic image scanning device and related computer-assisted schemes) is quite a challenging and difficult task. Since the 1980s, great research efforts have been made to develop fast and more reliable methods to assist clinical technicians in performing this important and time-consuming task. A number of computer-assisted methods including classical statistical methods, artificial neural networks and knowledge-based fuzzy logic systems, have been applied and tested. Based on the initial test using limited datasets, encouraging results in algorithm and system development have been demonstrated. Despite the significant research effort and progress made over the last two decades, computer-assisted chromosome detection and classification systems have not been routinely accepted and used in clinical laboratories. Further research and development is needed.

  13. Association of a Chromosomal Rearrangement Event with Mouse Posterior Polymorphous Corneal Dystrophy and Alterations in Csrp2bp, Dzank1, and Ovol2 Gene Expression

    PubMed Central

    Shen, Anna L.; Moran, Susan A.; Glover, Edward A.; Drinkwater, Norman R.; Swearingen, Rebecca E.; Teixeira, Leandro B.; Bradfield, Christopher A.

    2016-01-01

    We have previously described a mouse model of human posterior polymorphous corneal dystrophy (PPCD) and localized the causative mutation to a 6.2 Mbp region of chromosome 2, termed Ppcd1. We now show that the gene rearrangement linked to mouse Ppcd1 is a 3.9 Mbp chromosomal inversion flanked by 81 Kbp and 542 bp deletions. This recombination event leads to deletion of Csrp2bp Exons 8 through 11, Dzank1 Exons 20 and 21, and the pseudogene Znf133. In addition, we identified translocation of novel downstream sequences to positions adjacent to Csrp2bp Exon 7 and Dzank1 Exon 20. Twelve novel fusion transcripts involving Csrp2bp or Dzank1 linked to downstream sequences have been identified. Eight are expressed at detectable levels in PPCD1 but not wildtype eyes. Upregulation of two Csrp2bp fusion transcripts, as well as upregulation of the adjacent gene, Ovol2, was observed. Absence of the PPCD1 phenotype in animals haploinsufficient for Csrp2bp or both Csrp2bp and Dzank1 rules out haploinsufficiency of these genes as a cause of mouse PPCD1. Complementation experiments confirm that PPCD1 embryonic lethality is due to disruption of Csrp2bp expression. The ocular expression pattern of Csrp2bp is consistent with a role for this protein in corneal development and pathogenesis of PPCD1. PMID:27310661

  14. Ethanol alters proliferation and differentiation of normal and chromosomally abnormal human embryonic stem cell-derived neurospheres.

    PubMed

    Krishnamoorthy, Malini; Gerwe, Brian A; Scharer, Christopher D; Sahasranaman, Vanita; Eilertson, Carmen D; Nash, Rachel J; Usta, Sümeyra Naz; Kelly, Shasmine; Rose, Matthew; Peraza, Rene; Arumugham, Jagan; Stewart, Bethany; Stice, Steven L; Nash, Rodney J

    2013-06-01

    Ethanol is a powerful substance and, when consumed during pregnancy, has significant psychoactive and developmental effects on the developing fetus. These abnormalities include growth retardation, neurological deficits, and behavioral and cognitive deficiencies, commonly referred to as fetal alcohol spectrum disorder. The effect of ethanol has been reported to affect cellular development on the embryonic level, however, not much is known about mutations contributing to the influence of ethanol. The purpose of our study was to determine if mutation contribute to changes in differentiation patterning, cell-cycle regulatory gene expression, and DNA methylation in human embryonic stem cells after ethanol exposure. We exposed human embryonic stem cells (with and without know DNA mutations) to a low concentration (20 mM) of ethanol and measured neurosphere proliferation and differentiation, glial protein levels, expression of various cell-cycle genes, and DNA methylation. Ethanol altered cell-cycle gene expression between the two cell lines; however, gene methylation was not affected in ether lines.

  15. Detection of random alterations to time-varying musical instrument spectra

    NASA Astrophysics Data System (ADS)

    Horner, Andrew; Beauchamp, James; So, Richard

    2004-09-01

    The time-varying spectra of eight musical instrument sounds were randomly altered by a time-invariant process to determine how detection of spectral alteration varies with degree of alteration, instrument, musical experience, and spectral variation. Sounds were resynthesized with centroids equalized to the original sounds, with frequencies harmonically flattened, and with average spectral error levels of 8%, 16%, 24%, 32%, and 48%. Listeners were asked to discriminate the randomly altered sounds from reference sounds resynthesized from the original data. For all eight instruments, discrimination was very good for the 32% and 48% error levels, moderate for the 16% and 24% error levels, and poor for the 8% error levels. When the error levels were 16%, 24%, and 32%, the scores of musically experienced listeners were found to be significantly better than the scores of listeners with no musical experience. Also, in this same error level range, discrimination was significantly affected by the instrument tested. For error levels of 16% and 24%, discrimination scores were significantly, but negatively correlated with measures of spectral incoherence and normalized centroid deviation on unaltered instrument spectra, suggesting that the presence of dynamic spectral variations tends to increase the difficulty of detecting spectral alterations. Correlation between discrimination and a measure of spectral irregularity was comparatively low.

  16. Alteration/Deficiency in Activation 3 (ADA3) Protein, a Cell Cycle Regulator, Associates with the Centromere through CENP-B and Regulates Chromosome Segregation.

    PubMed

    Mohibi, Shakur; Srivastava, Shashank; Wang-France, Jun; Mirza, Sameer; Zhao, Xiangshan; Band, Hamid; Band, Vimla

    2015-11-20

    ADA3 (alteration/deficiency in activation 3) is a conserved component of several transcriptional co-activator and histone acetyltransferase (HAT) complexes. Recently, we generated Ada3 knock-out mice and demonstrated that deletion of Ada3 leads to early embryonic lethality. The use of Ada3(FL/FL) mouse embryonic fibroblasts with deletion of Ada3 using adenovirus Cre showed a critical role of ADA3 in cell cycle progression through mitosis. Here, we demonstrate an association of ADA3 with the higher order repeat region of the α-satellite region on human X chromosome centromeres that is consistent with its role in mitosis. Given the role of centromere proteins (CENPs) in mitosis, we next analyzed whether ADA3 associates with the centromere through CENPs. Both an in vivo proximity ligation assay and immunofluorescence studies confirmed the association of ADA3 with CENP-B protein, a highly conserved centromeric protein that binds to the 17-bp DNA sequences on α-satellite DNA. Deletional analysis showed that ADA3 directly associates with CENP-B through its N terminus, and a CENP-B binding-deficient mutant of ADA3 was incompetent in cell proliferation rescue. Notably, knockdown of ADA3 decreased binding of CENP-B onto the centromeres, suggesting that ADA3 is required for the loading of CENP-B onto the centromeres. Finally, we show that deletion of Ada3 from Ada3(FL/FL) mouse embryonic fibroblasts exhibited various chromosome segregation defects. Taken together, we demonstrate a novel ADA3 interaction with CENP-B-centromere that may account for its previously known function in mitosis. This study, together with its known function in maintaining genomic stability and its mislocalization in cancers, suggests an important role of ADA3 in mitosis.

  17. Alteration/Deficiency in Activation 3 (ADA3) Protein, a Cell Cycle Regulator, Associates with the Centromere through CENP-B and Regulates Chromosome Segregation*

    PubMed Central

    Mohibi, Shakur; Srivastava, Shashank; Wang-France, Jun; Mirza, Sameer; Zhao, Xiangshan; Band, Hamid; Band, Vimla

    2015-01-01

    ADA3 (alteration/deficiency in activation 3) is a conserved component of several transcriptional co-activator and histone acetyltransferase (HAT) complexes. Recently, we generated Ada3 knock-out mice and demonstrated that deletion of Ada3 leads to early embryonic lethality. The use of Ada3FL/FL mouse embryonic fibroblasts with deletion of Ada3 using adenovirus Cre showed a critical role of ADA3 in cell cycle progression through mitosis. Here, we demonstrate an association of ADA3 with the higher order repeat region of the α-satellite region on human X chromosome centromeres that is consistent with its role in mitosis. Given the role of centromere proteins (CENPs) in mitosis, we next analyzed whether ADA3 associates with the centromere through CENPs. Both an in vivo proximity ligation assay and immunofluorescence studies confirmed the association of ADA3 with CENP-B protein, a highly conserved centromeric protein that binds to the 17-bp DNA sequences on α-satellite DNA. Deletional analysis showed that ADA3 directly associates with CENP-B through its N terminus, and a CENP-B binding-deficient mutant of ADA3 was incompetent in cell proliferation rescue. Notably, knockdown of ADA3 decreased binding of CENP-B onto the centromeres, suggesting that ADA3 is required for the loading of CENP-B onto the centromeres. Finally, we show that deletion of Ada3 from Ada3FL/FL mouse embryonic fibroblasts exhibited various chromosome segregation defects. Taken together, we demonstrate a novel ADA3 interaction with CENP-B-centromere that may account for its previously known function in mitosis. This study, together with its known function in maintaining genomic stability and its mislocalization in cancers, suggests an important role of ADA3 in mitosis. PMID:26429915

  18. Detection of Y chromosome sequences in a 45,X/46,XXq - patient by Southern blot analysis of PCR-amplified DNA and fluorescent in situ hybridization (FISH)

    SciTech Connect

    Kocova, M.; Siegel, S.F.; Wenger, S.L.

    1995-02-13

    In some cases of gonadal dysgenesis, cytogenetic analysis seems to be discordant with the phenotype of the patients. We have applied techniques such as Southern blot analysis and fluorescent in situ hybridization (FISH) to resolve the phenotype/genotype discrepancy in a patient with ambiguous genitalia in whom the peripheral blood karotype was 45,X. Gonadectomy at age 7 months showed the gonadal tissue to be prepubertal testis on the left side and a streak gonad on the right. The karyotype obtained from the left gonad was 45,X/46,XXq- and that from the right gonad was 45,X. Three different techniques, PCR amplification, FISH, and chromosome painting for X and Y chromosomes, confirmed the presence of Y chromosome sequences. Five different tissues were evaluated. The highest percentage of Y chromosome positive cells were detected in the left gonad, followed by the peripheral blood lymphocytes, skin fibroblasts, and buccal mucosa. No Y chromosomal material could be identified in the right gonad. Since the Xq- chromosome is present in the left gonad (testis), it is likely that the Xq- contains Y chromosomal material. Sophisticated analysis in this patient showed that she has at least 2 cell lines, one of which contains Y chromosomal material. These techniques elucidated the molecular basis of the genital ambiguity for this patient. When Y chromosome sequences are present in patients with Ullrich-Turner syndrome or gonadal dysgenesis, the risk for gonadal malignancy is significantly increased. Hence, molecular diagnostic methods to ascertain for the presence of Y chromosome sequences may expedite the evaluation of patients with the ambiguous genitalia. 21 refs., 4 figs., 2 tabs.

  19. Detection of gene expression changes at chromosomal rearrangement breakpoints in evolution.

    PubMed

    Muñoz, Adriana; Sankoff, David

    2012-03-21

    We study the relation between genome rearrangements, breakpoints and gene expression. Genome rearrangement research has been concerned with the creation of breakpoints and their position in the chromosome, but the functional consequences of individual breakpoints remain virtually unknown, and there are no direct genome-wide studies of breakpoints from this point of view. A question arises of what the biological consequences of breakpoint creation are, rather than just their structural aspects. The question is whether proximity to the site of a breakpoint event changes the activity of a gene. We investigate this by comparing the distribution of distances to the nearest breakpoint of genes that are differentially expressed with the distribution of the same distances for the entire gene complement. We study this in data on whole blood tissue in human versus macaque, and in cerebral cortex tissue in human versus chimpanzee. We find in both data sets that the distribution of distances to the nearest breakpoint of "changed expression genes" differs little from this distance calculated for the rest of the gene complement. In focusing on the changed expression genes closest to the breakpoints, however, we discover that several of these have previously been implicated in the literature as being connected to the evolutionary divergence of humans from other primates. We conjecture that chromosomal rearrangements occasionally interrupt the regulatory configurations of genes close to the breakpoint, leading to changes in expression.

  20. Detection of gene expression changes at chromosomal rearrangement breakpoints in evolution

    PubMed Central

    2012-01-01

    Background We study the relation between genome rearrangements, breakpoints and gene expression. Genome rearrangement research has been concerned with the creation of breakpoints and their position in the chromosome, but the functional consequences of individual breakpoints remain virtually unknown, and there are no direct genome-wide studies of breakpoints from this point of view. A question arises of what the biological consequences of breakpoint creation are, rather than just their structural aspects. The question is whether proximity to the site of a breakpoint event changes the activity of a gene. Results We investigate this by comparing the distribution of distances to the nearest breakpoint of genes that are differentially expressed with the distribution of the same distances for the entire gene complement. We study this in data on whole blood tissue in human versus macaque, and in cerebral cortex tissue in human versus chimpanzee. We find in both data sets that the distribution of distances to the nearest breakpoint of "changed expression genes" differs little from this distance calculated for the rest of the gene complement. In focusing on the changed expression genes closest to the breakpoints, however, we discover that several of these have previously been implicated in the literature as being connected to the evolutionary divergence of humans from other primates. Conclusions We conjecture that chromosomal rearrangements occasionally interrupt the regulatory configurations of genes close to the breakpoint, leading to changes in expression. PMID:22536904

  1. Detection of B chromosomes in interphase hemolymph nuclei from living specimens of the grasshopper Eyprepocnemis plorans.

    PubMed

    Cabrero, J; Manrique-Poyato, M I; Camacho, J P M

    2006-01-01

    The two most important evolutionary properties of B chromosomes are their transmission rate (which suggests their selfishness when significantly higher than 0.5) and their net effects on carrier fitness (usually negative for parasitic Bs). The study of transmission rate unavoidably requires the analysis of many controlled crosses in order to accurately measure population average transmission rate. Therefore, getting a marker closely associated to B presence is of crucial importance to alleviate the load of performing many useless crosses between lacking B individuals. After investigating several cytogenetic techniques on several tissues that may be sampled without drastically damaging live specimens of the grasshopper Eyprepocnemis plorans, we report here the excellent results provided by the CMA3 fluorescence and C-banding techniques applied to hemocyte nuclei. These cells may be easily obtained from both males and females and provide information on B presence even during the interphase stage. The two cytogenetic techniques take advantage of the heterochromatic nature of the B chromosomes, so that Bs made predominantly of ribosomal DNA are revealed by CMA3 as bright bodies in the interphase hemocytes, and Bs mostly made of satellite DNA are visualized by C-banding as intensely stained bodies in these cells. 2006 S. Karger AG, Basel.

  2. Detection of Imprinting Effects for Quantitative Traits on X Chromosome Using Nuclear Families with Multiple Daughters.

    PubMed

    Yu, Kexin; Zhou, Ji-Yuan; Fung, Wing Kam

    2017-07-01

    Genomic imprinting is an epigenetic phenomenon in which the expression of an allele copy depends on its parental origin. This mechanism has been found to play an important role in many complex diseases. Statistical tests for imprinting effects have been developed for more than 15 years, but they are only suitable for autosomes. It was not until recently that the parental-asymmetry test on the X chromosome (XPAT) was proposed to test for imprinting effects. However, this test can only be used for qualitative traits. Therefore, in this article, we propose a class of PAT-type tests to test for imprinting for quantitative traits on the X chromosome in the presence of association, namely, Q-XPAT(c), Q-1-XPAT(c) and Q-C-XPAT(c), where c is a constant. These methods can accommodate complete and incomplete nuclear families with an arbitrary number of daughters. Extensive simulation studies demonstrate that the proposed tests control the size well under the null hypothesis of no imprinting effects and are powerful under various family structures. Moreover, by setting the inbreeding coefficient in females to be nonzero and using the assortative mating pattern in simulations, the proposed tests are shown to be valid under Hardy-Weinberg disequilibrium. © 2017 John Wiley & Sons Ltd/University College London.

  3. Unexpected heterogeneity of BCR-ABL fusion mRNA detected by polymerase chain reaction in Philadelphia chromosome-positive acute lymphoblastic leukemia

    SciTech Connect

    Hooberman, A.L.; Carrino, J.J.; Leibowitz, D.; Rowley, J.D.; Le Beau, M.M.; Arlin, Z.A.; Westbrook, C.A. )

    1989-06-01

    The Philadelphia (Ph{sup 1}) chromosome results in a fusion of portions of the BCR gene from chromosome 22 and the ABL gene from chromosome 9, producing a chimeric BCR-ABL mRNA and protein. In lymphoblastic leukemias, there are two molecular subtypes of the Ph{sup 1} chromosome, one with a rearrangement of the breakpoint cluster region (bcr) of the BCR gene, producing the same 8.5-kilobase BCR-ABL fusion mRNA seen in chronic myelogenous leukemia (CML), and the other, without a bcr rearrangement, producing a 7.0-kilobase BCR-ABL fusion mRNA that is seen only in acute lymphoblastic leukemia (ALL). The authors studied the molecular subtype of the Ph{sup 1} chromosome in 11 cases of Ph{sup 1}-positive ALL, including 2 with a previous diagnosis of CML, using a sensitive method to analyze the mRNA species based on the polymerase chain reaction (PCR). They observed unexpected heterogeneity in BCR-ABL mRNA in this population. They conclude that the PCR gives additional information about the Ph{sup 1} chromosome gene products that cannot be obtained by genomic analysis, but that it cannot be used as the sole means of detection of this chromosomal abnormality in ALL because of the high incidence of false negative results.

  4. Mate pair sequencing for the detection of chromosomal aberrations in patients with intellectual disability and congenital malformations

    PubMed Central

    Vergult, Sarah; Van Binsbergen, Ellen; Sante, Tom; Nowak, Silke; Vanakker, Olivier; Claes, Kathleen; Poppe, Bruce; Van der Aa, Nathalie; van Roosmalen, Markus J; Duran, Karen; Tavakoli-Yaraki, Masoumeh; Swinkels, Marielle; van den Boogaard, Marie-José; van Haelst, Mieke; Roelens, Filip; Speleman, Frank; Cuppen, Edwin; Mortier, Geert; Kloosterman, Wigard P; Menten, Björn

    2014-01-01

    Recently, microarrays have replaced karyotyping as a first tier test in patients with idiopathic intellectual disability and/or multiple congenital abnormalities (ID/MCA) in many laboratories. Although in about 14–18% of such patients, DNA copy-number variants (CNVs) with clinical significance can be detected, microarrays have the disadvantage of missing balanced rearrangements, as well as providing no information about the genomic architecture of structural variants (SVs) like duplications and complex rearrangements. Such information could possibly lead to a better interpretation of the clinical significance of the SV. In this study, the clinical use of mate pair next-generation sequencing was evaluated for the detection and further characterization of structural variants within the genomes of 50 ID/MCA patients. Thirty of these patients carried a chromosomal aberration that was previously detected by array CGH or karyotyping and suspected to be pathogenic. In the remaining 20 patients no causal SVs were found and only benign aberrations were detected by conventional techniques. Combined cluster and coverage analysis of the mate pair data allowed precise breakpoint detection and further refinement of previously identified balanced and (complex) unbalanced aberrations, pinpointing the causal gene for some patients. We conclude that mate pair sequencing is a powerful technology that can provide rapid and unequivocal characterization of unbalanced and balanced SVs in patient genomes and can be essential for the clinical interpretation of some SVs. PMID:24105367

  5. LANDSAT detection of hydrothermal alteration in the Nogal Canyon Cauldron, New Mexico

    NASA Technical Reports Server (NTRS)

    Vincent, R. K.; Rouse, G.

    1977-01-01

    In 1974 a circular-shaped iron oxide anomaly was observed in an image of a LANDSAT frame centered near Truth or Consequences, New Mexico. Field examination of the anomaly has shown that it coincides with a zone of hydrothermal alteration on the northern edge of the Nogal Canyon Cauldron. The altered area contains clay minerals ranging in colors from white to vivid red, the latter presumably resulting from hematite staining. In situ gas measurements showed no evidence of active hydrogen sulfide seepage. Preliminary geochemical analyses of grab samples have detected no significant amounts of mineralization. Whereas this area does not at present appear to be economically important, it provides an example of how LANDSAT can be utilized in reconnaissance mapping for cauldrons, calderas, and other volcanic features which display hydrothermal alteration.

  6. [Chromosomal instability in carcinogenesis of cervical cancer.

    PubMed

    de Los Santos-Munive, Victoria; Alonso-Avelino, Juan Angel

    2013-01-01

    In order to spot common chromosomal imbalances in early and late lesions of cervical cancer that might be used as progression biomarkers, we made a search of literature in PubMed from 1996 to 2011. The medical subject headings employed were chromosomal alterations, loss of heterozygosis, cervical cancer, cervical tumorigenesis, chromosomal aberrations, cervical intraepithelial neoplasm and low-grade squamous intraepithelial lesion. The common chromosomal imbalances were gains in 8q24 (77.7 %), 20q13 (66.9 %), 3q26 (47.1 %), Xp22 (43.8 %), and 5p15 (60 %), principally. On the other hand, integration of the high-risk human papillomavirus genome into the host chromosome has been associated with the development of neoplasia, but the chromosomal imbalances seem to precede and promote such integration. Chromosomal imbalances in 8q24, 20q13, 3q21-26 and 5p15-Xp22, determined by fluorescent in situ hybridization assay or comparative genomic hybridization assay for early detection of the presence of high-risk human papillomavirus, are promising markers of cervical cancer progression.

  7. Microarray based comparative genomic hybridisation (array-CGH) detects submicroscopic chromosomal deletions and duplications in patients with learning disability/mental retardation and dysmorphic features

    PubMed Central

    Shaw-Smith, C; Redon, R; Rickman, L; Rio, M; Willatt, L; Fiegler, H; Firth, H; Sanlaville, D; Winter, R; Colleaux, L; Bobrow, M; Carter, N

    2004-01-01

    The underlying causes of learning disability and dysmorphic features in many patients remain unidentified despite extensive investigation. Routine karyotype analysis is not sensitive enough to detect subtle chromosome rearrangements (less than 5 Mb). The presence of subtle DNA copy number changes was investigated by array-CGH in 50 patients with learning disability and dysmorphism, employing a DNA microarray constructed from large insert clones spaced at approximately 1 Mb intervals across the genome. Twelve copy number abnormalities were identified in 12 patients (24% of the total): seven deletions (six apparently de novo and one inherited from a phenotypically normal parent) and five duplications (one de novo and four inherited from phenotypically normal parents). Altered segments ranged in size from those involving a single clone to regions as large as 14 Mb. No recurrent deletion or duplication was identified within this cohort of patients. On the basis of these results, we anticipate that array-CGH will become a routine method of genome-wide screening for imbalanced rearrangements in children with learning disability. PMID:15060094

  8. Alteration mineral mapping in inaccessible regions using target detection algorithms to ASTER data

    NASA Astrophysics Data System (ADS)

    Pour, A. B.; Hashim, M.; Park, Y.

    2017-05-01

    In this study, the applications of target detection algorithms such as Constrained Energy Minimization (CEM), Orthogonal Subspace Projection (OSP) and Adaptive Coherence Estimator (ACE) to shortwave infrared bands of the Advanced Spaceborne Thermal Emission and Reflection Radiometer (ASTER) data was investigated to extract geological information for alteration mineral mapping in poorly exposed lithologies in inaccessible domains. The Oscar II coast area north-eastern Graham Land, Antarctic Peninsula (AP) was selected in this study to conduct a satellite-based remote sensing mapping technique. It is an inaccessible region due to the remoteness of many rock exposures and the necessity to travel over sever mountainous and glacier-cover terrains for geological field mapping and sample collection. Fractional abundance of alteration minerals such as muscovite, kaolinite, illite, montmorillonite, epidote, chlorite and biotite were identified in alteration zones using CEM, OSP and ACE algorithms in poorly mapped and unmapped zones at district scale for the Oscar II coast area. The results of this investigation demonstrated the applicability of ASTER shortwave infrared spectral data for lithological and alteration mineral mapping in poorly exposed lithologies and inaccessible regions, particularly using the image processing algorithms that are capable to detect sub-pixel targets in the remotely sensed images, where no prior information is available.

  9. Segmental Chromosomal Aberrations in Localized Neuroblastoma Can be Detected in Formalin-Fixed Paraffin-Embedded Tissue Samples and Are Associated With Recurrence

    PubMed Central

    Pinto, Navin; Mayfield, Jodi R.; Raca, Gordana; Applebaum, Mark A.; Chlenski, Alexandre; Sukhanova, Madina; Bagatell, Rochelle; Irwin, Meredith S.; Little, Anthony; Rawwas, Jawhar; Gosiengfiao, Yasmin; Delattre, Olivier; Janoueix-Lerosey, Isabelle; Lapouble, Eve; Schleiermacher, Gudrun; Cohn, Susan L.

    2016-01-01

    Background Array comparative genomic hybridization (CGH) analyses of frozen tumors have shown strong associations between the pattern of chromosomal aberrations and outcome in patients with advanced-stage neuroblastoma. New platforms for analyzing chromosomal aberrations using formalin-fixed paraffin-embedded (FFPE) tissue have recently been developed. We sought to determine whether chromosomal microarray analysis (CMA) using FFPE tumors is feasible and if segmental chromosomal aberrations were prognostic of recurrence in localized neuroblastoma. Methods Patients with MYCN nonamplified International Neuroblastoma Staging System stage 1 and 2 disease who recurred were identified. CMA was performed with diagnostic FFPE samples using OncoScan™ FFPE Express 2.0. The prognostic significance of chromosomal pattern was validated in 105 patients with available CGH results. Results In 26 evaluable patients, 11 recurred locally, nine had metastatic relapse, and six remained progression free >3 years from diagnosis. No chromosomal aberrations were identified in four tumors. Numerical chromosomal aberrations (NCAs) without segmental chromosomal aberration (SCA) were identified in 11 patients: six progressed locally, two had metastatic progression and 3 remained progression-free. Eleven patients had SCAs: four progressed locally, six developed metastatic progression and one remained progression-free. Five or more SCAs were only detected in tumors from patients who developed metastases (P = 0.0004). In the validation cohort, SCAs were associated with inferior event-free survival (EFS) compared to NCA (5-year EFS 68% ± 8.3% vs. 91% ± 3.6%, respectively; P = 0.0083). Conclusions It is feasible to evaluate chromosomal aberrations using FFPE neuroblastoma tissue. SCA is associated with inferior EFS in localized neuroblastoma patients, and multiple SCAs may be predictive of metastatic relapse. PMID:26864375

  10. 6q deletion detected by fluorescence in situ hybridization using bacterial artificial chromosome in chronic lymphocytic leukemia.

    PubMed

    Dalsass, Alessia; Mestichelli, Francesca; Ruggieri, Miriana; Gaspari, Paola; Pezzoni, Valerio; Vagnoni, Davide; Angelini, Mario; Angelini, Stefano; Bigazzi, Catia; Falcioni, Sadia; Troiani, Emanuela; Alesiani, Francesco; Catarini, Massimo; Attolico, Immacolata; Scortechini, Ilaria; Discepoli, Giancarlo; Galieni, Piero

    2013-07-01

    Deletions of the long arm of chromosome 6 are known to occur at relatively low frequency (3-6%) in chronic lymphocytic leukemia (CLL), and they are more frequently observed in 6q21. Few data have been reported regarding other bands on 6q involved by cytogenetic alterations in CLL. The cytogenetic study was performed in nuclei and metaphases obtained after stimulation with a combination of CpG-oligonucleotide DSP30 and interleukin-2. Four bacterial artificial chromosome (BAC) clones mapping regions in bands 6q16, 6q23, 6q25, 6q27 were used as probes for fluorescence in situ hybridization in 107 CLL cases in order to analyze the occurrence and localization of 6q aberrations. We identified 11 cases (10.2%) with 6q deletion of 107 patients studied with CLL. The trends of survival curves and the treatment-free intervals (TFI) of patients with deletion suggest a better outcome than the other cytogenetic risk groups. We observed two subgroups with 6q deletion as the sole anomaly: two cases with 6q16 deletion, and three cases with 6q25.2-27 deletion. There were differences of age, stage, and TFI between both subgroups. By using BAC probes, we observed that 6q deletion has a higher frequency in CLL and is linked with a good prognosis. In addition, it was observed that the deletion in 6q16 appears to be the most frequent and, if present as the only abnormality, it could be associated with a most widespread disease. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. An improved assay for the detection of alterations in bacterial DNA supercoiling in vivo.

