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Sample records for marker selection based

  1. A novel selectable marker based on Aspergillus niger arginase expression.

    PubMed

    Dave, Kashyap; Ahuja, Manmeet; Jayashri, T N; Sirola, Rekha Bisht; Punekar, Narayan S

    2012-06-10

    Selectable markers are valuable tools in transforming asexual fungi like Aspergillus niger. An arginase (agaA) expression vector and a suitable arginase-disrupted host would define a novel nutritional marker/selection for transformation. The development of such a marker was successfully achieved in two steps. The single