    PubMed

    Abu Mraheil, M; Heisig, A; Heisig, P

    2013-07-01

    Due to the increasing prevalence of antibiotic resistance and the yet low output of the genomics-based drug discovery approach novel strategies are urgently needed to detect new antibiotics. One such strategy uses known ubiquitous targets like DNA topoisomerases. However, to detect inhibitors of these enzymes by an in vitro assay time-consuming isolation of enzymes and DNA followed by electrophoretic separation of topoisomers are required. Instead, this study aimed at developing an in vivo assay for the detection of alterations in DNA supercoiling indicative of topoisomerase inhibition by a reporter gene assay. A pair of plasmids was developed which carry the reporter gene luc for firefly luciferase under control of either promoter ptopA (pPHB90) or pgyrA (pPHB91), whose activities are reciprocally affected by alterations of the supercoiling degree. Each plasmid is individually transferred into E. coli cells. The quotient of the luciferase activities determined using cells with either plasmid was taken as relative measure of the global supercoiling degree Qsc (quotient of supercoiling). Using isogenic reference strains with known alterations of the global DNA supercoiling degree due to mutations in either gyrB or topA, the reporter gene system was able to detect both a decrease and an increase of the negative supercoiling degree compared to the isogenic parent strain. Treating cells with known inhibitors of DNA gyrase, like fluoroquinolones, novobiocin as well as simocyclinone D8 from Streptomyces antibioticus which has been identified as an inhibitor of DNA gyrase in vitro, also caused decreases of the Qsc value in vivo. The suitability of this reporter gene system to screen for anti-topoisomerase I and II compounds from various natural sources like plant extracts by sensing alterations of the DNA supercoiling was demonstrated and offers a new application to identify novel compounds active against bacterial topoisomerases I and gyrase.

  12. B chromosome contains active genes and impacts the transcription of A chromosomes in maize (Zea mays L.).

    PubMed

    Huang, Wei; Du, Yan; Zhao, Xin; Jin, Weiwei

    2016-04-16

    The dispensable maize (Zea mays L.) B chromosome is highly heterochromatic and widely believed to be devoid of functional genes. Although low-copy B chromosome causes no obvious phenotype variation, its existence might influence A genome gene expression. Previous studies suggested that B chromosomes are evolved from standard chromosomes; therefore, they might contain genic regions showing homology with A chromosome sequences. Our data suggested that maize B chromosome influences the A-genome transcription with stronger effect associated with an increase in copy number of B chromosome. In total 130 differently expressed genes were detected in comparison between with and without B chromosome lines. These differentially expressed genes are mainly involved in cell metabolism and nucleotide binding. Using Starter + B, we amplified ten B chromosome loci with high sequence similarity to A-genome genes. Fluorescence in situ hybridization (FISH) confirmed that at least four ~5 kb-sized genes are located on the B chromosome. In addition, through de novo assembly of the reads not unmapped to maize B73 reference genome together with PCR validation, we found three B-located LTR; in particular, one of them, the 3.2 kb comp75688, is expressed in a B-dosage dependent manner. We found that in the presence of maize B chromosome, the transcription of A genome genes was altered, with more impact by the increase of the B chromosome number. The B-located transcriptionally active genes showed high similarity to their A-genome homologues, and retrotransposons on B chromosome also have partial homologous to A genome sequences. Our data shed more lights on the genome structure and evolution of the maize B chromosome.

  13. Chromosomal aberrations and aneuploidies of spermatozoa.

    PubMed

    Piomboni, Paola; Stendardi, Anita; Gambera, Laura

    2014-01-01

    Chromosomal abnormalities are relevant causes of human infertility, affecting 2 -14 % of infertile males. Patients with seminal anomalies could be affected by improper meiotic recombination and increased sperm chromosome aneuploidy. Since the transmission of a haploid chromosomal asset is fundamental for embryo vitality and development, the study of sperm chromosomes has become fundamental because intracytoplasmic sperm injection allows fertilization in cases of severe male infertility.In this chapter we summarize the data on the incidence of sperm aneuploidy, detected by fluorescence in situ hybridization (FISH), in infertile men with normal or abnormal karyotype. The possibility of reducing sperm chromosomal imbalance is also reported.Among control males, the lowest aneuploidy rate was detected (range: 0.09 -0.14 % for autosomes; 0.04 -0.10 % for gonosomes). In infertile patients with normal karyotype, the severity of semen alteration is correlated with the frequency of aneuploidy, particularly for X and Y chromosomes. Among patients with abnormal karyotype, 47,XXY and 47,XYY carriers showed a high variability of sperm aneuploidy both for gonosomes and autosomes. In Robertsonian translocation carriers, the increase in aneuploidy rate was particularly evident for total sex disomy, and resulted mainly from interchromosomal effect (ICE). In reciprocal translocation carriers, a high percentage of unbalanced sperm (approximately 50 %) was detected, perhaps mostly related to ICE.Sperm chromosomal constitution could be analyzed to obtain more accurate information about the causes of male infertility. It would be worthwhile to evaluate the benefits of a therapy with recombinant Follicle Stimulating Hormone (rFSH) on sperm chromosome segregation in selected infertile males.

  14. High Frequency of Genetic Alterations in Non-small Cell Lung Cancer Detected by Multi-target Fluorescence In Situ Hybridization

    PubMed Central

    Kang, Ji Un; Kwon, Kye Chul; Park, Jong Woo; Shin, So Youn; Kim, Jin Man; Jung, Sung Su

    2007-01-01

    Detection of genetic alterations could provide a tool as an adjuvant for the diagnosis of non-small cell lung cancer (NSCLC) and to define patients at risk for early relapse. In this study, a multi-target fluorescence in situ hybridization (FISH) assay was conducted to investigate the correlation between the alterations of chromosomes, including 5p15.2, 6p11.1-q11, 7p12, and 8q24.12-q24.13 (LaVysion Test), and clinicopathological variables, and to clarify the potential of the multi-target FISH assay in 37 NSCLC. The most notable finding was the higher frequency of a gain in chromosome 5p15.2 in early-stage (I+IIa) lung cancers. The frequency of the gain was 81.3% (16/22) in stage I tumors. The frequencies of gains in 6p11.1-q11 and 8q24.12-q24.13 were 61.5% (8/13) and 84.6% (11/13) in stage IIIa cancers, as compared with lower frequencies in stage I tumors at 25.0% and 31.3%, respectively. There was also a significant difference in the histological type. Our results suggest that a gain in 6p11.1-q11 and 8q24.12-q24.13 plays an important role in tumor progression and is associated with histological differentiation. On the other hand, gene amplification in the 5p region was one of the most consistent alterations in early-stage lung cancer, and thus a series of genes in the critical 5p15.2 region might potentially associated with the development of lung cancer. PMID:17923754

  15. High frequency of genetic alterations in non-small cell lung cancer detected by multi-target fluorescence in situ hybridization.

    PubMed

    Kang, Ji Un; Koo, Sun Hoe; Kwon, Kye Chul; Park, Jong Woo; Shin, So Youn; Kim, Jin Man; Jung, Sung Su

    2007-09-01

    Detection of genetic alterations could provide a tool as an adjuvant for the diagnosis of non-small cell lung cancer (NSCLC) and to define patients at risk for early relapse. In this study, a multi-target fluorescence in situ hybridization (FISH) assay was conducted to investigate the correlation between the alterations of chromosomes, including 5p15.2, 6p11.1-q11, 7p12, and 8q24.12-q24.13 (LaVysion Test), and clinicopathological variables, and to clarify the potential of the multi-target FISH assay in 37 NSCLC. The most notable finding was the higher frequency of a gain in chromosome 5p15.2 in early-stage (I+IIa) lung cancers. The frequency of the gain was 81.3% (16/22) in stage I tumors. The frequencies of gains in 6p11.1-q11 and 8q24.12-q24.13 were 61.5% (8/13) and 84.6% (11/13) in stage IIIa cancers, as compared with lower frequencies in stage I tumors at 25.0% and 31.3%, respectively. There was also a significant difference in the histological type. Our results suggest that a gain in 6p11.1-q11 and 8q24.12-q24.13 plays an important role in tumor progression and is associated with histological differentiation. On the other hand, gene amplification in the 5p region was one of the most consistent alterations in early-stage lung cancer, and thus a series of genes in the critical 5p15.2 region might potentially associated with the development of lung cancer.

  16. Sensitive Detection of Chromosomal Segments of Distinct Ancestry in Admixed Populations

    PubMed Central

    Price, Alkes L.; Tandon, Arti; Patterson, Nick; Barnes, Kathleen C.; Rafaels, Nicholas; Ruczinski, Ingo; Beaty, Terri H.; Mathias, Rasika; Reich, David; Myers, Simon

    2009-01-01

    Identifying the ancestry of chromosomal segments of distinct ancestry has a wide range of applications from disease mapping to learning about history. Most methods require the use of unlinked markers; but, using all markers from genome-wide scanning arrays, it should in principle be possible to infer the ancestry of even very small segments with exquisite accuracy. We describe a method, HAPMIX, which employs an explicit population genetic model to perform such local ancestry inference based on fine-scale variation data. We show that HAPMIX outperforms other methods, and we explore its utility for inferring ancestry, learning about ancestral populations, and inferring dates of admixture. We validate the method empirically by applying it to populations that have experienced recent and ancient admixture: 935 African Americans from the United States and 29 Mozabites from North Africa. HAPMIX will be of particular utility for mapping disease genes in recently admixed populations, as its accurate estimates of local ancestry permit admixture and case-control association signals to be combined, enabling more powerful tests of association than with either signal alone. PMID:19543370

  17. Novel Y-chromosome Short Tandem Repeat Variants Detected Through the Use of Massively Parallel Sequencing

    PubMed Central

    Warshauer, David H.; Churchill, Jennifer D.; Novroski, Nicole; King, Jonathan L.; Budowle, Bruce

    2015-01-01

    Massively parallel sequencing (MPS) technology is capable of determining the sizes of short tandem repeat (STR) alleles as well as their individual nucleotide sequences. Thus, single nucleotide polymorphisms (SNPs) within the repeat regions of STRs and variations in the pattern of repeat units in a given repeat motif can be used to differentiate alleles of the same length. In this study, MPS was used to sequence 28 forensically-relevant Y-chromosome STRs in a set of 41 DNA samples from the 3 major U.S. population groups (African Americans, Caucasians, and Hispanics). The resulting sequence data, which were analyzed with STRait Razor v2.0, revealed 37 unique allele sequence variants that have not been previously reported. Of these, 19 sequences were variations of documented sequences resulting from the presence of intra-repeat SNPs or alternative repeat unit patterns. Despite a limited sampling, two of the most frequently-observed variants were found only in African American samples. The remaining 18 variants represented allele sequences for which there were no published data with which to compare. These findings illustrate the great potential of MPS with regard to increasing the resolving power of STR typing and emphasize the need for sample population characterization of STR alleles. PMID:26391384

  18. Detection of obesity QTLs on mouse chromosomes 1 and 7 by selective DNA pooling

    SciTech Connect

    Taylor, B.A.; Phillips, S.J.

    1996-06-15

    The inheritance of obesity has been analyzed in an intercross between the lean 129/Sv mouse strain and the obesity-prone EL/Suz mouse strain. The weights of three major fat pads were determined on 4-month-old mice, and the sum of these weights, divided by body weight, was used as an adiposity index. The strategy of selective DNA pooling was used as a primary screen to identify putative quantitative trait loci (QTLs) affecting adiposity index. DNA pools representing the leanest 15% and fattest 15% of the F2 progeny were compared for differential allelic enrichment using widely dispersed microsatellite variants. To evaluate putative QTLs, individual genotyping and interval mapping were employed to estimate QTL effects and assess statistical significance. One QTL affecting adiposity index, which accounted for 12.3% of phenotypic variance in gender-merged data, was mapped to the central region of Chromosome (Chr) 7. The QTL allele inherited from EL conferred increased adiposity. A second QTL that accounts for 6.3% of phenotypic variance was identified on Chr 1 near D1Mitt211. At both QTLs, the data are consistent with dominant inheritance of the allele contributing to obesity. The possible relationships between these QTLs and previously described obesity QYLs, major obesity mutations, and candidate genes are discussed. 42 refs., 3 figs., 3 tabs.

  19. Rapid testing versus karyotyping in Down's syndrome screening: cost-effectiveness and detection of clinically significant chromosome abnormalities

    PubMed Central

    Gekas, Jean; van den Berg, David-Gradus; Durand, Audrey; Vallée, Maud; Wildschut, Hajo Izaäk Johannes; Bujold, Emmanuel; Forest, Jean-Claude; Rousseau, François; Reinharz, Daniel

    2011-01-01

    In all, 80% of antenatal karyotypes are generated by Down's syndrome screening programmes (DSSP). After a positive screening, women are offered prenatal foetus karyotyping, the gold standard. Reliable molecular methods for rapid aneuploidy diagnosis (RAD: fluorescence in situ hybridization (FISH) and quantitative fluorescence PCR (QF-PCR)) can detect common aneuploidies, and are faster and less expensive than karyotyping. In the UK, RAD is recommended as a standalone approach in DSSP, whereas the US guidelines recommend that RAD be followed up by karyotyping. A cost-effectiveness (CE) analysis of RAD in various DSSP is lacking. There is a debate over the significance of chromosome abnormalities (CA) detected with karyotyping but not using RAD. Our objectives were to compare the CE of RAD versus karyotyping, to evaluate the clinically significant missed CA and to determine the impact of detecting the missed CA. We performed computer simulations to compare six screening options followed by FISH, PCR or karyotyping using a population of 110 948 pregnancies. Among the safer screening strategies, the most cost-effective strategy was contingent screening with QF-PCR (CE ratio of $24 084 per Down's syndrome (DS) detected). Using karyotyping, the CE ratio increased to $27 898. QF-PCR missed only six clinically significant CA of which only one was expected to confer a high risk of an abnormal outcome. The incremental CE ratio (ICER) to find the CA missed by RAD was $66 608 per CA. These costs are much higher than those involved for detecting DS cases. As the DSSP are mainly designed for DS detection, it may be relevant to question the additional costs of karyotyping. PMID:20842178

  20. Rapid testing versus karyotyping in Down's syndrome screening: cost-effectiveness and detection of clinically significant chromosome abnormalities.

    PubMed

    Gekas, Jean; van den Berg, David-Gradus; Durand, Audrey; Vallée, Maud; Wildschut, Hajo Izaäk Johannes; Bujold, Emmanuel; Forest, Jean-Claude; Rousseau, François; Reinharz, Daniel

    2011-01-01

    In all, 80% of antenatal karyotypes are generated by Down's syndrome screening programmes (DSSP). After a positive screening, women are offered prenatal foetus karyotyping, the gold standard. Reliable molecular methods for rapid aneuploidy diagnosis (RAD: fluorescence in situ hybridization (FISH) and quantitative fluorescence PCR (QF-PCR)) can detect common aneuploidies, and are faster and less expensive than karyotyping.In the UK, RAD is recommended as a standalone approach in DSSP, whereas the US guidelines recommend that RAD be followed up by karyotyping. A cost-effectiveness (CE) analysis of RAD in various DSSP is lacking. There is a debate over the significance of chromosome abnormalities (CA) detected with karyotyping but not using RAD. Our objectives were to compare the CE of RAD versus karyotyping, to evaluate the clinically significant missed CA and to determine the impact of detecting the missed CA. We performed computer simulations to compare six screening options followed by FISH, PCR or karyotyping using a population of 110948 pregnancies. Among the safer screening strategies, the most cost-effective strategy was contingent screening with QF-PCR (CE ratio of $24084 per Down's syndrome (DS) detected). Using karyotyping, the CE ratio increased to $27898. QF-PCR missed only six clinically significant CA of which only one was expected to confer a high risk of an abnormal outcome. The incremental CE ratio (ICER) to find the CA missed by RAD was $66608 per CA. These costs are much higher than those involved for detecting DS cases. As the DSSP are mainly designed for DS detection, it may be relevant to question the additional costs of karyotyping.

  1. A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes

    PubMed Central

    Diaz-Perez, Silvia V.; Ferguson, David O.; Wang, Chen; Csankovszki, Gyorgyi; Wang, Chengming; Tsai, Shih-Chang; Dutta, Devkanya; Perez, Vanessa; Kim, SunMin; Eller, C. Daniel; Salstrom, Jennifer; Ouyang, Yan; Teitell, Michael A.; Kaltenboeck, Bernhard; Chess, Andrew; Huang, Sui; Marahrens, York

    2006-01-01

    The inactive X chromosome of female mammals displays several properties of heterochromatin including late replication, histone H4 hypoacetylation, histone H3 hypomethylation at lysine-4, and methylated CpG islands. We show that cre-Lox-mediated excision of 21 kb from both Xist alleles in female mouse fibroblasts led to the appearance of two histone modifications throughout the inactive X chromosome usually associated with euchromatin: histone H4 acetylation and histone H3 lysine-4 methylation. Despite these euchromatic properties, the inactive X chromosome was replicated even later in S phase than in wild-type female cells. Homozygosity for the deletion also caused regions of the active X chromosome that are associated with very high concentrations of LINE-1 elements to be replicated very late in S phase. Extreme late replication is a property of fragile sites and the 21-kb deletions destabilized the DNA of both X chromosomes, leading to deletions and translocations. This was accompanied by the phosphorylation of p53 at serine-15, an event that occurs in response to DNA damage, and the accumulation of γ-H2AX, a histone involved in DNA repair, on the X chromosome. The Xist locus therefore maintains the DNA stability of both X chromosomes. PMID:16980402

  2. Sparse representation and Bayesian detection of genome copy number alterations from microarray data

    PubMed Central

    Pique-Regi, Roger; Monso-Varona, Jordi; Ortega, Antonio; Seeger, Robert C.; Triche, Timothy J.; Asgharzadeh, Shahab

    2008-01-01

    Motivation: Genomic instability in cancer leads to abnormal genome copy number alterations (CNA) that are associated with the development and behavior of tumors. Advances in microarray technology have allowed for greater resolution in detection of DNA copy number changes (amplifications or deletions) across the genome. However, the increase in number of measured signals and accompanying noise from the array probes present a challenge in accurate and fast identification of breakpoints that define CNA. This article proposes a novel detection technique that exploits the use of piece wise constant (PWC) vectors to represent genome copy number and sparse Bayesian learning (SBL) to detect CNA breakpoints. Methods: First, a compact linear algebra representation for the genome copy number is developed from normalized probe intensities. Second, SBL is applied and optimized to infer locations where copy number changes occur. Third, a backward elimination (BE) procedure is used to rank the inferred breakpoints; and a cut-off point can be efficiently adjusted in this procedure to control for the false discovery rate (FDR). Results: The performance of our algorithm is evaluated using simulated and real genome datasets and compared to other existing techniques. Our approach achieves the highest accuracy and lowest FDR while improving computational speed by several orders of magnitude. The proposed algorithm has been developed into a free standing software application (GADA, Genome Alteration Detection Algorithm). Availability: http://biron.usc.edu/~piquereg/GADA Contact: jpei@chop.swmed.edu and rpique@ieee.org Supplementary information: Supplementary data are available at Bioinformatics online. PMID:18203770

  3. The human Y chromosome.

    PubMed Central

    Goodfellow, P; Darling, S; Wolfe, J

    1985-01-01

    long arm and sequences shared with autosomes. About two-thirds of XX males contain detectable Y derived sequences. As the amount of Y sequences present varies in different XX males, DNA from these subjects can be used to construct a map of the region around the sex determining gene. Assuming that XX males are usually caused by simple translocation, the sex determining genes cannot be located in the pericentric region. Although conventional genetic analysis of the Y chromosome is difficult, this chromosome is particularly suited to molecular analysis. Paradoxically, the Y chromosome may soon become the best defined human chromosome at the molecular level and may become the model for other chromosomes. Images PMID:3908683

  4. Spectral karyotyping identifies recurrent complex rearrangements of chromosomes 8, 17, and 20 in osteosarcomas.

    PubMed

    Bayani, Jane; Zielenska, Maria; Pandita, Ajay; Al-Romaih, Khaldoun; Karaskova, Jana; Harrison, Karen; Bridge, Julia A; Sorensen, Poul; Thorner, Paul; Squire, Jeremy A

    2003-01-01

    Conventional cytogenetic studies have shown that osteosarcomas (OSs) are often highly aneuploid, with a large number of both structural and numerical chromosomal alterations. To investigate the complexity of OS karyotypes in detail, we applied spectral karyotyping (SKY) to a series of 14 primary OS tumors and four established OS cell lines. A total of 531 rearrangements were identified by SKY, of which 300 breakpoints could be assigned to a specific chromosome band. There was an average of 38.5 breakpoints identified by SKY per primary tumor. Chromosome 20 was involved in a disproportionately high number of structural rearrangements, with 38 different aberrations being detected. Chromosomal rearrangements between chromosomes 20 and 8 were evident in four tumors. FISH analysis using a 20q13 subtelomeric probe identified frequent involvement of 20q in complex structural rearrangements of OS cell lines. Characterization of the structural aberrations of chromosomes 8 and 17 by use of SKY demonstrated frequent duplication or partial gains of chromosome bands 8q23-24 and 17p11-13. Other chromosomes frequently involved in structural alteration were chromosomes 1 (47 rearrangements) and 6 (38 rearrangements). Centromeric rearrangements often involving chromosomes 1, 6, 13, 14, 17, and 20 were present. Four of the 14 primary OS tumors were characterized by nonclonal changes that included both structural and numerical alterations. In summary, OS tumors have a very high frequency of structural and numerical alterations, compounded by gross changes in ploidy. This intrinsic karyotype instability leads to a diversity of rearrangements and the acquisition of composite chromosomal rearrangements, with the highest frequency of alteration leading to gain of 8q23-24 and 17p11-13 and rearrangement of 20q. These findings suggest that specific sequences mapping to these chromosomal regions will likely have a role in the development and progression of OS.

  5. Diffusion MRI and the detection of alterations following traumatic brain injury.

    PubMed

    Hutchinson, Elizabeth B; Schwerin, Susan C; Avram, Alexandru V; Juliano, Sharon L; Pierpaoli, Carlo

    2017-06-13

    This article provides a review of brain tissue alterations that may be detectable using diffusion magnetic resonance imaging MRI (dMRI) approaches and an overview and perspective on the modern dMRI toolkits for characterizing alterations that follow traumatic brain injury (TBI). Noninvasive imaging is a cornerstone of clinical treatment of TBI and has become increasingly used for preclinical and basic research studies. In particular, quantitative MRI methods have the potential to distinguish and evaluate the complex collection of neurobiological responses to TBI arising from pathology, neuroprotection, and recovery. dMRI provides unique information about the physical environment in tissue and can be used to probe physiological, architectural, and microstructural features. Although well-established approaches such as diffusion tensor imaging are known to be highly sensitive to changes in the tissue environment, more advanced dMRI techniques have been developed that may offer increased specificity or new information for describing abnormalities. These tools are promising, but incompletely understood in the context of TBI. Furthermore, model dependencies and relative limitations may impact the implementation of these approaches and the interpretation of abnormalities in their metrics. The objective of this paper is to present a basic review and comparison across dMRI methods as they pertain to the detection of the most commonly observed tissue and cellular alterations following TBI. © 2017 The Authors Journal of Neuroscience Research Published by Wiley Periodicals, Inc.

  6. Chromosomal Conditions

    MedlinePlus

    ... 150 babies is born with a chromosomal condition. Down syndrome is an example of a chromosomal condition. Because ... all pregnant women be offered prenatal tests for Down syndrome and other chromosomal conditions. A screening test is ...

  7. AB056. Establishing the procedure for detection of gr/gr deletions on the Y chromosome in Vietnamese infertile men

    PubMed Central

    Nguyễn, Trâm Bảo; Nguyễn, Ngọc-Anh Thị; Phạm, Thảo-Trang Nguyễn; Nguyễn, Trung Tấn

    2015-01-01

    Background and objective About 2-10% cases of both azoospermia or cryptospermia have a micro Y deletion. These deletions mainly locate at AZFa, b, or c which containing several crucial genes for normal spermatogenesis. Among various category of deletions is gr/gr deletions on AZFc. The influence on spermatogenesis mostly depend on characteristics of the Y chromosome which may vary among different ethnicity and geographical locations. In Vietnam, there is no study about gr/gr deletions. (I) To establish an approachable procedure to detect gr/gr deletions on the Y chromosome, which is applicable for hospitals and clinics in Vietnam; (II) to determine the percentage of gr/gr deletions carriers among infertile men (azoospermia and severe oligozoospermia). Methods Blood samples were collected for DNA extraction. Based on inclusive and exclusive criteria, 3 fertile and 32 infertile men (azoospermia and cryptozoospermia) were recruited to our study with informed consent. Sequence tagged sites (STSs) và primers were designed by using design software (Ape, PrimerPlex2, Oligoanalyzer IDT) based on the information of NCBI. DNA was extracted in silico from blood sample and applied to Multiplex PCR to detect gr/gr deletions. To ensure the true positive result of gr/gr deletions, MLPA was subjected to perform. Results and discussion With our designed primers, an approachable procedure was successfully established to detect gr/gr deletions that resulted at a high level of sensitivity (at 0.5ng DNA sample/reaction) and high level of stability. The percentage of gr/gr-deletion carriers among infertile men was 12.5% (4 positive samples), higher than 10.6% in an Asian population from a current study. All positive samples are confirmed significantly by MLPA technique. Conclusions An approachable procedure was successfully established to detect gr/gr deletions that resulted at a high level of sensitivity (at 0.5 ng DNA sample/reaction) and high level of stability. The percentage of gr

  8. Hyperspectral hybrid method classification for detecting altered mucosa of the human larynx

    PubMed Central

    2012-01-01

    Background In the field of earth observation, hyperspectral detector systems allow precise target detections of surface components from remote sensing platforms. This enables specific land covers to be identified without the need to physically travel to the areas examined. In the medical field, efforts are underway to develop optical technologies that detect altering tissue surfaces without the necessity to perform an excisional biopsy. With the establishment of expedient classification procedures, hyperspectral imaging may provide a non-invasive diagnostic method that allows determination of pathological tissue with high reliability. In this study, we examined the performance of a hyperspectral hybrid method classification for the automatic detection of altered mucosa of the human larynx. Materials and methods Hyperspectral Imaging was performed in vivo and 30 bands from 390 to 680 nm for 5 cases of laryngeal disorders (2x hemorrhagic polyp, 3x leukoplakia) were obtained. Image stacks were processed with unsupervised clustering (linear spectral unmixing), spectral signatures were extracted from unlabeled cluster maps and subsequently applied as end-members for supervised classification (spectral angle mapper) of further medical cases with identical diagnosis. Results Linear spectral unmixing clearly highlighted altered mucosa as single spectral clusters in all cases. Matching classes were identified, and extracted spectral signatures could readily be applied for supervised classifications. Automatic target detection performed well, as the considered classes showed notable correspondence with pathological tissue locations. Conclusions Using hyperspectral classification procedures derived from remote sensing applications for diagnostic purposes can create concrete benefits for the medical field. The approach shows that it would be rewarding to collect spectral signatures from histologically different lesions of laryngeal disorders in order to build up a spectral

  9. Exercise-induced hemostatic alterations are detectable by rotation thrombelastography (ROTEM): A marathon study.

    PubMed

    Sucker, Christoph; Zotz, Rainer B; Senft, Beate; Scharf, Rudiger E; Kröger, Knut; Erbel, Raimund; Möhlenkamp, Stefan

    2010-10-01

    Rotation thrombelastography (ROTEM) provides a whole blood assay that allows the assessment of plasmic- and platelet-related hemostasis in a single-step procedure. In our current study, we focused on the capability of the method to detect hemostatic alterations induced by physical exercise, enrolling 33 healthy participants of the Dusseldorf Marathon 2006. Venous blood drawn immediately before and after finishing the marathon was analyzed by a rotational thrombelastograph (Pentapharm, Munich, Germany). On initiation of blood coagulation by recalcification, standard ROTEM parameters were determined. Comparison of the results obtained before and after the physical exercise was performed using the Student t test for paired samples. As a result, the mean clotting time (CT) determined from blood samples obtained immediately after the marathon was significantly shorter (662.9 + or - 67.8 seconds vs 505.6 + or - 97.3 seconds, P = .002) and the mean maximal clot firmness was significantly broader (48.4 +/- 6.6 mm vs 51.5 +/- 4.5 mm, P = .0004) when compared to results obtained before the physical exercise. Differences between mean clot formation times (CFTs; 280.6 + 96 seconds vs 270.4 + or - 73.8 seconds) and mean alpha angles (45.9 degrees + or - 8 degrees vs 47.8 degrees + or - 5.8 degrees ) before and after the marathon were not statistically significant. Remarkably, some participants showed opposed results, particularly prolongation of CT and narrowing of maximum clot firmness (MCF). Our study demonstrates that ROTEM is sensitive to exercise-induced hemostatic alterations. The method appears to be capable of detecting even distinct changes in hemostasis in a single-step procedure. Further analyses are needed to clarify which hemostasis parameters influence ROTEM results and which ROTEM results are independent predictors of exercise-induced alterations of plasmic and platelet function. This might help to explain interindividual differences in exercise

  10. Two mutations in the locus control region hypersensitivity site-2 (5' HS-2) of haplotype 19 beta s chromosomes alter binding of trans-acting factors.

    PubMed

    Morgan, J C; Scott, D F; Lanclos, K D

    1996-01-01

    There are five major haplotypes associated with sickle cell anemia (SS). Individuals homozygous for haplotypes 3 (Senegal) and 31 (Saudi Arabian) have high fetal hemoglobin (HbF) levels (15 to 30% of total hemoglobin) whereas individuals homozygous for haplotypes 17 (Cameroon), 19 (Benin), and 20 (Bantu) have low HbF levels (1 to 10%). We previously identified several point mutations in the LCR 5'HS-2 that were specific for haplotype 19 beta s chromosomes (compared to the GenBank HUMHBB reference sequence, T-->G at position 8580, A-->G at position 8598, and A-->T at position 9114). We postulated that one or more of these mutations may alter the binding of specific trans-acting factors and ultimately affect the expression of HbF in these sickle cell patients. We performed gel mobility shift assays using 32P-end-labeled double-stranded 19mers corresponding to each of the LCR 5'HS-2 normal (GenBank) and mutant sequences. Nuclear extracts prepared from HeLa and HEL cells were used in our experiments and neither the normal nor mutant sequence at position 8580 bound trans-acting factors in either nuclear extract. The 8598 mutant increased binding of Sp1; using purified protein and both nuclear extracts. HEL extracts were used to quantify the increase in Sp1 binding to the 8598 mutation and we found an increase in binding of 66 and 47%, respectively, in two shifted bands. The 9114 mutation sharply decreased binding of an unknown trans-acting factor by 74%. This factor was present in both HeLa and HEL nuclear extracts.

  11. Satellite detection of vegetative damage and alteration caused by pollutants emitted by a zinc smelter

    NASA Technical Reports Server (NTRS)

    Mcmurtry, G. J.; Petersen, G. W. (Principal Investigator); Fritz, E. L.; Pennypacker, S. P.

    1974-01-01

    The author has identified the following significant results. Field observations and data collected by low flying aircraft were used to verify the accuracy of maps produced from the satellite data. Although areas of vegetation as small as six acres can accurately be detected, a white pine stand that was severely damaged by sulfur dioxide could not be differentiated from a healthy white pine stand because spectral differences were not large enough. When winter data were used to eliminate interference from herbaceous and deciduous vegetation, the damage was still undetectable. The analysis was able to produce a character map that accurately delineated areas of vegetative alteration due to high zinc levels accumulating in the soil. The map depicted a distinct gradient of less damage and alteration as the distance from the smelter increased. Although the satellite data will probably not be useful for detecting small acreages of damaged vegetation, it is concluded that the data may be very useful as an inventory tool to detect and delineate large vegetative areas possessing differing spectral signatures.

  12. Comparison of methods to detect copy number alterations in cancer using simulated and real genotyping data

    PubMed Central

    2012-01-01

    Background The detection of genomic copy number alterations (CNA) in cancer based on SNP arrays requires methods that take into account tumour specific factors such as normal cell contamination and tumour heterogeneity. A number of tools have been recently developed but their performance needs yet to be thoroughly assessed. To this aim, a comprehensive model that integrates the factors of normal cell contamination and intra-tumour heterogeneity and that can be translated to synthetic data on which to perform benchmarks is indispensable. Results We propose such model and implement it in an R package called CnaGen to synthetically generate a wide range of alterations under different normal cell contamination levels. Six recently published methods for CNA and loss of heterozygosity (LOH) detection on tumour samples were assessed on this synthetic data and on a dilution series of a breast cancer cell-line: ASCAT, GAP, GenoCNA, GPHMM, MixHMM and OncoSNP. We report the recall rates in terms of normal cell contamination levels and alteration characteristics: length, copy number and LOH state, as well as the false discovery rate distribution for each copy number under different normal cell contamination levels. Assessed methods are in general better at detecting alterations with low copy number and under a little normal cell contamination levels. All methods except GPHMM, which failed to recognize the alteration pattern in the cell-line samples, provided similar results for the synthetic and cell-line sample sets. MixHMM and GenoCNA are the poorliest performing methods, while GAP generally performed better. This supports the viability of approaches other than the common hidden Markov model (HMM)-based. Conclusions We devised and implemented a comprehensive model to generate data that simulate tumoural samples genotyped using SNP arrays. The validity of the model is supported by the similarity of the results obtained with synthetic and real data. Based on these results and

  13. Use of monoclonal antibodies in the detection of structural alterations occurring in lysozyme on heating.

    PubMed

    Kenett, D; Katchalski-Katzir, E; Fleminger, G

    1990-01-01

    Seven murine anti-hen egg-white lysozyme (HEL) monoclonal antibodies (MAbs), which recognize distinct epitopes of the native enzyme, were used as macromolecular probes to detect structural or conformational alterations occurring in HEL on heating at 95 degrees C, pH 5. As the interactions of the heat-treated HEL with its corresponding MAbs were carried out at room temperature, only irreversible structural and/or conformational alterations could be detected. The transformation of the native enzyme into its denatured form was followed electrophoretically and chromatographically. The denatured enzyme was more negatively charged at pH 8.4 and exhibited a longer retention time on reverse-phase HPLC than native HEL. Its specific catalytic activity was considerably lower than that of the native enzyme. Of the seven MAbs tested in competitive ELISA assays with native and heat-treated HEL only one, MAb D74.3, failed to recognize the heat-treated enzyme. This antibody, which is directed toward the active site region of the enzyme, was ineffective in inhibiting the catalytic activity of the heat-treated HEL using M. lysodeikticus as substrate. In contrast, the monoclonal antibody D1.3, which recognizes an epitope remote from the active site of HEL, inhibited the catalytic activity of the native as well as the heat-treated enzyme. The results indicate that the active site of HEL undergoes an irreversible structural alteration on heating for 2 hr at 95 degrees C, pH 5. No irreversible structural changes could be detected in the other regions of HEL recognized by the corresponding MAbs.

  14. Polymerase chain reaction detection of Y chromosome sequences in vaginal fluid: preliminary studies of a potential biomarker for sexual behavior.

    PubMed

    Zenilman, Jonathan M; Yuenger, Jeffrey; Galai, Noya; Turner, Charles F; Rogers, Susan M

    2005-02-01

    Self-reported measures of sexual behavior are subject to nontrivial reporting biases. The objective of this study was to develop a behavioral biomarker of recent sexual activity among females that is inexpensive, easily administered, and can be used in low sexually transmitted disease prevalence populations. We developed a polymerase chain reaction (PCR) assay to detect Y chromosome (Yc) fragments. The Yc primers were developed against a 200-basepair (bp) microsatellite repeat sequence, which is unique to the male genome. A standard PCR technique was used. Assay sensitivity was determined quantitatively using donated semen samples. To assess longevity of detectability, we recruited female subjects in monogamous relationships. Seventeen subjects had unprotected intercourse followed by 3 weeks of abstinence from vaginal intercourse. Self-administered vaginal swabs (SAVS) were collected every other day. In addition to the swabs, subjects kept daily sexual diaries. Swabs were processed by semiquantitative PCR, and Yc decay curves were determined for each subject. The half-life of Yc in vaginal fluid was calculated on the collection of individual decay curves by a random-effects regression model approach. The sensitivity of our Yc-PCR assay was determined to be 5 copies of Yc. In the longevity studies, Yc was detectable in SAVS up to 15 postcoital days (PCD). Mean Yc DNA concentration in SAVS eluate followed an exponential decay pattern for each subject. Mean concentrations were 66.7 ng/mL at PCD-1, 20.6 ng/mL at PCD-7, and 4.5 ng/mL at PCD-15. The estimated half-life for Yc clearance was 3.83 days. The swab-based Yc-DNA PCR assay can detect coitus in women for a 2-week retrospective period. This can be used to validate sexual behavior-reporting and condom use in women and promises to be a useful tool in sexual behavior research.

  15. Feature Selection for the Automated Detection of Metaphase Chromosomes: Performance Comparison Using a Receiver Operating Characteristic Method

    PubMed Central

    Qiu, Yuchen; Song, Jie; Lu, Xianglan; Li, Yuhua; Zheng, Bin; Li, Shibo; Liu, Hong

    2014-01-01

    Background. The purpose of this study is to identify a set of features for optimizing the performance of metaphase chromosome detection under high throughput scanning microscopy. In the development of computer-aided detection (CAD) scheme, feature selection is critically important, as it directly determines the accuracy of the scheme. Although many features have been examined previously, selecting optimal features is often application oriented. Methods. In this experiment, 200 bone marrow cells were first acquired by a high throughput scanning microscope. Then 9 different features were applied individually to group captured images into the clinically analyzable and unanalyzable classes. The performance of these different methods was assessed by a receiving operating characteristic (ROC) method. Results. The results show that using the number of labeled regions on each acquired image is suitable for the first on-line CAD scheme. For the second off-line CAD scheme, it would be suggested to combine four feature extraction methods including the number of labeled regions, average regions area, average region pixel value, and the standard deviation of either region distance or circularity. Conclusion. This study demonstrates an effective method of feature selection and comparison to facilitate the optimization of the CAD schemes for high throughput scanning microscope in the future. PMID:25763334

  16. Multicolor banding detects a complex three chromosome, seven breakpoint unbalanced rearrangement in an ICSI-derived fetus with multiple abnormalities.

    PubMed

    Seller, Mary J; Bint, Susan; Kavalier, Fred; Brown, Richard N; Ogilvie, Caroline Mackie

    2006-05-15

    We describe a fetus from an intracytoplasmic sperm injection (ICSI) pregnancy with severe facial clefts, receding jaw, preauricular skin tags, postaxial hexadactyly, bi-lobed right lung, supernumerary cranial bone, and dilated lateral ventricles of the brain. Using a combination of G-banding, fluorescence in situ hybridization (FISH), whole chromosome paints (WCPs), subtelomere probes, and multicolor banding (MCB), the karyotype was found to include a de novo unbalanced highly complex chromosome rearrangement (hCCR) involving chromosomes 3, 12, and 15 with seven breakpoints, and including monosomy for two separate regions of chromosome 12.

  17. Isovolumetric elasticity alteration in the human heart detected by in vivo time-harmonic elastography.

    PubMed

    Tzschätzsch, Heiko; Hättasch, Robert; Knebel, Fabian; Klaua, Robert; Schultz, Michael; Jenderka, Klaus-Vitold; Braun, Jürgen; Sack, Ingolf

    2013-12-01

    Time harmonic elastography (THE) has recently been introduced for measurement of the periodic alteration in myocardial shear modulus based on externally induced low-frequency acoustic vibrations produced by a loudspeaker. In this study, we propose further developments of cardiac THE toward a clinical modality including integration of the vibration source into the patient bed and automated parameter extraction from harmonic shear wave amplitudes, wall motion profiles and synchronized electrocardiographic records. This method has enabled us to evaluate the delay between wall motion and wave amplitude alteration for the measurement of isovolumetric times of elasticity alteration during contraction (τ(C)) and relaxation (τ(R)) in a group of 32 healthy volunteers. On average, the wave amplitudes changed between systole and diastole by a factor of 1.7 ± 0.3, with a τ(C) of 137 ± 61 ms and a τ(R) of 68 ± 73 ms, which agrees with results obtained with the more time-consuming and expensive cardiac magnetic resonance elastography. Furthermore, because of the high sampling rate, elasto-morphometric parameters such as transition times and the area of wave amplitude-cardiac motion cycles can be processed in an automated way for the future clinical detection of myocardial relaxation abnormalities.

  18. Diagnostic accuracy of the BACs-on-Beads™ assay versus karyotyping for prenatal detection of chromosomal abnormalities: a retrospective consecutive case series.

    PubMed

    Choy, K W; Kwok, Y K; Cheng, Y K Y; Wong, K M; Wong, H K; Leung, K O; Suen, K W; Adler, K; Wang, C C; Lau, T K; Schermer, M J; Lao, T T; Leung, T Y

    2014-09-01

    To evaluate the diagnostic performance of the BACs-on-Beads(™) (BoBs(™)) assay for prenatal detection of chromosomal abnormalities. Retrospective study. Tertiary prenatal diagnosis centre. Women referred for prenatal diagnosis. We retrieved 2153 archived DNA samples collected between January 2010 and August 2011 for the BoBs(™) assay. These samples had previously been tested by quantitative fluorescence polymerase chain reaction (QF-PCR) and karyotyping. In the BoBs(™) assay a sample was defined as normal disomic when the ratio of the fluorescence intensities in a chromosome locus lay within the threshold (mean ratio ± 2SD), and as deleted or duplicated when the ratio was below the lower threshold (0.6-0.8) or above the upper threshold (1.3-1.4), respectively. The BoBs(™) results were further validated by microarray and compared in a blinded manner with the original QF-PCR and karyotyping results. Concordance of any numerical, structural, and submicroscopic chromosomal abnormalities between the methods. BACs-on-Beads(™) was similar to karyotyping and QF-PCR in detecting trisomy 13, trisomy 18, trisomy 21, and sex chromosomal aneuploidies, and superior to QF-PCR in detecting major structural abnormalities (53.3 versus 13.3%) and mosaicism (28.6 versus 0%) involving chromosomal abnormalities other than the common aneuploidies. BoBs(™) detected six microdeletion syndromes missed by karyotyping and QF-PCR; however, BoBs(™) missed two cases of triploidy identified by QF-PCR. Therefore, the sensitivity of BoBs(™) is 96.7% (95% CI 92.6-98.7%), and its specificity is 100% (95% CI 99.8-100%). BACs-on-Beads(™) can replace QF-PCR for triaging in prenatal diagnosis, and gives a better diagnostic yield than current rapid aneuploidy tests. © 2014 Royal College of Obstetricians and Gynaecologists.

  19. Detection of aneuploid human sperm by fluorescence in situ hybridization: Evidence for a donor difference in frequency of sperm disomic for chromosomes 1 and Y

    SciTech Connect

    Robbins, W.A. Lawrence Livermore National Lab., CA ); Segraves, R.; Pinkel, D. ); Wyrobek, A.J. )

    1993-04-01

    Fluorescence in situ hybridization with repetitive-sequence DNA probes was used to detect human sperm disomic for chromosomes 1 and Y in three healthy men. Data on these same men had been obtained previously, using the human-sperm/hamster-egg cytogenetic technique, providing a cytogenetic reference for validating sperm hybridization measurements. Air-dried smears were prepared from semen samples and treated with DTT and lithium diiodosalicylate to expand sperm chromatin. Hybridization with fluorescently tagged DNA probes for chromosomes 1 (pUC177) or Y (pY3.4) yielded average frequencies of sperm with two fluorescent domains of 14.2[+-]2.4/10,000 and 5.6[+-]1.6/10,000 sperm, respectively. These frequencies did not differ statistically from frequencies of hyperploidy observed for these chromosomes with the hamster technique. In addition, frequencies of disomic sperm from one donor were elevated [approximately]2.5-fold above those of other donors, for both chromosomes 1 (P = .045) and Y (P = .01), consistent with a trend found with the hamster technique. The authors conclude that fluorescence in situ hybridization to sperm chromosomes provides a valid and promising measure of the frequency of disomic human sperm. 43 refs., 1 fig., 4 tabs.

  20. Specificity of inhibitory KIRs enables NK cells to detect changes in an altered peptide environment.

    PubMed

    Carrillo-Bustamante, Paola; de Boer, Rob J; Keşmir, Can

    2017-07-10

    The activity of natural killer (NK) cells is tightly regulated by inhibitory and activating receptors. Inhibitory killer immunoglobulin-like receptors (iKIRs) survey the surface of target cells by monitoring the expression of human leukocyte antigen (HLA) class I. The binding of iKIRs has been shown to be sensitive to the peptides presented by HLA class I, implying that iKIRs have the ability to detect the changes in the repertoire of peptide-HLA class I complexes (pHLA), a process occurring during viral infection and in tumor cells. To study how the pHLA repertoire changes upon infection, and whether an iKIR is able to detect these changes, we study peptides eluted from cells prior and after infection with measles virus (MV). Remarkably, most changes in the repertoire of potential iKIR ligands are predicted to be caused by the altered expression of self-peptides. We show that an iKIR can detect these changes in the presented peptides only if it is sufficiently specific, e.g., if iKIRs can distinguish between different amino acids in the contact residues (e.g., position 7 and 8). Our analysis further indicates that one single iKIR per host is not sufficient to detect changes in the peptide repertoire, suggesting that a multigene family encoding for different iKIRs is required for successful peptide recognition.

  1. Genetic aspects of human male infertility: the frequency of chromosomal abnormalities and Y chromosome microdeletions in severe male factor infertility.

    PubMed

    Vicdan, Arzu; Vicdan, Kubilay; Günalp, Serdar; Kence, Aykut; Akarsu, Cem; Işik, Ahmet Zeki; Sözen, Eran

    2004-11-10

    The main purpose of this study is to detect the frequency and type of both chromosomal abnormalities and Y chromosome microdeletions in patients with severe male factor infertility and fertile control subjects. The association between the genetic abnormality and clinical parameters was also evaluated. This study was carried out in 208 infertile and 20 fertile men. Results of 208 patients, 119 had non-obstructive azoospermia and 89 had severe oligoasthenoteratozoospermia (OAT). Seventeen out of 119 (14.3%) azoospermic patients and two out of 89 (2.2%) patients with OAT had Y chromosome microdeletions. In total, 19 cases with deletions were detected in 208 infertile men, with a frequency of 9.1%. The AZFc locus, mainly DAZ gene cluster was the most frequently deleted region. Five other cases with azoospermia (4.2%) and two cases with OAT (2.2%) had a chromosomal abnormality, with a total number of seven (3.4%). Including Y chromosome deletions and structural chromosome abnormalities, the rate of genetic abnormalities was 12.5% (26/208) in our patients. On the other hand, 20 men with proven fertility and fathers of five cases with microdeletions were genetically normal. Y chromosome deletions and chromosomal abnormalities were associated with various histological alterations in testis. Sertoli cell-only (SCO) syndrome and maturation arrest predominated in these cases, whereas hypospermatogenesis occurred more frequently in genetically normal patients. Various chromosomal abnormalities and deletions of Y chromosome can cause spermatogenic breakdown resulting in chromosomally derived infertility. All these findings strongly support the recommendation of genetic screening of infertile patients.

  2. Rapid Simultaneous Amplification and Detection of the MBR/JH Chromosomal Translocation by Fluorescence Melting Curve Analysis

    PubMed Central

    Bohling, Sandra D.; King, Thomas C.; Wittwer, Carl T.; Elenitoba-Johnson, Kojo S. J.

    1999-01-01

    Polymerase chain reaction (PCR) amplification and product analysis for the detection of chromosomal translocations, such as the t(14;18), has traditionally been a two-step process. PCR product detection has generally entailed gel electrophoresis and/or hybridization or sequencing for confirmation of assay specificity. Using a microvolume fluorimeter integrated with a thermal cycler and a PCR-compatible double-stranded DNA (dsDNA) binding fluorescent dye (SYBR Green I), we investigated the feasibility of simultaneous thermal amplification and detection of MBR/JH translocation products by fluorescence melting curve analysis. We analyzed DNA from 30 cases of lymphoproliferative disorders comprising 19 cases of previously documented MBR/JH-positive follicle center lymphoma and 11 reactive lymphadenopathies. The samples were coded and analyzed blindly for the presence of MBR/JH translocations by fluorescence melting curve analysis. We also performed dilutional assays using the MBR/JH-positive cell line SUDHL-6. Multiplex PCR for MBR/JH and β-globin was used to simultaneously assess sample adequacy. All (100%) of the 19 cases previously determined to be MBR/JH positive by conventional PCR analysis showed a characteristic sharp decrease in fluorescence at ∼90°C by melting curve analysis after amplification. Fluorescence melting peaks obtained by plotting the negative derivative of fluorescence over temperature (−dF/dT) versus temperature (T) showed melting temperatures (Tm) at 88.85 ± 1.15°C. In addition, multiplex assays using both MBR/JH and β-globin primers yielded easily distinguishable fluorescence melting peaks at ∼90°C and 81.2°C, respectively. Dilutional assays revealed that fluorescence melting curve analysis was more sensitive than conventional PCR and agarose gel electrophoresis with ultraviolet transillumination by as much as 100-fold. Simultaneous amplification and fluorescence melting curve analysis is a simple, reliable, and sensitive method

  3. Prenatally detected de novo apparently balanced chromosomal rearrangements: the effect on maternal worry, family functioning and intent of disclosure.

    PubMed

    Sinnerbrink, Ingrid B; Meiser, Bettina; Halliday, Jane; Sherwen, Amanda; Amor, David J; Waters, Elizabeth; Rea, Felicity; Evans, Elizabeth; Rahman, Belinda; Kirk, Edwin P

    2014-06-01

    This study aims to assess the impact of prenatal diagnosis of de novo apparently balanced chromosome rearrangements (ABCRs) on maternal stress, family functioning and maternal plans of disclosure of genetic information to their child. All liveborn children with prenatally detected de novo ABCRs in two Australian states over a 10-year period (1994-2003) were retrospectively ascertained. Of 39 eligible cases, 16 (41%) participated in the study. Mothers of these children completed a questionnaire using standardized measures to assess family functioning, parental distress, parent-child interaction and child characteristics, with open-ended questions regarding disclosure. The majority of mothers appeared to experience normal levels of parenting stress, quality of parent-child interaction and healthy family functioning. However, most mothers recalled experiencing a significant degree of worry at the time of receiving their prenatal test results, and some mothers (4/15) reported receiving uncertain or conflicting results. Most mothers (13/15) conveyed an understanding of the importance of disclosing this genetic information to their child, and 12/15 conveyed their intention to make this disclosure. Most mothers reported normal parenting stress and family functioning, despite experiencing significant worry upon receiving results. Some children are at risk of nondisclosure of their carrier status. © 2014 John Wiley & Sons, Ltd.

  4. Multiplexed chromosome conformation capture sequencing for rapid genome-scale high-resolution detection of long-range chromatin interactions.

    PubMed

    Stadhouders, Ralph; Kolovos, Petros; Brouwer, Rutger; Zuin, Jessica; van den Heuvel, Anita; Kockx, Christel; Palstra, Robert-Jan; Wendt, Kerstin S; Grosveld, Frank; van Ijcken, Wilfred; Soler, Eric

    2013-03-01

    Chromosome conformation capture (3C) technology is a powerful and increasingly popular tool for analyzing the spatial organization of genomes. Several 3C variants have been developed (e.g., 4C, 5C, ChIA-PET, Hi-C), allowing large-scale mapping of long-range genomic interactions. Here we describe multiplexed 3C sequencing (3C-seq), a 4C variant coupled to next-generation sequencing, allowing genome-scale detection of long-range interactions with candidate regions. Compared with several other available techniques, 3C-seq offers a superior resolution (typically single restriction fragment resolution; approximately 1-8 kb on average) and can be applied in a semi-high-throughput fashion. It allows the assessment of long-range interactions of up to 192 genes or regions of interest in parallel by multiplexing library sequencing. This renders multiplexed 3C-seq an inexpensive, quick (total hands-on time of 2 weeks) and efficient method that is ideal for the in-depth analysis of complex genetic loci. The preparation of multiplexed 3C-seq libraries can be performed by any investigator with basic skills in molecular biology techniques. Data analysis requires basic expertise in bioinformatics and in Linux and Python environments. The protocol describes all materials, critical steps and bioinformatics tools required for successful application of 3C-seq technology.

  5. Precise detection of chromosomal translocation or inversion breakpoints by whole-genome sequencing.

    PubMed

    Suzuki, Toshifumi; Tsurusaki, Yoshinori; Nakashima, Mitsuko; Miyake, Noriko; Saitsu, Hirotomo; Takeda, Satoru; Matsumoto, Naomichi

    2014-12-01

    Structural variations (SVs), including translocations, inversions, deletions and duplications, are potentially associated with Mendelian diseases and contiguous gene syndromes. Determination of SV-related breakpoints at the nucleotide level is important to reveal the genetic causes for diseases. Whole-genome sequencing (WGS) by next-generation sequencers is expected to determine structural abnormalities more directly and efficiently than conventional methods. In this study, 14 SVs (9 balanced translocations, 1 inversion and 4 microdeletions) in 9 patients were analyzed by WGS with a shallow (5 × ) to moderate read coverage (20 × ). Among 28 breakpoints (as each SV has two breakpoints), 19 SV breakpoints had been determined previously at the nucleotide level by any other methods and 9 were uncharacterized. BreakDancer and Integrative Genomics Viewer determined 20 breakpoints (16 translocation, 2 inversion and 2 deletion breakpoints), but did not detect 8 breakpoints (2 translocation and 6 deletion breakpoints). These data indicate the efficacy of WGS for the precise determination of translocation and inversion breakpoints.

  6. Shared Gene Expression Alterations in Nasal and Bronchial Epithelium for Lung Cancer Detection.

    PubMed

    2017-07-01

    We previously derived and validated a bronchial epithelial gene expression biomarker to detect lung cancer in current and former smokers. Given that bronchial and nasal epithelial gene expression are similarly altered by cigarette smoke exposure, we sought to determine if cancer-associated gene expression might also be detectable in the more readily accessible nasal epithelium. Nasal epithelial brushings were prospectively collected from current and former smokers undergoing diagnostic evaluation for pulmonary lesions suspicious for lung cancer in the AEGIS-1 (n = 375) and AEGIS-2 (n = 130) clinical trials and gene expression profiled using microarrays. All statistical tests were two-sided. We identified 535 genes that were differentially expressed in the nasal epithelium of AEGIS-1 patients diagnosed with lung cancer vs those with benign disease after one year of follow-up ( P  < .001). Using bronchial gene expression data from the AEGIS-1 patients, we found statistically significant concordant cancer-associated gene expression alterations between the two airway sites ( P  < .001). Differentially expressed genes in the nose were enriched for genes associated with the regulation of apoptosis and immune system signaling. A nasal lung cancer classifier derived in the AEGIS-1 cohort that combined clinical factors (age, smoking status, time since quit, mass size) and nasal gene expression (30 genes) had statistically significantly higher area under the curve (0.81; 95% confidence interval [CI] = 0.74 to 0.89, P  = .01) and sensitivity (0.91; 95% CI = 0.81 to 0.97, P  = .03) than a clinical-factor only model in independent samples from the AEGIS-2 cohort. These results support that the airway epithelial field of lung cancer-associated injury in ever smokers extends to the nose and demonstrates the potential of using nasal gene expression as a noninvasive biomarker for lung cancer detection.

  7. Insertion of reticuloendotheliosis virus long terminal repeat into a bacterial artificial chromosome clone of a very virulent Marek's disease virus alters its pathogenicity.

    PubMed

    Mays, Jody K; Silva, Robert F; Kim, Taejoong; Fadly, Aly

    2012-01-01

    Co-cultivation of the JM/102W strain of Marek's disease virus (MDV) with reticuloendotheliosis virus (REV) resulted in the generation of a recombinant MDV containing the REV long terminal repeat (LTR) named the RM1 strain of MDV, a strain that was highly attenuated for oncogenicity but induced severe bursal and thymic atrophy. We hypothesize that the phenotypic changes were solely due to the LTR insertion. Furthermore, we hypothesize that insertion of REV LTR into an analogous location in a different MDV would result in a similar phenotypic change. To test these hypotheses, we inserted the REV LTR into a bacterial artificial chromosome (BAC) clone of a very virulent strain of MDV, Md5, and designated the virus rMd5-RM1-LTR. The rMd5-RM1-LTR virus and the rMd5 virus were passaged in duck embryo fibroblast cells for up to 40 passages before pathogenicity studies. Susceptible chickens were inoculated intra-abdominally at hatch with the viruses rMd5-RM1-LTR, rMd5 BAC parental virus, wild-type strain Md5, or strain RM1 of MDV. The rMd5-RM1-LTR virus was attenuated at cell culture passage 40, whereas the rMd5 BAC without RM1 LTR retained its pathogenicity at cell culture passage 40. Using polymerase chain analysis, the RM1 LTR insert was detected in MDV isolated from buffy coat cells collected from chickens inoculated with rMd5-RM1-LTR, but only at 1 week post inoculation. The data suggest that the presence of the RM1 LTR insert within MDV genome for 1 week post inoculation with virus at hatch is sufficient to cause a reduction in pathogenicity of strain Md5 of MDV.

  8. Detection of fetal chromosomal anomalies: does nuchal translucency measurement have added value in the era of non-invasive prenatal testing?

    PubMed

    Lichtenbelt, K D; Diemel, B D M; Koster, M P H; Manten, G T R; Siljee, J; Schuring-Blom, G H; Page-Christiaens, G C M L

    2015-07-01

    The objective of this study is to determine what percentage of fetal chromosomal anomalies remains undetected when first trimester combined testing is replaced by non-invasive prenatal testing for trisomies 13, 18, and 21. We focused on the added clinical value of nuchal translucency (NT) measurement. Data on fetal karyotype, ultrasound findings, and pregnancy outcome of all pregnancies with an NT measurement ≥3.5 mm were retrospectively collected from a cohort of 25,057 singleton pregnancies in which first trimester combined testing was performed. Two hundred twenty-five fetuses (0.9 %) had an NT ≥3.5 mm. In 24 of these pregnancies, a chromosomal anomaly other than trisomy 13, 18, or 21 was detected. Eleven resulted in fetal demise, and ten showed fetal ultrasound anomalies. In three fetuses with normal ultrasound findings, a chromosomal anomaly was detected, of which one was a triple X. In three out of 25,057 pregnancies (0.01%), non-invasive prenatal testing and fetal ultrasound would have missed a chromosomal anomaly that would have been identified by NT measurement. © 2015 John Wiley & Sons, Ltd. © 2015 John Wiley & Sons, Ltd.

  9. Subtyping of renal cortical neoplasms in fine needle aspiration biopsies using a decision tree based on genomic alterations detected by fluorescence in situ hybridization

    PubMed Central

    Gowrishankar, Banumathy; Cahill, Lynnette; Arndt, Alexandra E; Al-Ahmadie, Hikmat; Lin, Oscar; Chadalavada, Kalyani; Chaganti, Seeta; Nanjangud, Gouri J; Murty, Vundavalli V; Chaganti, Raju S K; Reuter, Victor E; Houldsworth, Jane

    2014-01-01

    Objectives To improve the overall accuracy of diagnosis in needle biopsies of renal masses, especially small renal masses (SRMs), using fluorescence in situ hybridization (FISH), and to develop a renal cortical neoplasm classification decision tree based on genomic alterations detected by FISH. Patients and Methods Ex vivo fine needle aspiration biopsies of 122 resected renal cortical neoplasms were subjected to FISH using a series of seven-probe sets to assess gain or loss of 10 chromosomes and rearrangement of the 11q13 locus. Using specimen (nephrectomy)-histology as the ‘gold standard’, a genomic aberration-based decision tree was generated to classify specimens. The diagnostic potential of the decision tree was assessed by comparing the FISH-based classification and biopsy histology with specimen histology. Results Of the 114 biopsies diagnostic by either method, a higher diagnostic yield was achieved by FISH (92 and 96%) than histology alone (82 and 84%) in the 65 biopsies from SRMs (<4 cm) and 49 from larger masses, respectively. An optimized decision tree was constructed based on aberrations detected in eight chromosomes, by which the maximum concordance of classification achieved by FISH was 79%, irrespective of mass size. In SRMs, the overall sensitivity of diagnosis by FISH compared with histopathology was higher for benign oncocytoma, was similar for the chromophobe renal cell carcinoma subtype, and was lower for clear-cell and papillary subtypes. The diagnostic accuracy of classification of needle biopsy specimens (from SRMs) increased from 80% obtained by histology alone to 94% when combining histology and FISH. Conclusion The present study suggests that a novel FISH assay developed by us has a role to play in assisting in the yield and accuracy of diagnosis of renal cortical neoplasms in needle biopsies in particular, and can help guide the clinical management of patients with SRMs that were non-diagnostic by histology. PMID:24467611

  10. Breakage-fusion-bridge cycles and de novo telomere formation on broken chromosomes in maize callus cultures.

    PubMed

    Santos-Serejo, Janay A; Aguiar-Perecin, Margarida L R

    2016-06-01

    Breakpoints involved in chromosome alterations associated with heterochromatin have been detected in maize plants regenerated from callus culture. A cytogenetic analysis of plants regenerated from a maize callus was performed aiming to analyze the stability of a chromosome 7 bearing a deficiency-duplication (Df-Dp), which was interpreted as derived from a chromatid type breakage-fusion-bridge (BFB) cycle. The Df-Dp chromosome 7 was stable in mitotic and meiotic cells of the regenerated plants. Fluorescence in situ hybridization showed signals of telomeric sequences on the broken chromosome arm and provided evidence of de novo telomere formation. The stability of two types of altered chromosome 7 was investigated in C-banded metaphases from samples of the original callus that were collected during a period of 30-42 months after culture initiation. New alterations involving heterochromatic knobs of chromosomes 7 and 9 were observed. The aberrant chromosomes were stable in the subcultures, thus providing evidence of broken chromosome healing. The examination of anaphases showed the presence of bridges, which was consistent with the occurrence of BFB cycles. De novo telomere formation occurred in euchromatic and heterochromatic chromosome termini. The results point to events of chromosomal evolution that might occur in plants.

  11. Dramatyping: a generic algorithm for detecting reasonable temporal correlations between drug administration and lab value alterations

    PubMed Central

    2016-01-01

    According to the World Health Organization, one of the criteria for the standardized assessment of case causality in adverse drug reactions is the temporal relationship between the intake of a drug and the occurrence of a reaction or a laboratory test abnormality. This article presents and describes an algorithm for the detection of a reasonable temporal correlation between the administration of a drug and the alteration of a laboratory value course. The algorithm is designed to process normalized lab values and is therefore universally applicable. It has a sensitivity of 0.932 for the detection of lab value courses that show changes in temporal correlation with the administration of a drug and it has a specificity of 0.967 for the detection of lab value courses that show no changes. Therefore, the algorithm is appropriate to screen the data of electronic health records and to support human experts in revealing adverse drug reactions. A reference implementation in Python programming language is available. PMID:27042396

  12. 13C-phenylalanine breath test detects altered phenylalanine kinetics in schizophrenia patients

    PubMed Central

    Teraishi, T; Ozeki, Y; Hori, H; Sasayama, D; Chiba, S; Yamamoto, N; Tanaka, H; Iijima, Y; Matsuo, J; Kawamoto, Y; Kinoshita, Y; Hattori, K; Ota, M; Kajiwara, M; Terada, S; Higuchi, T; Kunugi, H

    2012-01-01

    Phenylalanine is an essential amino acid required for the synthesis of catecholamines including dopamine. Altered levels of phenylalanine and its metabolites in blood and cerebrospinal fluid have been reported in schizophrenia patients. This study attempted to examine for the first time whether phenylalanine kinetics is altered in schizophrenia using L-[1-13C]phenylalanine breath test (13C-PBT). The subjects were 20 chronically medicated schizophrenia patients (DSM-IV) and the same number of age- and sex-matched controls. 13C-phenylalanine (99 atom% 13C; 100 mg) was administered orally and the breath 13CO2 /12CO2 ratio was monitored for 120 min. The possible effect of antipsychotic medication (risperidone (RPD) or haloperidol (HPD) treatment for 21 days) on 13C-PBT was examined in rats. Body weight (BW), age and diagnostic status were significant predictors of the area under the curve of the time course of Δ13CO2 (‰) and the cumulative recovery rate (CRR) at 120 min. A repeated measures analysis of covariance controlled for age and BW revealed that the patterns of CRR change over time differed between the patients and controls and that Δ13CO2 was lower in the patients than in the controls at all sampling time points during the 120 min test, with an overall significant difference between the two groups. Chronic administration of RPD or HPD had no significant effect on 13C-PBT indices in rats. Our results suggest that 13C-PBT is a novel laboratory test that can detect altered phenylalanine kinetics in chronic schizophrenia patients. Animal experiments suggest that the observed changes are unlikely to be attributable to antipsychotic medication. PMID:22832963

  13. Detection of microvasculature alterations by synchrotron radiation in murine with delayed jellyfish envenomation syndrome.

    PubMed

    Wang, Beilei; Zhang, Bo; Huo, Hua; Wang, Tao; Wang, Qianqian; Wu, Yuanlin; Xiao, Liang; Ren, Yuqi; Zhang, Liming

    2014-04-01

    Using the tentacle extract (TE) from the jellyfish Cyanea capillata, we have previously established a delayed jellyfish envenomation syndrome (DJES) model, which is meaningful for clinical interventions against jellyfish stings. However, the mechanism of DJES still remains unclear. Thus, this study aimed to explore its potential mechanism by detecting TE-induced microvasculature alterations in vivo and ex vivo. Using a third-generation synchrotron radiation facility, we, for the first time, directly observed the blood vessel alterations induced by jellyfish venom in vivo and ex vivo. Firstly, microvasculature imaging of whole-body mouse in vivo indicated that the small blood vessel branches in the liver and kidney in the TE-treated group, seemed much thinner than those in the control group. Secondly, 3D imaging of kidney ex vivo showed that the kidneys in the TE-treated group had incomplete vascular trees where distal vessel branches were partly missing and disorderly disturbed. Finally, histopathological analysis found that obvious morphological changes, especially hemorrhagic effects, were also present in the TE-treated kidney. Thus, TE-induced microvasculature changes might be one of the important mechanisms of multiple organ dysfunctions in DJES. In addition, the methods we employed here will probably facilitate further studies on developing effective intervention strategies against DJES. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. First-Trimester Combined Screening Is Effective for the Detection of Unbalanced Chromosomal Translocations at 11 to 12 Weeks of Gestation

    PubMed Central

    Huang, ShangYu; Chang, ChiaLin; Hsiao, ChingHua; Soong, YungKuei; Duan, Tao

    2014-01-01

    The first trimester combined screening, which analyzes fetal nuchal translucency and levels of free β-human chorionic gonadotropin (β-hCG) and pregnancy-associated plasma protein A (PAPP-A) in maternal serum, is routinely used to detect abnormal pregnancies associated with Down syndrome and other trisomy aneuploidies. Based on the hypothesis that major chromosomal translocations could lead to similar biochemical and developmental outcomes during early embryo development, we compared these markers among pregnancies with normal, balanced, or unbalanced fetal karyotypes. Among the parents, 71 (73%) carry balanced reciprocal translocation and 26 (27%) have Robertsonian translocation. Of the 97 pregnancies tested, 39 (40%), 37 (37%), and 22 (23%) fetuses had normal karyotype, balanced chromosomal translocations, and unbalanced chromosomal translocations, respectively. Importantly, we found that pregnancies with an unbalanced translocation had significantly higher free β-hCG multiple of the median (MoM) and larger nuchal translucency thickness than those with normal karyotype or balanced translocations. Analysis showed that the area under a receiver operating characteristic curve (AUC) is 0.716, 0.820, and 0.936 for free β-hCG MoM, PAPP-A MoM, and fetal nuchal translucency, respectively. When these 3 independent factors were combined, the AUC reached 0.976. In addition, logistic regression showed that the most optimal model for predicting an unbalanced chromosomal translocation is a combination of PAPP-A and nuchal translucency with an AUC of 0.980. Therefore, the first trimester combined screening is not only effective in the screening of Down syndrome and other trisomy abnormalities but also has high sensitivity for the detection of unbalanced chromosomal translocations in fetuses. PMID:24177714

  15. Validation of targeted sequencing of single-nucleotide polymorphisms for non-invasive prenatal detection of aneuploidy of chromosomes 13, 18, 21, X, and Y.

    PubMed

    Nicolaides, K H; Syngelaki, A; Gil, M; Atanasova, V; Markova, D

    2013-06-01

    To assess the performance of cell-free DNA (cfDNA) testing in maternal blood for detection of fetal aneuploidy of chromosomes 13, 18, 21, X, and Y using targeted sequencing of single-nucleotide polymorphisms. Prospective study in 242 singleton pregnancies undergoing chorionic villus sampling at 11 to 13 weeks. Maternal blood was collected before chorionic villus sampling and sent to Natera (San Carlos, CA, USA). cfDNA was isolated from maternal plasma, and targeted multiplex PCR amplification followed by sequencing of 19 488 polymorphic loci covering chromosomes 13, 18, 21, X, and Y was performed. Sequencing data were analyzed using the NATUS algorithm that determines the copy number and calculates a sample-specific accuracy for each of the five chromosomes tested. Laboratory personnel were blinded to fetal karyotype. Results were provided for 229 (94.6%) of the 242 cases. Thirty-two cases were correctly identified as aneuploid, including trisomy 21 [n = 25; sensitivity = 100% (CI: 86.3-100%), specificity = 100% (CI: 98.2-100%)], trisomy 18 (n = 3), trisomy 13 (n = 1), Turner syndrome (n = 2), and triploidy (n = 1), with no false positive or false negative results. Median accuracy was 99.9% (range: 96.0-100%). cfDNA testing in maternal blood using targeted sequencing of polymorphic loci at chromosomes 13, 18, 21, X, and Y holds promise for accurate detection of fetal autosomal trisomies, sex chromosome aneuploidies, and triploidy. © 2013 John Wiley & Sons, Ltd.

  16. Structural chromosomal abnormalities in patients with mental retardation and/or multiple congenital anomalies: a new series of 24 patients.

    PubMed

    Tos, T; Karaman, A; Aksoy, A; Tukun, A

    2012-01-01

    Chromosomal abnormalities are a major cause of mental retardation and/or multiple congenital anomalies (MCA/MR). Screening for these chromosomal imbalances has mainly been done by standard karyotyping. The objective of this study was to report standard chromosome analysis and FISH screening of a series of 24 patients with MCA/MR. Structural chromosomal abnormalities were detected in 24 alterations and included 5 deletions, 2 duplications, 6 unbalanced translocations, 3 inversions, 2 insertions, 3 derivative chromosomes, 2 marker chromosomes and 1 isochromosome. We confirm that a high percentage of MCA/MR cases hitherto considered idiopathic is caused by chromosomal imbalances. We conclude that patients with MCA/MR should be routinely karyotyped.

  17. Rapid detection of sex chromosomal aneuploidies by QF-PCR: application in 200 men with severe oligozoospermia or azoospermia.

    PubMed

    Fodor, Flora; Kamory, Eniko; Csokay, Bela; Kopa, Zsolt; Kiss, Attila; Lantos, Istvan; Tisza, Timea

    2007-01-01

    Klinefelter syndrome is the most common genetic cause of severe male factor infertility. Cytogenetic evaluation of metaphase chromosomes generally has a long turnaround time. We describe a reliable molecular genetic method that can be completed in 2 working days to identify the presence of any extra X chromosomes. The quantitative fluorescent (QF) 5-plex PCR includes the amplification of amelogenin, which is present on both sex chromosomes in a biallelic form, a polymorphic short tandem repeat (STR) on the pseudoautosomal region of X and Y (X22), two polymorphic X-specific STRs (DXS6803, DXS6809), and a Y-specific marker (SY134), in a single tube. The presence of an extra X chromosome is recognized either by a supernumerary peak or an increased peak area based on criteria we have developed. The application of the method on 200 patients resulted in the identification of 14 patients (7%) with Klinefelter syndrome or a variant form (2 SRY-positive 46,XX men), as well as an additional patient with 47,XYY karyotype. The QF-PCR method, along with Y chromosome microdeletion testing, can be used as a first-step genetic analysis in azoospermic or severely oligozoospermic patients for the rapid identification of sex chromosome aneuploidies.

  18. Detection of Chromosomal Abnormalities with Different In Situ Hybridisation Techniques--the Usefulness in the Qualification of Cancer Patients for Molecularly-Targeted Therapies.

    PubMed

    Nicoś, Marcin; Wojas-Krawczyk, Kamila; Krawczyk, Paweł; Milanowski, Janusz

    2015-01-01

    Proper qualification of patients with cancer for an effective treatment regiment is essential to rationalize therapy benefit and costs. The early detection of genetic disorders that are responsible for the stimulation of uncontrolled cancer cells proliferation makes it possible to select a group of patients with a high probability of response to molecularly-targeted therapy. Data has shown that careful analysis of genes mutation using different PCR and sequencing techniques or chromosomal aberrations using in situ hybridization (ISH) techniques have a predictive value for drug targeted therapy. Overexpression of receptors and gene amplification has been reported in various cancers. Their detection is still a considerable challenge, which is connected with the unsatisfactory quality of DNA and low mutated cells percentage compared to cells with no genetic abnormalities in tested material. Different techniques of standardization were performed to prevent false negative results and to increase the sensitivity of qualitative and quantitative evaluation of chromosomal abnormalities. Immunohistochemistry (IHC) technique is useful in the screening of receptor expression in paraffin-embedded tissue samples in different malignant diseases. Whereas ISH techniques, especially fluorescence in situ hybridization (FISH), are now considered the diagnostic gold standard method in detection chromosomal aberrations. Moreover, molecular biology techniques, which are using molecular probes and real-time PCR and quantitative PCR techniques, were also applied for the detection of chromosomal changes. In order to identify the best genetic marker for treatment regiment, it is important to compare results of different studies, which are evaluating the sensitivity of diagnostic techniques and treatment response after a suitable selection factors based on genetic aberrations profile.

  19. Prenatal detection and outcome of congenital diaphragmatic hernia (CDH) associated with deletion of chromosome 15q26: two patients and review of the literature.

    PubMed

    Klaassens, M; Galjaard, R J H; Scott, D A; Brüggenwirth, H T; van Opstal, D; Fox, M V; Higgins, R R; Cohen-Overbeek, T E; Schoonderwaldt, E M; Lee, B; Tibboel, D; de Klein, A

    2007-09-15

    Congenital diaphragmatic hernia (CDH) is a severe birth defect characterized by a defect in the diaphragm with pulmonary hypoplasia and postnatal pulmonary hypertension. Approximately 50% of CDH cases are associated with other non-pulmonary congenital anomalies (so called non-isolated CDH) and in 5-10% of cases there is a chromosomal etiology. The majority of CDH cases are detected prenatally. In some cases prenatal chromosome analysis reveals a causative chromosomal anomaly, most often aneuploidy. Deletion of 15q26 is the most frequently described structural chromosomal aberration in patients with non-isolated CDH. In this paper we report on two patients with a deletion of 15q26 and phenotypes similar to other patients with CDH caused by 15q26 deletions. This phenotype consists of intra-uterine growth retardation, left-sided CDH, cardiac anomalies and characteristic facial features, similar to those seen in Fryns syndrome. We propose that when this combination of birth defects is identified, either pre- or postnatally, further investigations to confirm or exclude a deletion of 15q26 are indicated, since the diagnosis of this deletion will have major consequences for the prognosis and, therefore, can affect decision making.

  20. Detection of Phenotypic Alterations Using High-Content Analysis of Whole-Slide Images.

    PubMed

    Shirinifard, Abbas; Thiagarajan, Suresh; Vogel, Peter; Sablauer, András

    2016-05-01

    Tumors exhibit spatial heterogeneity, as manifested in immunohistochemistry (IHC) staining patterns. Current IHC quantification methods lose information by reducing this heterogeneity in each whole-slide image (WSI) or in selective fields of view to a single staining index. The aim of this study was to investigate the sensitivity of an IHC quantification method that uses this heterogeneity to reliably compare IHC staining patterns. We virtually partitioned WSIs by a grid of square tiles, and computed the staining index distributions to quantify heterogeneities. We used samples from these distributions as inputs to non-parametric statistical comparisons. We applied our grid method to fixed tumor samples from 26 tumors obtained from a double-blind preclinical study of a patient-derived orthotopic xenograft model of pediatric neuroblastoma in CD1 nude mice. We compared the results of our grid method to the results based on whole-slide indices, the current practice. We show that our grid method reliably detects phenotypic alterations that other tests based on whole-slide indices fail to detect. Based on robustness and increased sensitivity of statistical inference, we conclude that our method of whole-slide grid quantification is superior to existing whole-slide quantification techniques. © The Author(s) 2016.

  1. Maize (Zea mays) seeds can detect above-ground weeds; thiamethoxam alters the view.

    PubMed

    Afifi, Maha; Lee, Elizabeth; Lukens, Lewis; Swanton, Clarence

    2015-09-01

    Far red light is known to penetrate soil and delay seed germination. Thiamethoxam as a seed treatment has been observed to enhance seed germination. No previous work has explored the effect of thiamethoxam on the physiological response of buried maize seed when germinating in the presence of above-ground weeds. We hypothesised that the changes in red:far red reflected from above-ground weeds would be detected by maize seed phytochrome and delay seed germination by decreasing the level of GA and increasing ABA. We further hypothesised that thiamethoxam would overcome this delay in germination. Thiamethoxam enhanced seed germination in the presence of above-ground weeds by increasing GA signalling and downregulating DELLA protein and ABA signalling genes. An increase in amylase activity and a degradation of starch were also observed. Far red reflected from the above-ground weeds was capable of penetrating below the soil surface and was detected by maize seed phytochrome. Thiamethoxam altered the effect of far red on seed germination by stimulating GA and inhibiting ABA synthesis. This is the first study to suggest that the mode of action of thiamethoxam involves both GA synthesis and ABA inhibition. © 2014 Society of Chemical Industry.

  2. Detection of Genetically Altered Copper Levels in Drosophila Tissues by Synchrotron X-Ray Fluorescence Microscopy

    PubMed Central

    Lye, Jessica C.; Hwang, Joab E. C.; Paterson, David; de Jonge, Martin D.; Howard, Daryl L.; Burke, Richard

    2011-01-01

    Tissue-specific manipulation of known copper transport genes in Drosophila tissues results in phenotypes that are presumably due to an alteration in copper levels in the targeted cells. However direct confirmation of this has to date been technically challenging. Measures of cellular copper content such as expression levels of copper-responsive genes or cuproenzyme activity levels, while useful, are indirect. First-generation copper-sensitive fluorophores show promise but currently lack the sensitivity required to detect subtle changes in copper levels. Moreover such techniques do not provide information regarding other relevant biometals such as zinc or iron. Traditional techniques for measuring elemental composition such as inductively coupled plasma mass spectroscopy are not sensitive enough for use with the small tissue amounts available in Drosophila research. Here we present synchrotron x-ray fluorescence microscopy analysis of two different Drosophila tissues, the larval wing imaginal disc, and sectioned adult fly heads and show that this technique can be used to detect changes in tissue copper levels caused by targeted manipulation of known copper homeostasis genes. PMID:22053217

  3. Capturing Chromosome Conformation

    NASA Astrophysics Data System (ADS)

    Dekker, Job; Rippe, Karsten; Dekker, Martijn; Kleckner, Nancy

    2002-02-01

    We describe an approach to detect the frequency of interaction between any two genomic loci. Generation of a matrix of interaction frequencies between sites on the same or different chromosomes reveals their relative spatial disposition and provides information about the physical properties of the chromatin fiber. This methodology can be applied to the spatial organization of entire genomes in organisms from bacteria to human. Using the yeast Saccharomyces cerevisiae, we could confirm known qualitative features of chromosome organization within the nucleus and dynamic changes in that organization during meiosis. We also analyzed yeast chromosome III at the G1 stage of the cell cycle. We found that chromatin is highly flexible throughout. Furthermore, functionally distinct AT- and GC-rich domains were found to exhibit different conformations, and a population-average 3D model of chromosome III could be determined. Chromosome III emerges as a contorted ring.

  4. FISH-detected delay in replication timing of mutated FMR1 alleles on both active and inactive X-chromosomes.

    PubMed

    Yeshaya, J; Shalgi, R; Shohat, M; Avivi, L

    1999-01-01

    X-chromosome inactivation and the size of the CGG repeat number are assumed to play a role in the clinical, physical, and behavioral phenotype of female carriers of a mutated FMR1 allele. In view of the tight relationship between replication timing and the expression of a given DNA sequence, we have examined the replication timing of FMR1 alleles on active and inactive X-chromosomes in cell samples (lymphocytes or amniocytes) of 25 females: 17 heterozygous for a mutated FMR1 allele with a trinucleotide repeat number varying from 58 to a few hundred, and eight homozygous for a wild-type allele. We have applied two-color fluorescence in situ hybridization (FISH) with FMR1 and X-chromosome alpha-satellite probes to interphase cells of the various genotypes: the alpha-satellite probe was used to distinguish between early replicating (active) and late replicating (inactive) X-chromosomes, and the FMR1 probe revealed the replication pattern of this locus. All samples, except one with a large trinucleotide expansion, showed an early replicating FMR1 allele on the active X-chromosome and a late replicating allele on the inactive X-chromosome. In samples of mutation carriers, both the early and the late alleles showed delayed replication compared with normal alleles, regardless of repeat size. We conclude therefore that: (1) the FMR1 locus is subjected to X-inactivation; (2) mutated FMR1 alleles, regardless of repeat size, replicate later than wild-type alleles on both the active and inactive X-chromosomes; and (3) the delaying effect of the trinucleotide expansion, even with a low repeat size, is superimposed on the delay in replication associated with X-inactivation.

  5. Artifically inserting a reticuloendotheliosis virus long terminal repeat into a bacterial artificial chromosome clone of Marek's disease virus (MDV) alters expression of nearby MDV genes

    USDA-ARS?s Scientific Manuscript database

    The long terminal repeat (LTR) sequence of reticuloendotheliosis virus (REV) was inserted into the very virulent Marek’s disease virus (MDV) Md5 bacterial artificial chromosome clone. The insertion site was nearly identical to the REV LTR that was naturally inserted into the JM/102W strain of MDV fo...

  6. Molecular detection of altered X-inactivation patterns in the diagnosis of genetic disease.

    PubMed

    Malcolm, S

    1992-01-01

    It is widely assumed that when a female carrier of a genetic disorder exhibits clinical signs of the disorder it is due to chance non-random X-inactivation in particular tissues. Recently molecular methods have become available for the analysis of X-chromosome inactivation status. These are based either on the methylation patterns of DNA from the active and inactive chromosomes or on the rescue of active X chromosomes in somatic cell hybrids. As a consequence of the molecular studies, it has become obvious that there are some special cases of non-random X-inactivation patterns. These include females carrying X-linked immunodeficiencies and, sometimes, one of a pair of identical female twins.

  7. Microstructural white matter alterations in preclinical Alzheimer's disease detected using free water elimination diffusion tensor imaging.

    PubMed

    Hoy, Andrew R; Ly, Martina; Carlsson, Cynthia M; Okonkwo, Ozioma C; Zetterberg, Henrik; Blennow, Kaj; Sager, Mark A; Asthana, Sanjay; Johnson, Sterling C; Alexander, Andrew L; Bendlin, Barbara B

    2017-01-01

    Brain changes associated with Alzheimer's disease (AD) begin decades before disease diagnosis. While β-amyloid plaques and neurofibrillary tangles are defining features of AD, neuronal loss and synaptic pathology are closely related to the cognitive dysfunction. Brain imaging methods that are tuned to assess degeneration of myelinated nerve fibers in the brain (collectively called white matter) include diffusion tensor imaging (DTI) and related techniques, and are expected to shed light on disease-related loss of structural connectivity. Participants (N = 70, ages 47-76 years) from the Wisconsin Registry for Alzheimer's Prevention study underwent DTI and hybrid diffusion imaging to determine a free-water elimination (FWE-DTI) model. The study assessed the extent to which preclinical AD pathology affects brain white matter. Preclinical AD pathology was determined using cerebrospinal fluid (CSF) biomarkers. The sample was enriched for AD risk (APOE ε4 and parental history of AD). AD pathology assessed by CSF analyses was significantly associated with altered microstructure on both DTI and FWE-DTI. Affected regions included frontal, parietal, and especially temporal white matter. The f-value derived from the FWE-DTI model appeared to be the most sensitive to the relationship between the CSF AD biomarkers and microstructural alterations in white matter. These findings suggest that white matter degeneration is an early pathological feature of AD that may have utility both for early disease detection and as outcome measures for clinical trials. More complex models of microstructural diffusion properties including FWE-DTI may provide increased sensitivity to early brain changes associated with AD over standard DTI.

  8. Single-Gene Detection and Karyotyping Using Small-Target Fluorescence in Situ Hybridization on Maize Somatic Chromosomes

    PubMed Central

    Lamb, Jonathan C.; Danilova, Tatiana; Bauer, Matthew J.; Meyer, Julie M.; Holland, Jennifer J.; Jensen, Michael D.; Birchler, James A.

    2007-01-01

    Combined with a system for identifying each of the chromosomes in a genome, visualizing the location of individual genetic loci by fluorescence in situ hybridization (FISH) would aid in assembling physical and genetic maps. Previously, large genomic clones have been successfully used as FISH probes onto somatic chromosomes but this approach is complicated in species with abundant repetitive elements. In this study, repeat-free portions of sequences that were anchored to particular chromosomes including genes, gene clusters, large cDNAs, and portions of BACs obtained from public databases were used to label the corresponding physical location using FISH. A collection of probes that includes at least one marker on each chromosome in the maize complement was assembled, allowing a small-target karyotyping system to be developed. This set provides the foundation onto which additional loci could be added to strengthen further the ability to perform chromosomal identification in maize and its relatives. The probes were demonstrated to produce signals in several wild relatives of maize, including Zea luxurians, Z. diploperennis, and Tripsacum dactyloides. PMID:17237520

  9. Microdissected double-minute DNA detects variable patterns of chromosomal localizations and multiple abundantly expressed transcripts in normal and leukemic cells

    SciTech Connect

    Sen, S.; Zhou, Hongyi; Stass, S.A.; Sen, P. ); Mulac-Jericevic, B.; Pirrotta, V. )

    1994-02-01

    Double-minute (dm) chromosomes are cytogenetically resolvable DNA amplification-mediating acentric extrachromosomal structures that are commonly seen in primary tumors, tumor cell lines, and drug-resistant cells grown in vitro. Selective isolation of dm DNAs with standard molecular biological techniques is difficult, and thus, detailed studies to elucidate their structure, site of chromosomal origin, and chromosomal reintegration patterns have been limited. In those instances in which a gene has been localized on dms, characterization of the remainder of the DNA, which far exceeds the size of the gene identified, has remained inconclusive. dms seen in the acute myeloid leukemia cell line HL-60 have been shown to harbor the c-myc protooncogene. In this paper, the authors report the successful isolation of the dm-specific DNAs from these cells by the microdissection/polymerase chain reaction technique and demonstrate that the dm DNAs derived from a single discrete normal chromosome segment 8q24.1-q24.2 reintegrate at various specific locations in the leukemic cells. The microdissected dm DNA detects multiple abundantly expressed transcripts distinct from c-myc mRNA on Northern blots. By devising a [open quotes]transcript selection[close quotes] strategy, they cloned the partial genomic sequence of a gene from the microdissected DNA that encodes two of these RNAs. This strategy will be generally applicable for rapid cloning of unknown amplified genes harbored on dms. With DNA from 20 microdissected dms, they constructed a genomic library of about 20,000 recombinant microclones with an average insert size of about 450 bp. The microclones should help in isolating corresponding yeast artificial chromosome clones for high-resolution physical mapping of dms in HL-60 cells. Furthermore, application of the microdissection technique appears to be an extremely feasible approach to characterization of dms in other cell types. 42 refs., 6 figs., 1 tab.

  10. [Value of detection of cell-free fetal DNA in maternal plasma in the prenatal diagnosis of chromosomal abnormalities].

    PubMed

    Wang, Shu-juan; Gao, Zhi-ying; Lu, Yan-ping; Li, Ya-li; You, Yan-qin; Zhang, Li-wen; Wang, Long-xia; Xu, Hong

    2012-11-01

    To investigate the value of detection of fetal cell-free fetal DNA (cff-DNA) in maternal plasma in the prenatal diagnosis of chromosomal abnormalities. The plasma from 3200 gravidas (singleton with 20.3 ± 3.8 gestational weeks) was collected from April 1(st) 2011 to May 30(th) 2012. They were divided into 3 groups: (1) To tally 1720 cases were included in the high-risk serological screening group, in which women were younger than 35 years and got high-risk results in serological screening; (2) To tally 1310 cases were included in the advanced age group, in which women's age was more than 35 years; (3) To tally 170 cases were included in the supplementary group, in which women were younger than 35 years and got low-risk results in serological screening, or women who didn't take serological screening tests. All the 3030 gravidas in group 1 and 2 didn't take invasive prenatal diagnosis because of fear of abortion or short of prenatal diagnosis. Cff-DNA were detected by next generation sequencing in Shenzhen BGI Genomics Center for clinical laboratory. Amniocentesis and karyotype analysis were provided to the positive cases and women with negative results were followed-up by telephone. (1) The 3200 cases took cff-DNA detection, and 31 cases got positive results, including 27 cases of trisomy 21 and 4 cases of trisomy 18. Sixteen cases of trisomy 21 and 1 case of trisomy 18 were in the high-risk serological screening group. 7 cases of trisomy 21 and 2 cases of trisomy 18 were in the advanced age group. Four cases of trisomy 21 and 1 case of trisomy 18 were in the supplementary group. (2) And the 84% (26/31) cff-DNA detecting positive cases received amniocentesis. In the 27 trisomy 21 positive cases, 23 received amniocentesis and got karyotype of 47XN, +21, with the diagnostic accordance rate of 100%. In the 4 cases who didn't take karyotype analysis, fetal anomaly (ventricular septal defect, dextrocardia and choroid plexus cyst) was found in 1 case before 20

  11. Chromosomal Abnormalities in Non-Small Cell Lung Carcinomas and in Bronchial Epithelia of High-Risk Smokers Detected by Multi-Target Interphase Fluorescence in Situ Hybridization

    PubMed Central

    Santos Romeo, Maura; Sokolova, Irina A.; Morrison, Larry E.; Zeng, Chan; Barón, Anna E.; Hirsch, Fred R.; Miller, York E.; Franklin, Wilbur A.; Varella-Garcia, Marileila

    2003-01-01

    Human lung carcinogenesis is accompanied by complex chromosomal changes that may be detected in interphase cells by fluorescence in situ hybridization (FISH) assay using recently developed multitarget DNA probes. Touch preparations of 20 non-small cell lung carcinomas, sputum specimens from 3 patients with lung cancer and from 11 ex-smokers without lung cancer, and cultured benign bronchial epithelium of 42 high-risk smokers, 9 of whom had concurrent invasive carcinoma, were tested using a four-color FISH probe (LAVysion) targeting centromere 6, 5p15.2, 7p12 (EGFR), and 8q24 (MYC). Significantly high frequencies of abnormal cells were found in each of the 20 NSCLC (100%) and in the 3 sputum specimens from lung cancer patients. None of the cytologically normal sputa contained FISH abnormalities. Cultured bronchial epithelial cells from 11 of 42 patients (26%) were abnormal for at least one probe. Abnormal FISH patterns had no association with gender, presence of tumor or histology. Multicolor FISH can readily detect chromosomal abnormalities in imprints and sputa from lung carcinomas. Chromosomal aneusomy is also frequent in bronchial epithelial cells from long-term smokers. The prognostic significance of the multicolor LAVysion FISH probe set should be validated in a controlled clinical trial. PMID:12707375

  12. Chromosomal abnormalities in non-small cell lung carcinomas and in bronchial epithelia of high-risk smokers detected by multi-target interphase fluorescence in situ hybridization.

    PubMed

    Romeo, Maura Santos; Sokolova, Irina A; Morrison, Larry E; Zeng, Chan; Barón, Anna E; Hirsch, Fred R; Miller, York E; Franklin, Wilbur A; Varella-Garcia, Marileila

    2003-05-01

    Human lung carcinogenesis is accompanied by complex chromosomal changes that may be detected in interphase cells by fluorescence in situ hybridization (FISH) assay using recently developed multitarget DNA probes. Touch preparations of 20 non-small cell lung carcinomas, sputum specimens from 3 patients with lung cancer and from 11 ex-smokers without lung cancer, and cultured benign bronchial epithelium of 42 high-risk smokers, 9 of whom had concurrent invasive carcinoma, were tested using a four-color FISH probe (LAVysion) targeting centromere 6, 5p15.2, 7p12 (EGFR), and 8q24 (MYC). Significantly high frequencies of abnormal cells were found in each of the 20 NSCLC (100%) and in the 3 sputum specimens from lung cancer patients. None of the cytologically normal sputa contained FISH abnormalities. Cultured bronchial epithelial cells from 11 of 42 patients (26%) were abnormal for at least one probe. Abnormal FISH patterns had no association with gender, presence of tumor or histology. Multicolor FISH can readily detect chromosomal abnormalities in imprints and sputa from lung carcinomas. Chromosomal aneusomy is also frequent in bronchial epithelial cells from long-term smokers. The prognostic significance of the multicolor LAVysion FISH probe set should be validated in a controlled clinical trial.

  13. High-resolution analysis of DNA copy number alterations in patients with isolated sporadic keratoconus

    PubMed Central

    Hellani, Ali M.; Al Mansouri, Sameer M.; Kalantan, Hatem; Al-Muammar, Abdulrahman M.

    2011-01-01

    Purpose To determine whether patients with sporadic, non-familial keratoconus and no pathogenic mutations in the visual system homeobox 1 (VSX1) gene have evidence of chromosomal copy number alterations. Methods Twenty Saudi Arabian patients with isolated keratoconus, no family history of the disease and no mutations in VSX1 were recruited. Additionally, 10 ethnically-matched healthy controls were also recruited for this study. We screened patients for chromosomal copy number aberrations using the Agilent Human Genome CGH 244A Oligo Microarray Chip. Results None of the keratoconus patients screened had evidence of chromosomal copy number alterations when compared to normal ethnically matched controls. Conclusions Chromosomal deletions and/or duplications were not detected in any of the patients tested here. Other chromosomal imbalances such as translocations, inversions, and some ploidies cannot be detected by current array CGH technology and other nuclear genetic or epigenetic factors cannot be excluded as a possible contributing factor to keratoconus pathogenesis. PMID:21528002

  14. High-resolution analysis of DNA copy number alterations in patients with isolated sporadic keratoconus.

    PubMed

    Abu-Amero, Khaled K; Hellani, Ali M; Al Mansouri, Sameer M; Kalantan, Hatem; Al-Muammar, Abdulrahman M

    2011-03-30

    To determine whether patients with sporadic, non-familial keratoconus and no pathogenic mutations in the visual system homeobox 1 (VSX1) gene have evidence of chromosomal copy number alterations. Twenty Saudi Arabian patients with isolated keratoconus, no family history of the disease and no mutations in VSX1 were recruited. Additionally, 10 ethnically-matched healthy controls were also recruited for this study. We screened patients for chromosomal copy number aberrations using the Agilent Human Genome CGH 244A Oligo Microarray Chip. None of the keratoconus patients screened had evidence of chromosomal copy number alterations when compared to normal ethnically matched controls. Chromosomal deletions and/or duplications were not detected in any of the patients tested here. Other chromosomal imbalances such as translocations, inversions, and some ploidies cannot be detected by current array CGH technology and other nuclear genetic or epigenetic factors cannot be excluded as a possible contributing factor to keratoconus pathogenesis.

  15. CloneCNA: detecting subclonal somatic copy number alterations in heterogeneous tumor samples from whole-exome sequencing data.

    PubMed

    Yu, Zhenhua; Li, Ao; Wang, Minghui

    2016-08-19

    Copy number alteration is a main genetic structural variation that plays an important role in tumor initialization and progression. Accurate detection of copy number alterations is necessary for discovering cancer-causing genes. Whole-exome sequencing has become a widely used technology in the last decade for detecting various types of genomic aberrations in cancer genomes. However, there are several major issues encountered in these detection problems, including normal cell contamination, tumor aneuploidy, and intra-tumor heterogeneity. Especially, deciphering the intra-tumor heterogeneity is imperative for identifying clonal and subclonal copy number alterations. We introduce CloneCNA, a novel bioinformatics tool for efficiently addressing these issues and automatically detecting clonal and subclonal somatic copy number alterations from heterogeneous tumor samples. CloneCNA fully explores the log ratio of read counts between paired tumor-normal samples and tumor B allele frequency of germline heterozygous SNP positions, further employs efficient statistical models to quantitatively represent copy number status of tumor sample containing multiple clones. We examine CloneCNA on simulated heterogeneous and real tumor samples, and the results demonstrate that CloneCNA has higher power to detect copy number alterations than existing methods. CloneCNA, a novel algorithm is developed to efficiently and accurately identify somatic copy number alterations from heterogeneous tumor samples. We demonstrate the statistical framework of CloneCNA represents a remarkable advance for tumor whole-exome sequencing data. We expect that CloneCNA will promote cancer-focused studies for investigating the role of clonal evolution and elucidating critical events benefiting tumor tumourigenesis and progression.

  16. Detection of aneuploidy in sperm of an ataxia telangiectasia patient using three-chromosome fluorescence in situ hybridization

    SciTech Connect

    Lowe, X.R.; Baulch, J.E.; Arnheim, N.

    1994-09-01

    Ataxia telangiectasia (A-T) is an inherited, recessive, cancer-prone disorder. Fluorescence in situ hybridization (FISH) with DNA probes specific for three chromosomes was applied to sperm of an A-T patient to determine if there may be an increased germinal risk for aneuploidy. Air-dried sperm smears were treated with proteinase K and were decondensed with DTT and LIS. The slides were then hybridized with fluorescently labeled repetitive DNA probes specific for chromosomes X, Y and 8, and a total of 11,825 sperm cells were scored. The ratio of sperm bearing X-8 and Y-8 was 1:1, as predicted. The frequencies of hyperhaploidy were 3.9, 1.0, 17.6 and 7.8 per 10,000 cells for categories X-X-8, Y-Y-8, X-Y-8 and 8-8-(X or Y), respectively, In addition, the frequency of diploidy (X-Y-8-8) was 18.6 and auto-diploidies (X-X-8-8 and Y-Y-8-8) were 1.0 and 2.0, respectively. These frequencies were not significantly different when compared with levels in healthy men (p > 0.1). Our finding suggests that chromosome X, Y and 8 aneuploidies are not elevated in the sperm of A-T patients, but studies with additional patients and chromosomes are needed.

  17. Alterations in candidate genes PHF2, FANCC, PTCH1 and XPA at chromosomal 9q22.3 region: Pathological significance in early- and late-onset breast carcinoma

    PubMed Central

    Sinha, Satyabrata; Singh, Ratnesh K; Alam, Neyaz; Roy, Anup; Roychoudhury, Susanta; Panda, Chinmay Kumar

    2008-01-01

    Introduction Younger women with breast carcinoma (BC) exhibits more aggressive pathologic features compared to older women; young age could be an independent predictor of adverse prognosis. To find any existing differences in the molecular pathogenesis of BC in both younger and older women, alterations at chromosomal (chr.) 9q22.32-22.33 region were studied owing to its association in wide variety of tumors. Present work focuses on comparative analysis of alterations of four candidate genes; PHF2, FANCC, PTCH1 and XPA located within 4.4 Mb region of the afore-said locus in two age groups of BC, as well as the interrelation and prognostic significance of alterations of these genes. Methods Deletion analysis of PHF2, FANCC, PTCH1 and XPA were examined in a subset of 47 early-onset (group-A: ≤ 40 years) and 59 late-onset (group-B: > 40 years) breast carcinomas using both microsatellite and exonic markers. Methylation Sensitive Restriction analysis (MSRA) was done to check for promoter methylation. Quantitative real-time polymerase chain reaction (Q-PCR) and immunohistochemisty (IHC) was done in some genes to see their relative mRNA and protein expressions respectively. Clinico-pathological correlation of different parameters as well as patient survival was calculated using different statistical softwares like EpiInfo 6.04b, SPSS 10.0 etc. Results Either age group exhibited high frequency of overall alterations in PHF2, FANCC and PTCH1 compared to XPA. Samples with alteration (deletion/methylation) in these genes showed reduced level of mRNA expression as seen by Q-PCR. Immunohistochemical analysis of FANCC and PTCH1 also supported this observation. Poor patient survival was noted in both age groups having alterations in FANCC. Similar result was also seen with PTCH1 and XPA alterations in group-A and PHF2 alterations in group-B. This reflected their roles as prognostic tools in the respective groups in which they were altered. Conclusion Overall alterations of PHF2

  18. Allometric Analysis Detects Brain Size-Independent Effects of Sex and Sex Chromosome Complement on Human Cerebellar Organization.

    PubMed

    Mankiw, Catherine; Park, Min Tae M; Reardon, P K; Fish, Ari M; Clasen, Liv S; Greenstein, Deanna; Giedd, Jay N; Blumenthal, Jonathan D; Lerch, Jason P; Chakravarty, M Mallar; Raznahan, Armin

    2017-03-17

    The cerebellum is a large hindbrain structure that is increasingly recognized for its contribution to diverse domains of cognitive and affective processing in human health and disease. Although several of these domains are sex-biased, our fundamental understanding of cerebellar sex differences - including their spatial distribution, potential biological determinants, and independence from brain volume variation - lags far behind that for the cerebrum. Here, we harness automated neuroimaging methods for cerebellar morphometrics in 417 individuals to (i) localize normative male-female differences in raw cerebellar volume, (ii) compare these to sex chromosome effects estimated across five rare X-/Y-chromosome aneuploidy (SCA) syndromes, and (iii) clarify brain size-independent effects of sex and SCA on cerebellar anatomy using a generalizable allometric approach which considers scaling relationships between regional cerebellar volume and brain volume in health. Integration of these approaches shows that (i) sex and SCA effects on raw cerebellar volume are large and distributed, but regionally heterogeneous, (ii) human cerebellar volume scales with brain volume in a highly non-linear and regionally heterogeneous fashion that departs from documented patterns of cerebellar scaling in phylogeny, and (iii) cerebellar organization is modified in a brain size-independent manner by sex (relative expansion of total cerebellum, flocculus, and Crus II-lobule VIIIB volumes in males) and SCA (contraction of total cerebellar, lobule IV and Crus I volumes with additional X- or Y-chromosomes; X-specific contraction of Crus II-lobule VIIIB). Our methods and results clarify the shifts in human cerebellar organization that accompany interwoven variations in sex, sex chromosome complement, and brain size.SIGNIFICANCE STATEMENTCerebellar systems are implicated in diverse domains of sex-biased behavior and pathology, but we lack a basic understanding of how sex differences in the human

  19. Alterations of ATM and CADM1 in chromosomal 11q22.3-23.2 region are associated with the development of invasive cervical carcinoma.

    PubMed

    Mazumder Indra, Dipanjana; Mitra, Sraboni; Roy, Anup; Mondal, Ranajit Kumar; Basu, Partha Sarathi; Roychoudhury, Susanta; Chakravarty, Runu; Panda, Chinmay Kumar

    2011-12-01

    To understand the importance of chr11q22.3-23.2 region in the development of cervical cancer, we have studied the genetic and epigenetic alterations of the candidate genes ATM, PPP2R1B, SDHD and CADM1 in cervical intraepithelial neoplasia (CIN) and cervical carcinoma (CACX) samples. Our study revealed low expression and high alterations (methylation/deletion) (55-59%) of ATM and CADM1 genes along with poor patient outcome. The alterations of ATM and CADM1 are associated with the progression of tumor from CIN to Stage I/II, thus implying their role in early invasiveness. The two genes, PPP2R1B and SDHD, lying in between ATM and CADM1, have low frequency of alterations, and majority of the alterations are in CACX samples, indicating that their alterations might be associated with disease progression. Expressions (mRNA/protein) of the genes showed concordance with their molecular alterations. Significant co-alteration of ATM and CADM1 points to their synergic action for the development of CACX. Mutation is, however, a rare phenomenon for inactivation of ATM. Association between the alteration of ATM and CHEK1 and poor survival of the patients having co-alterations of ATM and CHEK1 points to the DNA damage response pathway disruption in development of CACX. Thus, our data suggest that inactivation of ATM-CHEK1-associated DNA damage response pathway and CADM1-associated signaling network might have an important role in the development of CACX.

  20. Human Spermatozoa as a Model for Detecting Missing Proteins in the Context of the Chromosome-Centric Human Proteome Project.

    PubMed

    Jumeau, Fanny; Com, Emmanuelle; Lane, Lydie; Duek, Paula; Lagarrigue, Mélanie; Lavigne, Régis; Guillot, Laëtitia; Rondel, Karine; Gateau, Alain; Melaine, Nathalie; Guével, Blandine; Sergeant, Nicolas; Mitchell, Valérie; Pineau, Charles

    2015-09-04

    The Chromosome-Centric Human Proteome Project (C-HPP) aims at cataloguing the proteins as gene products encoded by the human genome in a chromosome-centric manner. The existence of products of about 82% of the genes has been confirmed at the protein level. However, the number of so-called "missing proteins" remains significant. It was recently suggested that the expression of proteins that have been systematically missed might be restricted to particular organs or cell types, for example, the testis. Testicular function, and spermatogenesis in particular, is conditioned by the successive activation or repression of thousands of genes and proteins including numerous germ cell- and testis-specific products. Both the testis and postmeiotic germ cells are thus promising sites at which to search for missing proteins, and ejaculated spermatozoa are a potential source of proteins whose expression is restricted to the germ cell lineage. A trans-chromosome-based data analysis was performed to catalog missing proteins in total protein extracts from isolated human spermatozoa. We have identified and manually validated peptide matches to 89 missing proteins in human spermatozoa. In addition, we carefully validated three proteins that were scored as uncertain in the latest neXtProt release (09.19.2014). A focus was then given to the 12 missing proteins encoded on chromosomes 2 and 14, some of which may putatively play roles in ciliation and flagellum mechanistics. The expression pattern of C2orf57 and TEX37 was confirmed in the adult testis by immunohistochemistry. On the basis of transcript expression during human spermatogenesis, we further consider the potential for discovering additional missing proteins in the testicular postmeiotic germ cell lineage and in ejaculated spermatozoa. This project was conducted as part of the C-HPP initiatives on chromosomes 14 (France) and 2 (Switzerland). The mass spectrometry proteomics data have been deposited with the Proteome

  1. Detection of hydrothermal alteration at Virginia City, Nevada using Airborne Imaging Spectrometry (AIS)

    NASA Technical Reports Server (NTRS)

    Hutsinpiller, A.; Taranik, J. V.

    1986-01-01

    Airborne Imaging Spectrometer (AIS) data were collected over Virginia City, Nevada; an area of gold and silver mineralization with extensive surface exposures of altered volcanic rocks. The data were corrected for atmospheric effects by a flat-field method, and compared to library spectra of various alteration minerals using a spectral analysis program SPAM. Areas of strong clay alteration were identified on the AIS images that were mapped as kaolinitic, illitic, and sericitic alterations zones. Kaolinitic alteration is distinguishable in the 2.1 to 2.4 and 1.2 to 1.5 micrometer wavelength regions. Montmorillonite, illite, and sericite have absorption features similar to each other at 2.2 micrometer wavelength. Montnorillonite and illite also may be present in varying proportions within one Ground Instantaneous Field of View (GIFOV). In general AIS data is useful in identifying alteration zones that are associated with or lie above precious metal mineralization at Virginia City.

  2. Detection of hydrothermal alteration at Virginia City, Nevada using Airborne Imaging Spectrometry (AIS)

    NASA Technical Reports Server (NTRS)

    Hutsinpiller, A.; Taranik, J. V.

    1986-01-01

    Airborne Imaging Spectrometer (AIS) data were collected over Virginia City, Nevada; an area of gold and silver mineralization with extensive surface exposures of altered volcanic rocks. The data were corrected for atmospheric effects by a flat-field method, and compared to library spectra of various alteration minerals using a spectral analysis program SPAM. Areas of strong clay alteration were identified on the AIS images that were mapped as kaolinitic, illitic, and sericitic alterations zones. Kaolinitic alteration is distinguishable in the 2.1 to 2.4 and 1.2 to 1.5 micrometer wavelength regions. Montmorillonite, illite, and sericite have absorption features similar to each other at 2.2 micrometer wavelength. Montnorillonite and illite also may be present in varying proportions within one Ground Instantaneous Field of View (GIFOV). In general AIS data is useful in identifying alteration zones that are associated with or lie above precious metal mineralization at Virginia City.

  3. Epilepsy and chromosomal abnormalities

    PubMed Central

    2010-01-01

    Background Many chromosomal abnormalities are associated with Central Nervous System (CNS) malformations and other neurological alterations, among which seizures and epilepsy. Some of these show a peculiar epileptic and EEG pattern. We describe some epileptic syndromes frequently reported in chromosomal disorders. Methods Detailed clinical assessment, electrophysiological studies, survey of the literature. Results In some of these congenital syndromes the clinical presentation and EEG anomalies seems to be quite typical, in others the manifestations appear aspecific and no strictly linked with the chromosomal imbalance. The onset of seizures is often during the neonatal period of the infancy. Conclusions A better characterization of the electro clinical patterns associated with specific chromosomal aberrations could give us a valuable key in the identification of epilepsy susceptibility of some chromosomal loci, using the new advances in molecular cytogenetics techniques - such as fluorescent in situ hybridization (FISH), subtelomeric analysis and CGH (comparative genomic hybridization) microarray. However further studies are needed to understand the mechanism of epilepsy associated with chromosomal abnormalities. PMID:20438626

  4. Richter's syndrome with identification of marker chromosomes.

    PubMed

    Fitzgerald, P H; McEwan, C M; Hamer, J W; Beard, M E

    1980-07-01

    A case is presented of a man with Richter's syndrome with diffuse histiocytic lymphoma following a ten-year history of untreated chronic lymphatic leukemia. He did not respond to therapy. The lymphoma cells had 61 chromosomes with aneuploidy of 13 chromosomes and 11 structurally altered chromosomes. Only chromosomes 8, 9, 12, 14, and 18 were diploid and without abnormality. Significantly, part of chromsome 1q was duplicated as in the partial trisomy of 1q reported to characterize a number of hematologic neoplasms. A large marker chromosome with subterminal centromere was a tandem duplication of chromosome 4q.

  5. STUDIES ON THE ISOLATION OF METAPHASE CHROMOSOMES

    PubMed Central

    ChoraŻy, M.; Bendich, A.; Borenfreund, E.; Hutchison, D. J.

    1963-01-01

    A method for the isolation of metaphase chromosomes from mouse L1210 leukemia cells has been developed. Cells, arrested at metaphase with colchicine, were exposed to hypotonic solution and the pH was then adjusted to 5.6 to stabilize the chromosomes. The metaphase figures were subsequently disrupted and the chromosomes isolated by a series of differential centrifugations in sucrose. The isolated chromosomes were well preserved, as judged by morphological criteria. The effect of various enzymes and chemical agents on the isolated chromosomes was studied. Chymotrypsin, trypsin, and deoxyribonuclease caused a marked disintegration of the chromosomes, whereas treatment with pepsin and ribonuclease induced no significant morphological alterations. PMID:14069802

  6. Detection of alterations in testicular and epididymal function in laboratory animals.

    PubMed Central

    Amann, R P

    1986-01-01

    The potential impact of an agent altering male reproductive function is greater for humans than for animals. Consequently, it is essential that sensitive criteria be used to look for effects on a multiplicity of target sites when an agent is evaluated using an animal model. No animal model has reproductive characteristics similar to those of humans, but this does not negate the validity of using animal models. Classic methodologies for reproductive toxicology are limited by the approaches used for subjective evaluation of testicular histology and use of natural mating for fertility tests. After dosing for an interval at least equal to six times the duration of one cycle of the seminiferous epithelium, sperm from ejaculated semen or the cauda epididymidis can be evaluated for normalcy of morphology or function and should be used for artificial insemination of females to critically evaluate fertility. Normal males of animal models ejaculate a great excess of sperm. A 50 or 90% reduction in the number of fertile sperm deposited during mating probably will not markedly reduce fertility. Artificial insemination of a critical number of sperm, selected to result in slightly less than maximal fertility for control animals, will maximize the probability of detecting a decrease in fertility if the same critical number of sperm is inseminated for treated animals as for control animals. Testicular function should be evaluated by objective, rather than subjective, criteria. For each male, a piece of testicular tissue should be appropriately fixed and an aliquot of parenchyma should be homogenized to allow enumeration of homogenization-resistant spermatids. Among the more sensitive criteria of testicular function are the minor diameter of essentially round seminiferous tubules, the ratio of leptotene spermatocytes to Sertoli cells, the corrected numbers of germ cells per seminiferous tubule cross section, and the number of homogenization-resistant spermatids per testis. PMID

  7. Thermal nociceptive threshold testing detects altered sensory processing in broiler chickens with spontaneous lameness.

    PubMed

    Hothersall, Becky; Caplen, Gina; Parker, Richard M A; Nicol, Christine J; Waterman-Pearson, Avril E; Weeks, Claire A; Murrell, Joanna C

    2014-01-01

    Lameness is common in commercially reared broiler chickens but relationships between lameness and pain (and thus bird welfare) have proved complex, partly because lameness is often partially confounded with factors such as bodyweight, sex and pathology. Thermal nociceptive threshold (TNT) testing explores the neural processing of noxious stimuli, and so can contribute to our understanding of pain. Using an acute model of experimentally induced articular pain, we recently demonstrated that TNT was reduced in lame broiler chickens, and was subsequently attenuated by administration of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs). This study extended these findings to a large sample of commercial broilers. It examined factors affecting thermal threshold (Part 1) and the effect of an NSAID drug (meloxicam, 5 mg/kg) and of an opioid (butorphanol; 4 mg/kg) (Part 2). Spontaneously lame and matched non-lame birds (n=167) from commercial farms were exposed to ramped thermal stimulations via a probe attached to the lateral aspect of the tarsometatarsus. Baseline skin temperature and temperature at which a behavioural avoidance response occurred (threshold) were recorded. In Part 1 bird characteristics influencing threshold were modelled; In Part 2 the effect of subcutaneous administration of meloxicam or butorphanol was investigated. Unexpectedly, after accounting for other influences, lameness increased threshold significantly (Part 1). In Part 2, meloxicam affected threshold differentially: it increased further in lame birds and decreased in non-lame birds. No effect of butorphanol was detected. Baseline skin temperature was also consistently a significant predictor of threshold. Overall, lameness significantly influenced threshold after other bird characteristics were taken into account. This, and a differential effect of meloxicam on lame birds, suggests that nociceptive processing may be altered in lame birds, though mechanisms for this require further investigation.

  8. Detecting truly clonal alterations from multi-region profiling of tumours

    NASA Astrophysics Data System (ADS)

    Werner, Benjamin; Traulsen, Arne; Sottoriva, Andrea; Dingli, David

    2017-03-01

    Modern cancer therapies aim at targeting tumour-specific alterations, such as mutations or neo-antigens, and maximal treatment efficacy requires that targeted alterations are present in all tumour cells. Currently, treatment decisions are based on one or a few samples per tumour, creating uncertainty on whether alterations found in those samples are actually present in all tumour cells. The probability of classifying clonal versus sub-clonal alterations from multi-region profiling of tumours depends on the earliest phylogenetic branching event during tumour growth. By analysing 181 samples from 10 renal carcinoma and 11 colorectal cancers we demonstrate that the information gain from additional sampling falls onto a simple universal curve. We found that in colorectal cancers, 30% of alterations identified as clonal with one biopsy proved sub-clonal when 8 samples were considered. The probability to overestimate clonal alterations fell below 1% in 7/11 patients with 8 samples per tumour. In renal cell carcinoma, 8 samples reduced the list of clonal alterations by 40% with respect to a single biopsy. The probability to overestimate clonal alterations remained as high as 92% in 7/10 renal cancer patients. Furthermore, treatment was associated with more unbalanced tumour phylogenetic trees, suggesting the need of denser sampling of tumours at relapse.

  9. Detecting truly clonal alterations from multi-region profiling of tumours

    PubMed Central

    Werner, Benjamin; Traulsen, Arne; Sottoriva, Andrea; Dingli, David

    2017-01-01

    Modern cancer therapies aim at targeting tumour-specific alterations, such as mutations or neo-antigens, and maximal treatment efficacy requires that targeted alterations are present in all tumour cells. Currently, treatment decisions are based on one or a few samples per tumour, creating uncertainty on whether alterations found in those samples are actually present in all tumour cells. The probability of classifying clonal versus sub-clonal alterations from multi-region profiling of tumours depends on the earliest phylogenetic branching event during tumour growth. By analysing 181 samples from 10 renal carcinoma and 11 colorectal cancers we demonstrate that the information gain from additional sampling falls onto a simple universal curve. We found that in colorectal cancers, 30% of alterations identified as clonal with one biopsy proved sub-clonal when 8 samples were considered. The probability to overestimate clonal alterations fell below 1% in 7/11 patients with 8 samples per tumour. In renal cell carcinoma, 8 samples reduced the list of clonal alterations by 40% with respect to a single biopsy. The probability to overestimate clonal alterations remained as high as 92% in 7/10 renal cancer patients. Furthermore, treatment was associated with more unbalanced tumour phylogenetic trees, suggesting the need of denser sampling of tumours at relapse. PMID:28344344

  10. International, collaborative assessment of limitations of chromosome-specific probes (CSP) and fluorescent in situ hybridization (FISH): Analysis of expected detections in 73,000 prenatal cases

    SciTech Connect

    Evans, M.I.; Henry, G.P.; Miller, W.A.

    1994-09-01

    FISH and CSP have been proposed to reduce karyotyping need. The purpose of this study was to assess the potential efficacy of CSP-FISH using currently available probes (13, 18, 21, X, & Y) in large, prenatal diagnostic centers. Results (1990-1993) from 7 centers in 4 countries were divided by those expected to be detectable by currently available probes, and those which would be missed assuming 10% probe efficacy. 72,994 karyotypes included 699 trisomy 21`s, 352 trisomy 18`s, 136 trisomy 13`s, 358 sex chromosome aneuploidies, 70 triploidies, and 855 others (translocations, inversions, deletions, markers). Of 2,613 abnormalities, 1,745 would be detectable (66.8%). [Detroit 55.7%, Stockholm 68.3%, Boston 52.6%, Denver 61.3%, Muenster 77.0%, London 84.5%, Philadelphia 69.4%]. Centers with high proportions of referrals for ultrasound anomalies had the highest CSP-FISH positives secondary to increased T 18 & 13. We conclude: (1) 73,000 karyotypes show relatively consistent incidences of the common trisomies, sex chromosome abnormalities, and other chromosome abnormalities among the centers. (2) The proportion expected detectable by FISH-CSP technology varies from 52.6% to 84.5%, averaging 66.8%. (3) 1/3 of the karyotypic abnormalities would be missed, and therefore, replacement of complete karyotyping with FISH would have unacceptably high false-negative rates for routine evaluation. (4) FISH-CSP, while useful when positive for anomalies, is not sufficient when negative to obviate the need for a complete karyotype.

  11. Miller-Dieker syndrome resulting from rearrangement of a familial chromosome 17 inversion detected by fluorescence in situ hybridisation.

    PubMed Central

    Kingston, H M; Ledbetter, D H; Tomlin, P I; Gaunt, K L

    1996-01-01

    We report a case of Miller-Dieker syndrome (MDS) owing to an unbalanced rearrangement of a familial pericentric inversion of chromosome 17 (inv(17) (p13.3q25.1)). In addition to lissencephaly and the facial features of MDS, the affected child had other congenital malformations consistent with distal 17q duplication. Initial cytogenetic analysis failed to show any abnormality and fluorescence in situ hybridisation (FISH) studies confirmed the 17p deletion in the proband and identified the chromosome 17 inversion in his mother. FISH studies were performed in other relatives and enabled first trimester prenatal diagnosis by chorionic villus sampling in a subsequent pregnancy of the proband's mother. These findings underline the value of FISH in the investigation of MDS families. Images PMID:8825053

  12. Chromosomal Flexibility

    ERIC Educational Resources Information Center

    Journal of College Science Teaching, 2005

    2005-01-01

    Scientists have shown that a genetic element on one chromosome may direct gene activity on another. Howard Hughes Medical Institute (HHMI) researchers report that a multitasking master-control region appears to over-see both a set of its own genes and a related gene on a nearby chromosome. The findings reinforce the growing importance of location…

  13. Chromosomal Flexibility

    ERIC Educational Resources Information Center

    Journal of College Science Teaching, 2005

    2005-01-01

    Scientists have shown that a genetic element on one chromosome may direct gene activity on another. Howard Hughes Medical Institute (HHMI) researchers report that a multitasking master-control region appears to over-see both a set of its own genes and a related gene on a nearby chromosome. The findings reinforce the growing importance of location…

  14. Modeling Chromosomes

    ERIC Educational Resources Information Center

    Robertson, Carol

    2016-01-01

    Learning about chromosomes is standard fare in biology classrooms today. However, students may find it difficult to understand the relationships among the "genome", "chromosomes", "genes", a "gene locus", and "alleles". In the simple activity described in this article, which follows the 5E approach…

  15. Modeling Chromosomes

    ERIC Educational Resources Information Center

    Robertson, Carol

    2016-01-01

    Learning about chromosomes is standard fare in biology classrooms today. However, students may find it difficult to understand the relationships among the "genome", "chromosomes", "genes", a "gene locus", and "alleles". In the simple activity described in this article, which follows the 5E approach…

  16. DETECTING STREAM INVERTEBRATE COMMUNITY ALTERATION DUE TO MID TO LOW LEVELS OF WATERSHED LANDSCAPE MODIFICATION

    EPA Science Inventory

    As part of an investigation into the effects of watershed landscape alteration on stream ecosystems, quantitative invertebrate samples were collected from riffles in 26 second and third order south shore Lake Superior streams. Nonmetric multidimensional scaling (NMDS) ordination ...

  17. Chromosomal abnormalities in couples with repeated fetal loss: An Indian retrospective study

    PubMed Central

    Sheth, Frenny J; Liehr, Thomas; Kumari, Pritti; Akinde, Ralph; Sheth, Harsh J; Sheth, Jayesh J

    2013-01-01

    BACKGROUND: Recurrent pregnancy loss is a common occurrence and a matter of concern for couples planning the pregnancy. Chromosomal abnormalities, mainly balanced rearrangements, are common in couples with repeated miscarriages. PURPOSE: The purpose of this study is to evaluate the contribution of chromosomal anomalies causing repeated spontaneous miscarriages and provide detailed characterization of a few structurally altered chromosomes. MATERIALS AND METHODS: A retrospective cytogenetic study was carried out on 4859 individuals having a history of recurrent miscarriages. The cases were analyzed using G-banding and fluorescence in situ hybridization wherever necessary. RESULTS: Chromosomal rearrangements were found in 170 individuals (3.5%). Translocations were seen in 72 (42.35%) cases. Of these, reciprocal translocations constituted 42 (24.70%) cases while Robertsonian translocations were detected in 30 (17.64%) cases. 7 (4.11%) cases were mosaic, 8 (4.70%) had small supernumerary marker chromosomes and 1 (0.6%) had an interstitial microdeletion. Nearly, 78 (1.61%) cases with heteromorphic variants were seen of which inversion of Y chromosome (57.70%) and chromosome 9 pericentromeric variants (32.05%) were predominantly involved. CONCLUSIONS: Chromosomal analysis is an important etiological investigation in couples with repeated miscarriages. Characterization of variants/marker chromosome enable calculation of a more precise recurrent risk in a subsequent pregnancy thereby facilitating genetic counseling and deciding further reproductive options. PMID:24497706

  18. Flow karyotyping and sorting of human chromosomes

    SciTech Connect

    Gray, J.W.; Lucas, J.; Peters, D.; Pinkel, D.; Trask, B.; van den Engh, G.; Van Dilla, M.A.

    1986-07-16

    Flow cytometry and sorting are becoming increasingly useful as tools for chromosome classfication and for the detection of numerical and structural chromosome aberrations. Chromosomes of a single type can be purified with these tools to facilitate gene mapping or production of chromosome specific recombinant DNA libraries. For analysis of chromosomes with flow cytometry, the chromosomes are extracted from mitotic cells, stained with one or more fluorescent dyes and classified one-by-one according to their dye content(s). Thus, the flow approach is fundamentally different than conventional karyotyping where chromosomes are classified within the context of a metaphase spread. Flow sorting allows purification of chromosomes that can be distinguished flow cytometrically. The authors describe the basic principles of flow cytometric chromosome classification i.e. flow karyotyping, and chromosome sorting and describe several applications. 30 refs., 8 figs.

  19. MRD detection in B-cell non-Hodgkin lymphomas using Ig gene rearrangements and chromosomal translocations as targets for real-time quantitative PCR.

    PubMed

    Pott, Christiane; Brüggemann, Monika; Ritgen, Matthias; van der Velden, Vincent H J; van Dongen, Jacques J M; Kneba, Michael

    2013-01-01

    Minimal residual disease (MRD) diagnostics is of high clinical relevance in patients with indolent B-cell Non-Hodgkin lymphomas (B-NHL) and serves as a surrogate parameter to evaluate treatment effectiveness and long-term prognosis. MRD diagnostics performed by real-time quantitative PCR (RQ-PCR) is the gold-standard and currently the most sensitive and the most broadly applied method in follicular lymphoma (FL) and mantle cell lymphoma (MCL). RQ-PCR analysis of the junctional regions of the rearranged immunoglobulin heavy-chain gene (IgH) serves as the most broadly applicable MRD target in B-NHL (∼80%). Chromosomal translocations as t(14;18) translocation in FL and t(11;14) translocation in MCL can be used in selected lymphoma subtypes. In patients with B-cell chronic lymphocytic leukemia, both flow-cytometry as well as RQ-PCR are equally suitable for MRD assessment as long as a sensitivity of ≤10(-4) shall be achieved.MRD diagnostics targeting the IgH gene is complex and requires extensive knowledge and experience because the junctional regions of each lymphoma have to be identified before the patient-specific RQ-PCR assays can be designed for MRD monitoring. Furthermore, somatic mutations of the IgH region occurring during B-cell development of germinal center and post-germinal center lymphomas may hamper appropriate primer binding leading to false negative results. The translocations mentioned above have the advantage that consensus forward primers and probes, both placed in the breakpoint regions of chromosome 18 in FL and chromosome 11 in MCL, can be used in combination with a reverse primer placed in the IgH joining region of chromosome 14. RQ-PCR-based methods can reach a good sensitivity (≤10(-4)). This chapter provides all relevant background information and technical aspects for the complete laboratory process from detection of the clonal IgH gene rearrangement and the chromosomal translocations at diagnosis to the actual MRD measurements in

  20. Molecular genetic approach to human meningioma: loss of genes on chromosome 22

    SciTech Connect

    Seizinger, B.R.; De La Monte, S.; Atkins, L.; Gusella, J.F.; Martuza, R.L.

    1987-08-01

    A molecular genetic approach employing polymorphic DNA markers has been used to investigate the role of chromosomal aberrations in meningioma, one of the most common tumors of the human nervous system. Comparison of the alleles detected by DNA markers in tumor DNA versus DNA from normal tissue revealed chromosomal alterations present in primary surgical specimens. In agreement with cytogenetic studies of cultured meningiomas, the most frequent alteration detected was loss of heterozygosity on chromosome 22. Forty of 51 patients were constitutionally heterozygous for at least one chromosome 22 DNA marker. Seventeen of the 40 constitutionally heterozygotic patients (43%) displayed hemizygosity for the corresponding marker in their meningioma tumor tissues. Loss of heterozygosity was also detected at a significantly lower frequency for markers on several other autosomes. In view of the striking association between acoustic neuroma and meningioma in bilateral acoustic neurofibromatosis and the discovery that acoustic neuromas display specific loss of genes on chromosome 22, the authors propose that a common mechanism involving chromosome 22 is operative in the development of both tumor types. Fine-structure mapping to reveal partial deletions in meningiomas may provide the means to clone and characterize a gene (or genes) of importance for tumorigenesis in this and possibly other clinically associated tumors of the human nervous system.

  1. The hippocampi of children with chromosome 22q11.2 deletion syndrome have localized anterior alterations that predict severity of anxiety

    PubMed Central

    Scott, Julia A.; Goodrich-Hunsaker, Naomi; Kalish, Kristopher; Lee, Aaron; Hunsaker, Michael R.; Schumann, Cynthia M.; Carmichael, Owen T.; Simon, Tony J.

    2016-01-01

    Background Individuals with 22q11.2 deletion syndrome (22q11.2DS) have an elevated risk for schizophrenia, which increases with history of childhood anxiety. Altered hippocampal morphology is a common neuroanatomical feature of 22q11.2DS and idiopathic schizophrenia. Relating hippocampal structure in children with 22q11.2DS to anxiety and impaired cognitive ability could lead to hippocampus-based characterization of psychosis-proneness in this at-risk population. Methods We measured hippocampal volume using a semiautomated approach on MRIs collected from typically developing children and children with 22q11.2DS. We then analyzed hippocampal morphology with Localized Components Analysis. We tested the modulating roles of diagnostic group, hippocampal volume, sex and age on local hippocampal shape components. Lastly, volume and shape components were tested as covariates of IQ and anxiety. Results We included 48 typically developing children and 69 children with 22q11.2DS in our study. Hippocampal volume was reduced bilaterally in children with 22q11.2DS, and these children showed greater variation in the shape of the anterior hippocampus than typically developing children. Children with 22q11.2DS had greater inward deformation of the anterior hippocampus than typically developing children. Greater inward deformation of the anterior hippocampus was associated with greater severity of anxiety, specifically fear of physical injury, within the 22q11.2DS group. Limitations Shape alterations are not specific to hippocampal subfields. Conclusion Alterations in the structure of the anterior hippocampus likely affect function and may impact limbic circuitry. We suggest these alterations potentially contribute to anxiety symptoms in individuals with 22q11.2DS through modulatory pathways. Altered hippocampal morphology may be uniquely linked to anxiety risk factors for schizophrenia, which could be a powerful neuroanatomical marker of schizophrenia risk and hence protection

  2. Genome‐wide significant schizophrenia risk variation on chromosome 10q24 is associated with altered cis‐regulation of BORCS7, AS3MT, and NT5C2 in the human brain

    PubMed Central

    Duarte, Rodrigo R. R.; Troakes, Claire; Nolan, Matthew; Srivastava, Deepak P.; Murray, Robin M.

    2016-01-01

    Chromosome 10q24.32‐q24.33 is one of the most robustly supported risk loci to emerge from genome‐wide association studies (GWAS) of schizophrenia. However, extensive linkage disequilibrium makes it difficult to distinguish the actual susceptibility gene(s) at the locus, limiting its value for improving biological understanding of the condition. In the absence of coding changes that can account for the association, risk is likely conferred by altered regulation of one or more genes in the region. We, therefore, used highly sensitive measures of allele‐specific expression to assess cis‐regulatory effects associated with the two best‐supported schizophrenia risk variants (SNP rs11191419 and indel ch10_104957618_I/rs202213518) on the primary positional candidates BORCS7, AS3MT, CNNM2, and NT5C2 in the human brain. Heterozygosity at rs11191419 was associated with increased allelic expression of BORCS7 and AS3MT in the fetal and adult brain, and with reduced allelic expression of NT5C2 in the adult brain. Heterozygosity at ch10_104957618_I was associated with reduced allelic expression of NT5C2 in both the fetal and adult brain. Comparisons between cDNA ratios in heterozygotes and homozygotes for the risk alleles indicated that cis‐effects on NT5C2 expression in the adult dorsolateral prefrontal cortex could be largely accounted for by genotype at these two risk variants. While not excluding effects on other genes in the region, this study implicates altered neural expression of BORCS7, AS3MT, and NT5C2 in susceptibility to schizophrenia arising from genetic variation at the chromosome 10q24 locus. © 2016 The Authors. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics Published by Wiley Periodicals, Inc. PMID:27004590

  3. Genome-wide significant schizophrenia risk variation on chromosome 10q24 is associated with altered cis-regulation of BORCS7, AS3MT, and NT5C2 in the human brain.

    PubMed

    Duarte, Rodrigo R R; Troakes, Claire; Nolan, Matthew; Srivastava, Deepak P; Murray, Robin M; Bray, Nicholas J

    2016-09-01

    Chromosome 10q24.32-q24.33 is one of the most robustly supported risk loci to emerge from genome-wide association studies (GWAS) of schizophrenia. However, extensive linkage disequilibrium makes it difficult to distinguish the actual susceptibility gene(s) at the locus, limiting its value for improving biological understanding of the condition. In the absence of coding changes that can account for the association, risk is likely conferred by altered regulation of one or more genes in the region. We, therefore, used highly sensitive measures of allele-specific expression to assess cis-regulatory effects associated with the two best-supported schizophrenia risk variants (SNP rs11191419 and indel ch10_104957618_I/rs202213518) on the primary positional candidates BORCS7, AS3MT, CNNM2, and NT5C2 in the human brain. Heterozygosity at rs11191419 was associated with increased allelic expression of BORCS7 and AS3MT in the fetal and adult brain, and with reduced allelic expression of NT5C2 in the adult brain. Heterozygosity at ch10_104957618_I was associated with reduced allelic expression of NT5C2 in both the fetal and adult brain. Comparisons between cDNA ratios in heterozygotes and homozygotes for the risk alleles indicated that cis-effects on NT5C2 expression in the adult dorsolateral prefrontal cortex could be largely accounted for by genotype at these two risk variants. While not excluding effects on other genes in the region, this study implicates altered neural expression of BORCS7, AS3MT, and NT5C2 in susceptibility to schizophrenia arising from genetic variation at the chromosome 10q24 locus. © 2016 The Authors. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics Published by Wiley Periodicals, Inc.

  4. Detection of alteration associated with a porphyry copper deposit in southern Arizona

    NASA Technical Reports Server (NTRS)

    Abrams, M. J.; Siegal, B. S.

    1977-01-01

    Computer processing of Landsat MSS data was performed using contrast stretching and band-to-band ratioing. A false color ratio composite picture showed color anomalies which coincided with known areas of alteration on and about Red Mountain. A helicopter survey of the study area was undertaken using a portable field reflectance spectrometer. One hundred fifty-six spectra were obtained in the 0.4 to 2.5 micrometer wavelength region. The spectra were digitized, and contour maps for 24 wavelength intervals were produced; no spectral anomalies were evident for the known altered areas. A contour map produced from the 1.6 and 2.2 micrometer ratio generally delineated the alteration areas. The 1.3, 1.6, and 2.2 micrometer wavelength data were canonically transformed using a transformation empirically derived from discriminant function analysis of altered and unaltered materials for the Goldfield, Nevada region, and a contour map was produced for the first canonical variable. The known areas of alteration were clearly defined on the contour map.

  5. Detecting long-range chromatin interactions using the chromosome conformation capture sequencing (4C-seq) method.

    PubMed

    Gheldof, Nele; Leleu, Marion; Noordermeer, Daan; Rougemont, Jacques; Reymond, Alexandre

    2012-01-01

    Eukaryotic transcription is tightly regulated by transcriptional regulatory elements, even though these elements may be located far away from their target genes. It is now widely recognized that these regulatory elements can be brought in close proximity through the formation of chromatin loops, and that these loops are crucial for transcriptional regulation of their target genes. The chromosome conformation capture (3C) technique presents a snapshot of long-range interactions, by fixing physically interacting elements with formaldehyde, digestion of the DNA, and ligation to obtain a library of unique ligation products. Recently, several large-scale modifications to the 3C technique have been presented. Here, we describe chromosome conformation capture sequencing (4C-seq), a high-throughput version of the 3C technique that combines the 3C-on-chip (4C) protocol with next-generation Illumina sequencing. The method is presented for use in mammalian cell lines, but can be adapted to use in mammalian tissues and any other eukaryotic genome.

  6. Chromosomal aberrations in human hepatocellular carcinomas associated with hepatitis C virus infection detected by comparative genomic hybridization

    PubMed Central

    Sakakura, C; Hagiwara, A; Taniguchi, H; Yamaguchi, T; Yamagishi, H; Takahashi, T; Koyama, K; Nakamura, Y; Abe, T; Inazawa, J

    1999-01-01

    Thirty-five hepatocellular carcinomas (HCCs) associated with hepatitis C virus (HCV) were analysed by comparative genomic hybridization (CGH), to screen for changes in copy-number of DNA sequences. Chromosomal losses were noted in 1p34–36 (37%), 4q12–21 (48%), 5q13–21 (35%), 6q13–16 (23%), 8p21–23 (28%), 13q (20%), 16q (33%) and 17p13 (37%). Gains were noted in 1q (46%), 6p (20%), 8q21–24 (31%) and 17q (43%). High level gains indicative of gene amplifications were found in 7q31 (3%), 11q13 (3%), 14q12 (6%) and 17q12 (3%); amplification at 14q12 may be characteristic for HCCs. No significant difference in chromosomal aberrations was noted between carcinomas associated with HCV-infection in our study and those reported earlier in HCCs infected with hepatitis B virus (HBV), indicating that both HBV- and HCV-related carcinomas may progress through a similar cascade of molecular events. © 1999 Cancer Research Campaign PMID:10471057

  7. Chromosome Abnormalities

    MedlinePlus

    ... division. There are two kinds of cell division, mitosis and meiosis. Mitosis results in two cells that are duplicates of ... 23 paired sets of chromosomes in nonreproductive cells. Mitosis: Cell division resulting in cells that have paired ...

  8. Identification of discrete chromosomal deletion by binary recursive partitioning of microarray differential expression data.

    PubMed

    Zhou, X; Cole, S W; Rao, N P; Cheng, Z; Li, Y; McBride, J; Wong, D T W

    2005-05-01

    DNA copy number abnormalities (CNA) are characteristic of tumours, and are also found in association with congenital anomalies and mental retardation. The ultimate impact of copy number abnormalities is manifested by the altered expression of the encoded genes. We previously developed a statistical method for the detection of simple chromosomal amplification using microarray expression data. In this study, we significantly advanced those analytical techniques to allow detection of localised chromosomal deletions based on differential gene expression data. Using three cell lines with known chromosomal deletions as model system, mRNA expression in those cells was compared with that observed in diploid cell lines of matched tissue origin. Results show that genes from deleted chromosomal regions are substantially over-represented (p<0.000001 by chi2) among genes identified as underexpressed in deletion cell lines relative to normal matching cells. Using a likelihood based statistical model, we were able to identify the breakpoint of the chromosomal deletion and match with the karyotype data in each cell line. In one such cell line, our analyses refined a previously identified 10p chromosomal deletion region. The deletion region was mapped to between 10p14 and 10p12, which was further confirmed by subtelomeric fluorescence in situ hybridisation. These data show that microarray differential expression data can be used to detect and map the boundaries of submicroscopic chromosomal deletions.

  9. [Chromosome analysis and genetic testing].

    PubMed

    Isobe, Yasushi; Miura, Ikuo

    2014-03-01

    Chromosomal and genetic tests are essential to establish correct diagnoses of the lymphoma. When the tissue examination is planned, these should be done simultaneously with the morphological and immunophenotypic evaluations. Chromosome analyses can identify the genomic alterations of tumor cells. Some chromosome abnormalities define disease subtypes. For example, recurrent 14q32 translocations involving the immunoglobulin heavy chain locus support the diagnosis of B-cell lymphoma, and their translocation partners identify the types. In contrast, genetic testings are performed to confirm the presence of certain abnormalities including gene rearrangements, mutations, amplifications and deletions in each case. These results provide us detailed information for diagnosis, prognosis, and choice of therapy.

  10. Prenatal diagnosis and carrier detection of a cryptic translocation by using DNA markers from the short arm of chromosome 5.

    PubMed Central

    Overhauser, J; Bengtsson, U; McMahon, J; Ulm, J; Butler, M G; Santiago, L; Wasmuth, J J

    1989-01-01

    DNA markers from the short arm of chromosome 5 were used to examine a large family in which a microscopically undetectable translocation was segregating. In addition to confirming that three retarded children were hemizygous for loci distal to 5p14, these analyses identified five individuals as being carriers of the balanced translocation. The use of molecular probes provided informed genetic counseling to the family for the first time. With the DNA markers from 5p, prenatal diagnosis was performed on two fetal chorionic villus samples, both of which were found to have unbalanced karyotypes. The identification of translocation carriers was complicated by recombination between the small translocated segment of 5p and the corresponding region on the normal homologue, which changed the haplotype of the translocated 5p segment. Images Figure 6 Figure 2 Figure 4 PMID:2667351

  11. A new essential hypertension susceptibility locus on chromosome 2p24-p25, detected by genomewide search.

    PubMed

    Angius, Andrea; Petretto, Enrico; Maestrale, Giovanni Battista; Forabosco, Paola; Casu, Giuseppina; Piras, Daniela; Fanciulli, Manuela; Falchi, Mario; Melis, Paola Maria; Palermo, Mario; Pirastu, Mario

    2002-10-01

    Essential hypertension (EH) is a complex disorder that results from the interaction of a number of susceptibility genes and environmental factors. We studied an isolated Sardinian village (Talana) in which the prevalence of hypertension is comparable to that in most Western populations. Talana exhibits features, such as slow demographic growth, high inbreeding, a low number of founders, stable lifestyle and culture, and accurate genealogical records, that make it suitable for the study of complex disorders. Clinical assessment of the entire adult population (N= approximately 1,000) identified approximately 100 hypertensive subjects. For our study, we selected the individuals with the most-severe EH (i.e., diastolic blood pressure >100 mm Hg), belonging to a single deep-rooted pedigree (12 generations), whose common ancestors lived in the 17th century. We performed a three-stage genomewide search using 36 affected individuals, by means of parametric linkage and allele-sharing approaches. LOD scores >1 were observed on chromosomes 1, 2, 13, 15, 17, and 19 (stage I). The most striking result was found in a 7.57-cM region on chromosome 2p24-p25. All five nonparametric linkage statistics estimated by the SimWalk2 program lie above the significance threshold of P<.008 for the whole region. Similar significance was obtained for 2p24-25 when parametric linkage (LOD score 1.99) and linkage disequilibrium mapping (P=.00006) were used, suggesting that a hypertension-susceptibility locus is located between D2S2278 and D2S168. This finding is strengthened by a recent report of linkage with marker D2S168 in a hypertensive sib-pair sample from China.

  12. Genome-wide scan for serum ghrelin detects linkage on chromosome 1p36 in Hispanic children: results from the Viva La Familia study.

    PubMed

    Voruganti, V Saroja; Göring, Harald H H; Diego, Vincent P; Cai, Guowen; Mehta, Nitesh R; Haack, Karin; Cole, Shelley A; Butte, Nancy F; Comuzzie, Anthony G

    2007-10-01

    This study was conducted to investigate genetic influence on serum ghrelin and its relationship with adiposity-related phenotypes in Hispanic children (n=1030) from the Viva La Familia study (VFS). Anthropometric measurements and levels of serum ghrelin were estimated and genetic analyses conducted according to standard procedures. Mean age, body mass index (BMI), and serum ghrelin were 11+/-0.13 y, 25+/-0.24 kg/m2 and 38+/-0.5 ng/mL, respectively. Significant heritabilities (p<0.001) were obtained for BMI, weight, fat mass, percent fat, waist circumference, waist-to-height ratio, and ghrelin. Bivariate analyses of ghrelin with adiposity traits showed significant negative genetic correlations (p<0.0001) with weight, BMI, fat mass, percent fat, waist circumference, and waist-to-height ratio. A genome-wide scan for ghrelin detected significant linkage on chromosome 1p36.2 between STR markers D1S2697 and D1S199 (LOD=3.2). The same region on chromosome 1 was the site of linkage for insulin (LOD=3.3), insulinlike growth factor binding protein 1 (IGFBP1) (LOD=3.4), homeostatic model assessment method (HOMA) (LOD=2.9), and C-peptide (LOD=2.0). Several family-based studies have reported linkages for obesity-related phenotypes in the region of 1p36. These results indicate the importance of this region in relation to adiposity in children from the VFS.

  13. PTEN loss and chromosome 8 alterations in Gleason grade 3 prostate cancer cores predicts the presence of un-sampled grade 4 tumor: implications for Active Surveillance

    PubMed Central

    Trock, Bruce J.; Fedor, Helen; Gurel, Bora; Jenkins, Robert B.; Knudsen, BS; Fine, Samson W.; Said, Jonathan W.; Carter, H. Ballentine; Lotan, Tamara L.; De Marzo, Angelo M.

    2016-01-01

    Men who enter active surveillance because their biopsy exhibits only Gleason grade 3 (G3) frequently have higher grade tumor missed by biopsy. Thus, biomarkers are needed that, when measured on G3 tissue, can predict the presence of higher grade tumor in the whole prostate. We evaluated whether PTEN loss, chromosome 8q gain (MYC) and/or 8p loss (LPL) measured only on G3 cores is associated with un-sampled G4 tumor. A tissue microarray was constructed of prostatectomy tissue from patients whose prostates exhibited only Gleason score 3+3, only 3+4, or only 4+3 tumor (n=50 per group). Cores sampled only from areas of G3 were evaluated for PTEN loss by immunohistochemistry, and PTEN deletion, LPL/8p loss, and MYC/8q gain by fluorescence in situ hybridization (FISH). Biomarker results were compared between Gleason score 6 vs. 7 tumors using conditional logistic regression. PTEN protein loss, odds ratio=4.99, p=.033, MYC/8q gain, odds ratio=5.36, p=.010, and LPL/8p loss, odds ratio=3.96, p=.003 were significantly more common in G3 cores derived from Gleason 7 vs. Gleason 6 tumors. PTEN gene deletion was not statistically significant. Associations were stronger comparing Gleason 4+3 vs. 6 than for Gleason 3+4 vs. 6. MYC/8q gain, LPL/8p loss, and PTEN protein loss measured in G3 tissue microarray cores strongly differentiate whether the core comes from a Gleason 6 or Gleason 7 tumor. If validated to predict upgrading from G3 biopsy to prostatectomy these biomarkers could reduce the likelihood of enrolling high risk men and facilitate safe patient selection for active surveillance. PMID:27080984

  14. Effects of metric and harmonic rhythm on the detection of pitch alterations in melodic sequences.

    PubMed

    Smith, K C; Cuddy, L L

    1989-08-01

    Tested response time to alterations. Metric rhythm and harmonic rhythm of 13-note tonal sequences were either matched or mismatched. Metric rhythm (3/4 or 4/4 meter) was induced by dynamic accents. Harmonic rhythm was induced by implied chord progressions initiated on the first note and on either every third or every fourth note. Responses were not always faster for matched rhythms or for alterations occurring on the dynamic accent. Responses were consistently faster for sequences presented in 4/4 meter. Musically untrained Ss performed similarly to trained Ss, but were slower and more variable. Accuracy of recall on a music dictation task also favored 4/4 meter rather than matched rhythms. Coding of pitch content may have been facilitated by the structural framework of 4/4 meter rather than by expectancies arising from the match of temporal and pitch organization.

  15. Utility of linearly amplified RNA for RT-PCR detection of chromosomal translocations: validation using the t(2;5)(p23;q35) NPM-ALK chromosomal translocation.

    PubMed

    Schumacher, Jonathan A; Jenson, Stephen D; Elenitoba-Johnson, Kojo S J; Lim, Megan S

    2004-02-01

    The requirement for sufficient quantities of starting RNA has limited the ability to evaluate multiple transcripts using reverse transcriptase-polymerase chain reaction (RT-PCR). In this study, we demonstrate the utility of linear RNA amplification for RT-PCR analysis of multiple gene transcripts including a chromosomal translocation, using the t(2;5)(p23;q35) as a model. RNA from the t(2;5)-positive cell line, SU-DHL-1, and the t(2;5)-negative cell line, HUT-78, was extracted and exposed to two rounds of linear amplification. RT-PCR using cDNA from the resultant amplified (a) RNA and total RNA resulted in the 177 bp NPM-ALK fusion gene product from the SU-DHL-1 cell line, but not from aRNA or total RNA from the HUT-78 cell line. DNA sequencing of the RT-PCR products from total and aRNA of SU-DHL-1 cells demonstrated identical sequences corresponding to the NPM-ALK fusion gene. Evaluation of 25 snap-frozen tissue samples, including eight NPM-ALK-positive ALCLs demonstrated 100% concordance of t(2;5) detection between cDNA from total RNA and that from aRNA. Our results show that linear amplification of RNA can enhance starting RNA greater than 200-fold and can be used for rapid and specific detection of multiplex gene expression from a variety of sources. This method can generate a renewable archive of representative cDNA, which can be used for retrospective screening of stored samples as well as positive controls for the clinical molecular diagnostic laboratory.

  16. CHROMOSOME MICROMANIPULATION

    PubMed Central

    Nicklas, R. Bruce; Koch, Carol A.

    1969-01-01

    Kinetochore reorientation is the critical process ensuring normal chromosome distribution. Reorientation has been studied in living grasshopper spermatocytes, in which bivalents with both chromosomes oriented to the same pole (unipolar orientation) occur but are unstable: sooner or later one chromosome reorients, the stable, bipolar orientation results, and normal anaphase segregation to opposite poles follows. One possible source of stability in bipolar orientations is the normal spindle forces toward opposite poles, which slightly stretch the bivalent. This tension is lacking in unipolar orientations because all the chromosomal spindle fibers and spindle forces are directed toward one pole. The possible role of tension has been tested directly by micromanipulation of bivalents in unipolar orientation to artificially create the missing tension. Without exception, such bivalents never reorient before the tension is released; a total time "under tension" of over 5 hr has been accumulated in experiments on eight bivalents in eight cells. In control experiments these same bivalents reoriented from a unipolar orientation within 16 min, on the average, in the absence of tension. Controlled reorientation and chromosome segregation can be explained from the results of these and related experiments. PMID:5824068

  17. Tumor-induced lymph node alterations detected by MRI lymphography using gadolinium nanoparticles.

    PubMed

    Partridge, S C; Kurland, B F; Liu, C-L; Ho, R J Y; Ruddell, A

    2015-10-26

    Contrast-enhanced MRI lymphography shows potential to identify alterations in lymph drainage through lymph nodes (LNs) in cancer and other diseases. MRI studies have typically used low molecular weight gadolinium contrast agents, however larger gadolinium-loaded nanoparticles possess characteristics that could improve the specificity and sensitivity of lymphography. The performance of three gadolinium contrast agents with different sizes and properties was compared by 3T MRI after subcutaneous injection. Mice bearing B16-F10 melanoma footpad tumors were imaged to assess tumor-induced alterations in lymph drainage through tumor-draining popliteal and inguinal LNs versus contralateral uninvolved drainage. Gadolinium lipid nanoparticles were able to identify tumor-induced alterations in contrast agent drainage into the popliteal LN, while lower molecular weight or albumin-binding gadolinium agents were less effective. All of the contrast agents distributed in foci around the cortex and medulla of tumor-draining popliteal LNs, while they were restricted to the cortex of non-draining LNs. Surprisingly, second-tier tumor-draining inguinal LNs exhibited reduced uptake, indicating that tumors can also divert LN drainage. These characteristics of tumor-induced lymph drainage could be useful for diagnosis of LN pathology in cancer and other diseases. The preferential uptake of nanoparticle contrasts into tumor-draining LNs could also allow selective targeting of therapies to tumor-draining LNs.

  18. Tumor-induced lymph node alterations detected by MRI lymphography using gadolinium nanoparticles

    PubMed Central

    Partridge, S. C.; Kurland, B. F.; Liu, C.-L.; Ho, R. J. Y.; Ruddell, A.

    2015-01-01

    Contrast-enhanced MRI lymphography shows potential to identify alterations in lymph drainage through lymph nodes (LNs) in cancer and other diseases. MRI studies have typically used low molecular weight gadolinium contrast agents, however larger gadolinium-loaded nanoparticles possess characteristics that could improve the specificity and sensitivity of lymphography. The performance of three gadolinium contrast agents with different sizes and properties was compared by 3T MRI after subcutaneous injection. Mice bearing B16-F10 melanoma footpad tumors were imaged to assess tumor-induced alterations in lymph drainage through tumor-draining popliteal and inguinal LNs versus contralateral uninvolved drainage. Gadolinium lipid nanoparticles were able to identify tumor-induced alterations in contrast agent drainage into the popliteal LN, while lower molecular weight or albumin-binding gadolinium agents were less effective. All of the contrast agents distributed in foci around the cortex and medulla of tumor-draining popliteal LNs, while they were restricted to the cortex of non-draining LNs. Surprisingly, second-tier tumor-draining inguinal LNs exhibited reduced uptake, indicating that tumors can also divert LN drainage. These characteristics of tumor-induced lymph drainage could be useful for diagnosis of LN pathology in cancer and other diseases. The preferential uptake of nanoparticle contrasts into tumor-draining LNs could also allow selective targeting of therapies to tumor-draining LNs. PMID:26497382

  19. A 14-year follow-up of a case detected prenatally of partial trisomy 13q21.32-qter and monosomy 18q22.3-qter as a result of a maternal complex chromosome rearrangement involving chromosomes 6, 13, and 18.

    PubMed

    Quadrelli, Roberto; Quadrelli, Andrea; Milunsky, Aubrey; Zou, Ying S; Huang, Xin-Li; Viera, Estela; Mechoso, Búrix; Bellini, Sylvia; Costabel, Mariana; Vaglio, Alicia

    2009-06-01

    A balanced complex chromosome rearrangement (CCR) involving three chromosomes is rare and may lead to different types of aneuploid germ cells. We report here a 14-year follow-up of a boy with a karyotype defined as 46,XY,der(18)t(6;13;18)(q21;q21.32;q22.3).ish der(18)(13qter+,18qter-) characterized by multiple congenital abnormalities, including distinctive minor facial anomalies, short neck, abnormalities of the extremities, anogenital abnormalities, flexion contractures, especially at extremities, and severe mental and growth retardation. Chromosome analysis in the mother showed a CCR involving chromosomes 6, 13, and 18. This CCR was the result of a three-break rearrangement, and the derivative chromosome 13 consisted of parts of chromosomes 18 and 13. The karyotype of the child was not balanced, and resulted in partial trisomy for 13q and partial monosomy for 18q detected prenatally by conventional and molecular cytogenetics. Although such a karyotype and its phenotype have not previously been reported, we have compared the clinical and cytogenetic data from our patient with previously described cases of partial trisomy 13q and monosomy 18q despite different break points. We are presenting a new CCR in a woman with normal phenotype with a history of four early abortions and a long follow-up of her malformed newborn with partial 13q trisomy and 18q monosomy.

  20. Remote Detection and Geochemical Studies for Finding and Understanding Hydrocarbon-induced Rock Alterations

    NASA Astrophysics Data System (ADS)

    Petrovic, Ana

    The main objective of this work was to determine if there are characteristic mineral assemblages and chemical changes in areas affected by hydrocarbon microseepages. For this purpose remote sensing was utilized for mapping surficial rock alterations, and geochemical tools were used to understand the alteration processes. The key area chosen for this type of work were altered and unaltered Wingate Sandstone outcrops in Lisbon Valley, Utah. The Spectral Angle Mapper method was applied on HyMap hyperspectral images to classify the extent of altered and unaltered outcrops, as well as to map the changes in mineral content within the outcrops. The Spectral Feature Fitting and Band Ratio methods were used to identify lithological changes in the area. The commonly used band ratios for identification of calcite and kaolinite were modified for future use with multispectral data. Reflectance spectroscopy, thin section studies, major, minor, and trace element analyses, and stable carbon and oxygen studies on both bleached (altered) and unbleached (unaltered) samples were successfully used to delineate areas of similar rock composition and relate changes due to hydrocarbons leaking from underlying petroleum reservoirs. Unbleached Wingate Sandstone samples had higher hematite and feldspar content than bleached Wingate samples, which were characterized by larger amounts of clay, calcite, and pyrite. Some bleached samples also had higher concentrations of elements (U, Mo) characteristic of hydrocarbon-related reducing environments, and were depleted in 13C when compared to the unbleached samples. Based on these results, the following model of chemical reactions is suggested for diagnostic changes within Wingate Sandstone. Hydrocarbon-induced reducing environment caused the transformation of sulfate ion (obtained from groundwater or from oxidation of H2S) to sulfide ion, resulting in the reduction of hematite to pyrite, The released hydrogen ion from this reaction reacted with

  1. Alteration mineralogy and geochemistry as an exploration tool for detecting basement heat sources in sedimentary basins

    NASA Astrophysics Data System (ADS)

    Uysal, Tonguc; Gasparon, Massimo; van Zyl, Jacobus; Wyborn, Doone

    2010-05-01

    The Cooper Basin located in South Australia and Queensland hosts some of the hottest granites in the world at economic drilling depths (240°C at 3.5 km). Investigating the mechanism of heat-producing element enrichment in the Cooper Basin granite is crucial for understanding hot-dry rock geothermal systems and developing exploration strategies. Trace element (by ICP-MS) and stable isotope geochemistry of whole rock granite samples and hydrothermal phyllosilicate alteration minerals separated from the granite and overlying sandstones and mudstones of the Cooper Basin were examined in detail. Granite core samples from relatively shallow depths in Moomba 1 and Big Lake 1 are strongly altered with pervasive sericite (illite) and quartz precipitation, probably associated with intense micro-fracturing and veining. The intensity of hydrothermal alteration is less in deeper samples from Mcleod 1, Jolokia and Habanero 1. Highly altered granites from former holes are substantially enriched in lithophile elements, particularly in Cs, Rb, Be, Th, U and rare earth elements (REE) relative to the upper continental crust (UCC). U and Th contents with concentrations of up to 30 and 144 ppm, respectively, are 10 and 13 times higher than those of the UCC. Comparison of the trace element composition of the same samples dissolved by open beaker acid digestion and high-pressure acid bomb digestion (to dissolve zircon) shows that zircon is not the main repository of U and Th in the Cooper Basin granite. Instead, we propose that the enrichment of heat-producing elements was promoted by a regional hydrothermal event leading to the precipitation of U and Th- bearing minerals such as illite, K-feldspar and thorite. Crystallinity index (illite crystallinity) of the sericite indicates hydrothermal temperatures ranging from 250°C (in Moomba 1 and Big Lake 1) to 350°C (in McLeod 1 and Jolokia 1). In the overlying sedimentary rocks, crystallinity of authigenic illites translates to lower

  2. Sex Chromosome Drive

    PubMed Central

    Helleu, Quentin; Gérard, Pierre R.; Montchamp-Moreau, Catherine

    2015-01-01

    Sex chromosome drivers are selfish elements that subvert Mendel's first law of segregation and therefore are overrepresented among the products of meiosis. The sex-biased progeny produced then fuels an extended genetic conflict between the driver and the rest of the genome. Many examples of sex chromosome drive are known, but the occurrence of this phenomenon is probably largely underestimated because of the difficulty to detect it. Remarkably, nearly all sex chromosome drivers are found in two clades, Rodentia and Diptera. Although very little is known about the molecular and cellular mechanisms of drive, epigenetic processes such as chromatin regulation could be involved in many instances. Yet, its evolutionary consequences are far-reaching, from the evolution of mating systems and sex determination to the emergence of new species. PMID:25524548

  3. Nuclear organisation in totipotent human nuclei and its relationship to chromosomal abnormality.

    PubMed

    Finch, Katie A; Fonseka, Gothami; Ioannou, Dimitris; Hickson, Nicholas; Barclay, Zoe; Chatzimeletiou, Katerina; Mantzouratou, Anna; Handyside, Alan; Delhanty, Joy; Griffin, Darren K

    2008-03-01

    Studies of nuclear organisation, most commonly determining the nuclear location of chromosome territories and individual loci, have furthered our understanding of nuclear function, differentiation and disease. In this study, by examining eight loci on different chromosomes, we tested hypotheses that: (1) totipotent human blastomeres adopt a nuclear organisation akin to that of committed cells; (2) nuclear organisation is different in chromosomally abnormal blastomeres; and (3) human blastomeres adopt a ;chromocentre' pattern. Analysis of in vitro fertilisation (IVF) conceptuses permits valuable insight into the cell biology of totipotent human nuclei. Here, extrapolations from images of preimplantation genetic screening (PGS) cases were used to make comparisons between totipotent blastomeres and several committed cells, showing some differences and similarities. Comparisons between chromosomally abnormal nuclei and those with no detected abnormality (NDA) suggest that the former display a significant non-random pattern for all autosomal loci, but there is a less distinct, possibly random, pattern in 'NDA' nuclei. No evidence was found that the presence of an extra chromosome is accompanied by an altered nuclear location for that chromosome. Centromeric loci on chromosomes 15 and 16 normally seen at the nuclear periphery were mostly centrally located in aneuploid cells, providing some evidence of a 'chromocentre'; however, the chromosome-18 centromere was more peripheral, similar to committed cells. Our results provide clues to the nature of totipotency in human cells and might have future applications for preimplantation diagnosis and nuclear transfer.

  4. Chromosome Cohesion Established by Rec8-Cohesin in Fetal Oocytes Is Maintained without Detectable Turnover in Oocytes Arrested for Months in Mice.

    PubMed

    Burkhardt, Sabrina; Borsos, Máté; Szydlowska, Anna; Godwin, Jonathan; Williams, Suzannah A; Cohen, Paula E; Hirota, Takayuki; Saitou, Mitinori; Tachibana-Konwalski, Kikuë

    2016-03-07

    Sister chromatid cohesion mediated by the cohesin complex is essential for chromosome segregation in mitosis and meiosis [1]. Rec8-containing cohesin, bound to Smc3/Smc1α or Smc3/Smc1β, maintains bivalent cohesion in mammalian meiosis [2-6]. In females, meiotic DNA replication and recombination occur in fetal oocytes. After birth, oocytes arrest at the prolonged dictyate stage until recruited to grow into mature oocytes that divide at ovulation. How cohesion is maintained in arrested oocytes remains a pivotal question relevant to maternal age-related aneuploidy. Hypothetically, cohesin turnover regenerates cohesion in oocytes. Evidence for post-replicative cohesion establishment mechanism exists, in yeast and invertebrates [7, 8]. In mouse fetal oocytes, cohesin loading factor Nipbl/Scc2 localizes to chromosome axes during recombination [9, 10]. Alternatively, cohesion is maintained without turnover. Consistent with this, cohesion maintenance does not require Smc1β transcription, but unlike Rec8, Smc1β is not required for establishing bivalent cohesion [11, 12]. Rec8 maintains cohesion without turnover during weeks of oocyte growth [3]. Whether the same applies to months or decades of arrest is unknown. Here, we test whether Rec8 activated in arrested mouse oocytes builds cohesion revealed by TEV cleavage and live-cell imaging. Rec8 establishes cohesion when activated during DNA replication in fetal oocytes using tamoxifen-inducible Cre. In contrast, no new cohesion is detected when Rec8 is activated in arrested oocytes by tamoxifen despite cohesin synthesis. We conclude that cohesion established in fetal oocytes is maintained for months without detectable turnover in dictyate-arrested oocytes. This implies that women's fertility depends on the longevity of cohesin proteins that established cohesion in utero.

  5. Copy-number variations on the X chromosome in Japanese patients with mental retardation detected by array-based comparative genomic hybridization analysis.

    PubMed

    Honda, Shozo; Hayashi, Shin; Imoto, Issei; Toyama, Jun; Okazawa, Hitoshi; Nakagawa, Eiji; Goto, Yu-Ichi; Inazawa, Johji

    2010-09-01

    X-linked mental retardation (XLMR) is a common, clinically complex and genetically heterogeneous disease arising from many mutations along the X chromosome. Although research during the past decade has identified >90 XLMR genes, many more remain uncharacterized. In this study, copy-number variations (CNVs) were screened in individuals with MR from 144 families by array-based comparative genomic hybridization (aCGH) using a bacterial artificial chromosome-based X-tiling array. Candidate pathogenic CNVs (pCNVs) were detected in 10 families (6.9%). Five of the families had pCNVs involving known XLMR genes, duplication of Xq28 containing MECP2 in three families, duplication of Xp11.22-p11.23 containing FTSJ1 and PQBP1 in one family, and deletion of Xp11.22 bearing SHROOM4 in one family. New candidate pCNVs were detected in five families as follows: identical complex pCNVs involved in dup(X)(p22.2) and dup(X)(p21.3) containing part of REPS2, NHS and IL1RAPL1 in two unrelated families, duplication of Xp22.2 including part of FRMPD4, duplication of Xq21.1 including HDX and deletion of Xq24 noncoding region in one family, respectively. Both parents and only mother samples were available in six and three families, respectively, and pCNVs were inherited from each of their mothers in those families other than a family of the proband with deletion of SHROOM4. This study should help to identify the novel XLMR genes and mechanisms leading to MR and reveal the clinical conditions and genomic background of XLMR.

  6. Support for schizophrenia susceptibility locus on chromosome 2q detected in a Swedish isolate using a dense map of microsatellites and SNPs.

    PubMed

    Aberg, Karolina; Axelsson, Elin; Saetre, Peter; Jiang, Lin; Wetterberg, Lennart; Pettersson, Ulf; Lindholm, Eva; Jazin, Elena

    2008-10-05

    Extended pedigrees are not only very useful to identify disease genes for rare Mendelian conditions, but they may also help unravel the genetics of complex diseases such as schizophrenia. In this study we performed genome-wide multipoint non-parametric linkage (NPL) score calculations using 825 microsatellites and 5,366 single nucleotide polymorphisms (SNPs), respectively, and searched for haplotypes shared by affected individuals, in three multiplex families including 29 genotyped affected individuals which in total contains 49 relative pairs useful for linkage studies. The most consistent results for microsatellites and SNPs were observed on 2q12.3-q14.1 (NPL scores 2.0, empirical P-value 0.009). However, the overall highest NPL score was observed on chromosome 2q33.3 using SNPs (NPL score 2.2, empirical P-value 0.007). Other chromosomal regions were detected on 5q15-q22.1, with microsatellites (NPL scores 1.7, empirical P-value 0.021) and with SNPs (NPL scores 2.0, empirical P-value 0.010) and on 5q23.1 (NPL score 1.9, empirical P-value 0.012) and 8q24.1-q24.2 (NPL score 2.1, empirical P-value 0.009) when using SNPs. The analysis of extended pedigrees allowed the search for haplotypes inherited identical by decent (IBD) by affected individuals. In all regions with NPL score >1.9 we found haplotypes inherited IBD by multiple cases. However, no common haplotypes were found for affected individuals in all families. In conclusion our NPL results support earlier findings suggesting that 2q and possibly 5q and 8q contain susceptibility loci for schizophrenia. Haplotype sharing in families helped to delimit the detected regions that potentially are susceptibility loci for schizophrenia.

  7. Chromosome Territories

    PubMed Central

    Cremer, Thomas; Cremer, Marion

    2010-01-01

    Chromosome territories (CTs) constitute a major feature of nuclear architecture. In a brief statement, the possible contribution of nuclear architecture studies to the field of epigenomics is considered, followed by a historical account of the CT concept and the final compelling experimental evidence of a territorial organization of chromosomes in all eukaryotes studied to date. Present knowledge of nonrandom CT arrangements, of the internal CT architecture, and of structural interactions with other CTs is provided as well as the dynamics of CT arrangements during cell cycle and postmitotic terminal differentiation. The article concludes with a discussion of open questions and new experimental strategies to answer them. PMID:20300217

  8. Automated clinical system for chromosome analysis

    NASA Technical Reports Server (NTRS)

    Castleman, K. R.; Friedan, H. J.; Johnson, E. T.; Rennie, P. A.; Wall, R. J. (Inventor)

    1978-01-01

    An automatic chromosome analysis system is provided wherein a suitably prepared slide with chromosome spreads thereon is placed on the stage of an automated microscope. The automated microscope stage is computer operated to move the slide to enable detection of chromosome spreads on the slide. The X and Y location of each chromosome spread that is detected is stored. The computer measures the chromosomes in a spread, classifies them by group or by type and also prepares a digital karyotype image. The computer system can also prepare a patient report summarizing the result of the analysis and listing suspected abnormalities.

  9. Simultaneous Detection of Both Single Nucleotide Variations and Copy Number Alterations by Next-Generation Sequencing in Gorlin Syndrome

    PubMed Central

    Morita, Kei-ichi; Naruto, Takuya; Tanimoto, Kousuke; Yasukawa, Chisato; Oikawa, Yu; Masuda, Kiyoshi; Imoto, Issei; Inazawa, Johji; Omura, Ken; Harada, Hiroyuki

    2015-01-01

    Gorlin syndrome (GS) is an autosomal dominant disorder that predisposes affected individuals to developmental defects and tumorigenesis, and caused mainly by heterozygous germline PTCH1 mutations. Despite exhaustive analysis, PTCH1 mutations are often unidentifiable in some patients; the failure to detect mutations is presumably because of mutations occurred in other causative genes or outside of analyzed regions of PTCH1, or copy number alterations (CNAs). In this study, we subjected a cohort of GS-affected individuals from six unrelated families to next-generation sequencing (NGS) analysis for the combined screening of causative alterations in Hedgehog signaling pathway-related genes. Specific single nucleotide variations (SNVs) of PTCH1 causing inferred amino acid changes were identified in four families (seven affected individuals), whereas CNAs within or around PTCH1 were found in two families in whom possible causative SNVs were not detected. Through a targeted resequencing of all coding exons, as well as simultaneous evaluation of copy number status using the alignment map files obtained via NGS, we found that GS phenotypes could be explained by PTCH1 mutations or deletions in all affected patients. Because it is advisable to evaluate CNAs of candidate causative genes in point mutation-negative cases, NGS methodology appears to be useful for improving molecular diagnosis through the simultaneous detection of both SNVs and CNAs in the targeted genes/regions. PMID:26544948

  10. Simultaneous Detection of Both Single Nucleotide Variations and Copy Number Alterations by Next-Generation Sequencing in Gorlin Syndrome.

    PubMed

    Morita, Kei-ichi; Naruto, Takuya; Tanimoto, Kousuke; Yasukawa, Chisato; Oikawa, Yu; Masuda, Kiyoshi; Imoto, Issei; Inazawa, Johji; Omura, Ken; Harada, Hiroyuki

    2015-01-01

    Gorlin syndrome (GS) is an autosomal dominant disorder that predisposes affected individuals to developmental defects and tumorigenesis, and caused mainly by heterozygous germline PTCH1 mutations. Despite exhaustive analysis, PTCH1 mutations are often unidentifiable in some patients; the failure to detect mutations is presumably because of mutations occurred in other causative genes or outside of analyzed regions of PTCH1, or copy number alterations (CNAs). In this study, we subjected a cohort of GS-affected individuals from six unrelated families to next-generation sequencing (NGS) analysis for the combined screening of causative alterations in Hedgehog signaling pathway-related genes. Specific single nucleotide variations (SNVs) of PTCH1 causing inferred amino acid changes were identified in four families (seven affected individuals), whereas CNAs within or around PTCH1 were found in two families in whom possible causative SNVs were not detected. Through a targeted resequencing of all coding exons, as well as simultaneous evaluation of copy number status using the alignment map files obtained via NGS, we found that GS phenotypes could be explained by PTCH1 mutations or deletions in all affected patients. Because it is advisable to evaluate CNAs of candidate causative genes in point mutation-negative cases, NGS methodology appears to be useful for improving molecular diagnosis through the simultaneous detection of both SNVs and CNAs in the targeted genes/regions.

  11. Global warming alters sound transmission: differential impact on the prey detection ability of echolocating bats

    PubMed Central

    Luo, Jinhong; Koselj, Klemen; Zsebők, Sándor; Siemers, Björn M.; Goerlitz, Holger R.

    2014-01-01

    Climate change impacts the biogeography and phenology of plants and animals, yet the underlying mechanisms are little known. Here, we present a functional link between rising temperature and the prey detection ability of echolocating bats. The maximum distance for echo-based prey detection is physically determined by sound attenuation. Attenuation is more pronounced for high-frequency sound, such as echolocation, and is a nonlinear function of both call frequency and ambient temperature. Hence, the prey detection ability, and thus possibly the foraging efficiency, of echolocating bats and susceptible to rising temperatures through climate change. Using present-day climate data and projected temperature rises, we modelled this effect for the entire range of bat call frequencies and climate zones around the globe. We show that depending on call frequency, the prey detection volume of bats will either decrease or increase: species calling above a crossover frequency will lose and species emitting lower frequencies will gain prey detection volume, with crossover frequency and magnitude depending on the local climatic conditions. Within local species assemblages, this may cause a change in community composition. Global warming can thus directly affect the prey detection ability of individual bats and indirectly their interspecific interactions with competitors and prey. PMID:24335559

  12. Global warming alters sound transmission: differential impact on the prey detection ability of echolocating bats.

    PubMed

    Luo, Jinhong; Koselj, Klemen; Zsebok, Sándor; Siemers, Björn M; Goerlitz, Holger R

    2014-02-06

    Climate change impacts the biogeography and phenology of plants and animals, yet the underlying mechanisms are little known. Here, we present a functional link between rising temperature and the prey detection ability of echolocating bats. The maximum distance for echo-based prey detection is physically determined by sound attenuation. Attenuation is more pronounced for high-frequency sound, such as echolocation, and is a nonlinear function of both call frequency and ambient temperature. Hence, the prey detection ability, and thus possibly the foraging efficiency, of echolocating bats and susceptible to rising temperatures through climate change. Using present-day climate data and projected temperature rises, we modelled this effect for the entire range of bat call frequencies and climate zones around the globe. We show that depending on call frequency, the prey detection volume of bats will either decrease or increase: species calling above a crossover frequency will lose and species emitting lower frequencies will gain prey detection volume, with crossover frequency and magnitude depending on the local climatic conditions. Within local species assemblages, this may cause a change in community composition. Global warming can thus directly affect the prey detection ability of individual bats and indirectly their interspecific interactions with competitors and prey.

  13. Fetal MRI detects early alterations of brain development in Tetralogy of Fallot.

    PubMed

    Schellen, Christoph; Ernst, Schwartz; Gruber, Gerlinde M; Mlczoch, Elisabeth; Weber, Michael; Brugger, Peter C; Ulm, Barbara; Langs, Georg; Salzer-Muhar, Ulrike; Prayer, Daniela; Kasprian, Gregor

    2015-09-01

    Prenatal imaging has identified alterations of brain growth in fetuses with congenital heart disease. However, little is known about the timing of altered brain development and its occurrence in specific congenital heart disease subgroups. This magnetic resonance imaging study aimed to identify early (median, 25 gestational weeks [GW]) changes in fetal total brain (TBV), gray matter (GMV), and subcortical brain (SBV) volumes in Tetralogy of Fallot (TOF) cases in utero. Fetal magnetic resonance imaging (1.5 Tesla) was performed in 24 fetuses who were diagnosed with TOF and 24 normal age-matched control fetuses (20-34 GW). TBV, GMV, SBV, intracranial cavity, cerebellar, ventricular, and external cerebrospinal fluid volumes were quantified by manual segmentation based on coronal T2-weighted sequences. Mixed model analyses of variance and t-tests were conducted to compare cases and control fetuses. TBV was significantly lower (P < .001) in early (<25 GW) and late TOF cases. Both GMV (P = .003) and SBV (P = .001) were affected. The GMV-to-SBV ratio declined in fetuses with TOF (P = .026). Compared with normal fetuses, ventricular volume was increased (P = .0048). External cerebrospinal fluid was enlarged in relation to head size (P < .001). Intracranial cavity volume (P = .314) and cerebellar volume (P = .074) were not significantly reduced in fetuses with TOF. TOF is associated with smaller volumes of gray and white matter and enlarged cerebrospinal fluid spaces. These changes are present at ≤25 GW and indicate altered fetal brain growth in this pathophysiologic entity during early stages of human brain development. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. Real-time, label-free isothermal solid-phase amplification/detection (ISAD) device for rapid detection of genetic alteration in cancers.

    PubMed

    Shin, Yong; Perera, Agampodi Promoda; Kim, Kyung Woo; Park, Mi Kyoung

    2013-06-07

    Here, we first present an isothermal solid-phase amplification/detection (ISAD) technique for the detection of single-point mutations that can be performed without labelling in real-time by utilizing both silicon microring-based solid-phase amplification and isothermal recombinase polymerase amplification (RPA). The ISAD technique was performed on a silicon microring device with a plastic chamber containing 10 μL of the reaction mixture, and characterized with an assay for the detection of the HRAS (Harvey RAS) gene single-point mutation. For the solid-phase amplification, the primer of the gene was directly attached to the surface of the device via an amine modification reaction. The amplified DNA was detected, without a label, by measuring the optical wavelength shift of the silicon microring resonator during the reaction. We demonstrated that the sensitivity of the ISAD technique was 100-times higher than that of RPA and conventional PCR methods. Moreover, this technique can be used to distinguish a single-point mutation of the HRAS gene via target amplification. This novel DNA amplification/detection technique will be useful for the detection of sequence alterations such as mutations and single-nucleotide polymorphisms as DNA biomarkers in human diseases.

  15. Detection of TTV in peripheral blood cells from patients with altered ALT and AST levels.

    PubMed

    de Oliveira, Jaqueline Carvalho; Nasser, Thiago Franco; Oda, Julie Massayo Maeda; Aoki, Mateus Nóbrega; Carneiro, Juliana Laino do Val; Barbosa, Décio Sabbatini; Reiche, Edna Maria Vissoci; Watanabe, Maria Angelica Ehara

    2008-04-01

    This work analyzes the prevalence of TTV DNA in peripheral blood cells from patients with hepatic alterations and healthy blood donors and measures levels of sodium, potassium, urea, creatinine, phosphatase alkaline, total and direct bilirubin, gamma glutamyl transferase (GGT), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in certain randomly selected patients. DNA samples from 111 individuals were evaluated. They were divided into two groups, "A" (study) and "B" (control), including 54 patients with liver enzyme alterations (ALT/AST) presenting non-B-non-C hepatitis and 57 blood donors, respectively. TTV DNA was determined by nested PCR. Certain products of the second-round PCR were sequenced. Serum biochemical assay was performed and disclosed TTV in 31.48% (17/54) of patients in group A and 5.26% (3/57) in the control group B. TTV prevalence was significantly higher in patients with liver disease than in healthy donors. In group A, sodium, potassium, urea, creatinine, phosphatase alkaline, total and direct bilirubin, gamma glutamyl transferase (GGT), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were analyzed in certain randomly selected patients and no significant difference in biochemical levels (p>0.05) was found when TTV infected and noninfected individuals were compared. Knowledge related to TTV has rapidly increased, but many fundamental aspects remain unclear. This led us to question the role of TTV and doubt remains as to whether or not it is just a commensal virus. Further studies are necessary to confirm and extend these findings.

  16. The heart in Duchenne muscular dystrophy: early detection of contractile performance alteration

    PubMed Central

    Wagner, Sören; Knipp, Stephan; Weber, Cornelia; Hein, Selina; Schinkel, Stefanie; Walther, Andreas; Bekeredjian, Raffi; Müller, Oliver J; Friedrich, Oliver

    2012-01-01

    Progressive cardiomyopathy is a major cause of death in Duchenne muscular dystrophy (DMD) patients. Coupling between Ca2+ handling and contractile properties in dystrophic hearts is poorly understood. It is also not clear whether developing cardiac failure is dominated by alterations in Ca2+ pathways or more related to the contractile apparatus. We simultaneously recorded force and Ca2+ transients in field-stimulated papillary muscles from young (10–14 weeks) wild-type (wt) and dystrophic mdx mice. Force amplitudes were fivefold reduced in mdx muscles despite only 30 % reduction in fura-2 ratio amplitudes. This indicated mechanisms other than systolic Ca2+ to additionally account for force decrements in mdx muscles. pCa-force relations revealed decreased mdx myofibrillar Ca2+ sensitivity. ‘In vitro’ motility assays, studied in mdx hearts here for the first time, showed significantly slower sliding velocities. mdx MLC/MHC isoforms were not grossly altered. Dystrophic hearts showed echocardiography signs of early ventricular wall hypertrophy with a significantly enlarged end-diastolic diameter ‘in vivo’. However, fractional shortening was still comparable to wt mice. Changes in the contractile apparatus satisfactorily explained force drop in mdx hearts. We give first evidence of early hypertrophy in mdx mice and possible mechanisms for already functional impairment of cardiac muscle in DMD. PMID:22970922

  17. Alterations in body fluid content can be detected by bioelectrical impedance analysis.

    PubMed

    Scheltinga, M R; Jacobs, D O; Kimbrough, T D; Wilmore, D W

    1991-05-01

    The electrical resistance across the whole body and its segments to the conduction of a weak alternating current was determined in human subjects under three different conditions: (1) during bed rest, (2) during infusion of 1 liter of saline, and (3) during donation of 1 unit of blood. During bed rest, extracellular and total body water were measured by dilution of bromide and heavy water, respectively. Electrical resistance obtained from electrodes placed on proximal portions of extremities ("proximal resistance") accounted for less than 50% of that determined by electrodes positioned on routinely used portions of a hand and foot ("whole body resistance"). Following saline infusion, resistance determined from the whole body and all its segments fell (P less than 0.001); the magnitude of the drop in both proximal and whole body resistance was inversely related to the volume of total body water (TBW) (r = -0.82, P less than 0.002, and r = -0.73, P less than 0.01, respectively). In contrast, blood donation was associated with significantly increased resistance at both measurement sites. TBW predicted from anthropometrics was inversely related to both proximal (r = -0.90, P less than 0.001) and whole body resistance (r = -0.75, P less than 0.001). Bioelectrical impedance analysis is a simple technique which may be useful in monitoring minimal alterations in TBW. Furthermore, altered fluid status may be predicted more accurately by changes in proximal resistance compared to changes in traditionally used whole body resistance.

  18. What's in your buffer? Solute altered millisecond motions detected by solution NMR.

    PubMed

    Wong, Madeline; Khirich, Gennady; Loria, J Patrick

    2013-09-17

    To date, little work has been conducted on the relationship between solute and buffer molecules and conformational exchange motion in enzymes. This study uses solution NMR to examine the effects of phosphate, sulfate, and acetate in comparison to MES- and HEPES-buffered references on the chemical shift perturbation and millisecond, chemical, or conformational exchange motions in the enzyme ribonuclease A (RNase A), triosephosphate isomerase (TIM) and HisF. The results indicate that addition of these solutes has a small effect on (1)H and (15)N chemical shifts for RNase A and TIM but a significant effect for HisF. For RNase A and TIM, Carr-Purcell-Meiboom-Gill relaxation dispersion experiments, however, show significant solute-dependent changes in conformational exchange motions. Some residues show loss of millisecond motions relative to the reference sample upon addition of solute, whereas others experience an enhancement. Comparison of exchange parameters obtained from fits of dispersion data indicates changes in either or both equilibrium populations and chemical shifts between conformations. Furthermore, the exchange kinetics are altered in many cases. The results demonstrate that common solute molecules can alter observed enzyme millisecond motions and play a more active role than what is routinely believed.

  19. Congenital Hypothyroidism with Neurological and Respiratory Alterations: A Case Detected Using a Variable Diagnostic Threshold for TSH

    PubMed Central

    Barreiro, Jesús; Castro-Feijoo, Lidia; Colón, Cristóbal; Cabanas, Paloma; Heredia, Claudia; Castaño, Luis Antonio; Gómez-Lado, Carmen; Couce, M.Luz; Pombo, Manuel

    2011-01-01

    We report a case of congenital hypothyroidism (CH) with neurological and respiratory alterations due to a heterozygotic c.374-1G > A mutation of TITF1/NKX2-1. The hypothyroidism was detected using a neonatal screening protocol in which the thyroid stimulating hormone (TSH) threshold is re-set each day on the basis of within-day variability and between-day variation. In this case, the threshold on the day of the initial analysis was 8.2 mIU/L, and the measured TSH level in heel-prick blood was 8.3 mIU/L. Conflict of interest:None declared. PMID:22155464

  20. NOR sites detected by Ag-dAPI staining of an unusual autosome chromosome of Bradysia hygida (Diptera:Sciaridae) colocalize with C-banded heterochromatic region.

    PubMed

    Gaspar, Vanessa Pinatto; Borges, Alex Rodrigues; Fernandez, Maria Aparecida

    2002-01-01

    The study of chromosomes in insects is a good tool in mitotic process analysis, zoographic localization and evolution investigation. Among them, the Sciaridae offers a karyotype with a small number of chromosomes, where the heterochromatin and nucleolar organizer region, NOR, are easily analyzed in metaphase chromosomes obtained from cerebral ganglia squashes. In this work, the heterochromatic regions on Bradysia hygida mitotic chromosomes, revealed by C-banding, were identified as centromeric blocks on A and C chromosomes and as dark interstitial region in B and X chromosomes. By Ag-DAPI staining, active nucleolus organizer region, NOR, was revealed associated to the constitutive heterochromatin in the end of the C autosome chromosome. The C-band regions and the unusual ribosomal site localization are discussed.

  1. Integrated Copy Number and Expression Analysis Identifies Profiles of Whole-Arm Chromosomal Alterations and Subgroups with Favorable Outcome in Ovarian Clear Cell Carcinomas

    PubMed Central

    Uehara, Yuriko; Oda, Katsutoshi; Ikeda, Yuji; Koso, Takahiro; Tsuji, Shingo; Yamamoto, Shogo; Asada, Kayo; Sone, Kenbun; Kurikawa, Reiko; Makii, Chinami; Hagiwara, Otoe; Tanikawa, Michihiro; Maeda, Daichi; Hasegawa, Kosei; Nakagawa, Shunsuke; Wada-Hiraike, Osamu; Kawana, Kei; Fukayama, Masashi; Fujiwara, Keiichi; Yano, Tetsu; Osuga, Yutaka; Fujii, Tomoyuki; Aburatani, Hiroyuki

    2015-01-01

    Ovarian clear cell carcinoma (CCC) is generally associated with chemoresistance and poor clinical outcome, even with early diagnosis; whereas high-grade serous carcinomas (SCs) and endometrioid carcinomas (ECs) are commonly chemosensitive at advanced stages. Although an integrated genomic analysis of SC has been performed, conclusive views on copy number and expression profiles for CCC are still limited. In this study, we performed single nucleotide polymorphism analysis with 57 epithelial ovarian cancers (31 CCCs, 14 SCs, and 12 ECs) and microarray expression analysis with 55 cancers (25 CCCs, 16 SCs, and 14 ECs). We then evaluated PIK3CA mutations and ARID1A expression in CCCs. SNP array analysis classified 13% of CCCs into a cluster with high frequency and focal range of copy number alterations (CNAs), significantly lower than for SCs (93%, P < 0.01) and ECs (50%, P = 0.017). The ratio of whole-arm to all CNAs was higher in CCCs (46.9%) than SCs (21.7%; P < 0.0001). SCs with loss of heterozygosity (LOH) of BRCA1 (85%) also had LOH of NF1 and TP53, and LOH of BRCA2 (62%) coexisted with LOH of RB1 and TP53. Microarray analysis classified CCCs into three clusters. One cluster (CCC-2, n = 10) showed more favorable prognosis than the CCC-1 and CCC-3 clusters (P = 0.041). Coexistent alterations of PIK3CA and ARID1A were more common in CCC-1 and CCC-3 (7/11, 64%) than in CCC-2 (0/10, 0%; P < 0.01). Being in cluster CCC-2 was an independent favorable prognostic factor in CCC. In conclusion, CCC was characterized by a high ratio of whole-arm CNAs; whereas CNAs in SC were mainly focal, but preferentially caused LOH of well-known tumor suppressor genes. As such, expression profiles might be useful for sub-classification of CCC, and might provide useful information on prognosis. PMID:26043110

  2. Utilization of the rpoB Gene as a Specific Chromosomal Marker for Real-Time PCR Detection of Bacillus anthracis

    PubMed Central

    Qi, Yuan; Patra, Guy; Liang, Xudong; Williams, Leanne E.; Rose, Sharon; Redkar, Rajendra J.; DelVecchio, Vito G.

    2001-01-01

    The potential use of Bacillus anthracis as a weapon of mass destruction poses a threat to humans, domesticated animals, and wildlife and necessitates the need for a rapid and highly specific detection assay. We have developed a real-time PCR-based assay for the specific detection of B. anthracis by taking advantage of the unique nucleotide sequence of the B. anthracis rpoB gene. Variable region 1 of the rpoB gene was sequenced from 36 Bacillus strains, including 16 B. anthracis strains and 20 other related bacilli, and four nucleotides specific for B. anthracis were identified. PCR primers were selected so that two B. anthracis-specific nucleotides were at their 3′ ends, whereas the remaining bases were specific to the probe region. This format permitted the PCR reactions to be performed on a LightCycler via fluorescence resonance energy transfer (FRET). The assay was found to be specific for 144 B. anthracis strains from different geographical locations and did not cross-react with other related bacilli (175 strains), with the exception of one strain. The PCR assay can be performed on isolated DNA as well as crude vegetative cell lysates in less than 1 h. Therefore, the rpoB-FRET assay could be used as a new chromosomal marker for rapid detection of B. anthracis. PMID:11472954

  3. Automatic segmentation of overlapping and touching chromosomes

    NASA Astrophysics Data System (ADS)

    Yuan, Zhiqiang; Chen, Xiaohua; Zhang, Renli; Yu, Chang

    2001-09-01

    This paper describes a technique to segment overlapping and touching chromosomes of human metaphase cells. Automated chromosome classification has been an important pattern recognition problem for decades, numerous attempts were made in the past to characterize chromosome band patterns. But successful separation between touching and overlapping chromosomes is vital for correct classification. Since chromosomes are non-rigid objects, common methods for separation between touching chromosomes are not usable. We proposed a method using shape concave and convex information, topology analysis information, and band pale paths for segmentation of touching and overlapping chromosomes. To detect shape concave and convex information, we should first pre-segment the chromosomes and get the edge of overlapping and touching chromosomes. After filtering the original image using edge-preserving filter, we adopt the Otsu's segmentation method and extract the boundary of chromosomes. Hence the boundary can be used for segment the overlapping and touching chromosomes by detecting the concave and convex information based on boundary information. Most of the traditional boundary-based algorithms detect corners based on two steps: the first step is to acquire the smoothed version of curvature at every point along the contour, and the second step is to detect the positions where curvature maximal occur and threshold the curvature as corner points. Recently wavelet transform has been adopted into corner detection algorithms. Since the metaphase overlapping chromosomes has multi-scale corners, we adopt a multi-scale corner detection method based on Hua's method for corner detection. For touching chromosomes, it is convenient to split them using pale paths. Starting from concave corner points, a search algorithm is represented. The searching algorithm traces three pixels into the object in the direction of the normal vector in order to avoid stopping at the initial boundary until it

  4. Phenobarbital reduces EEG amplitude and propagation of neonatal seizures but does not alter performance of automated seizure detection.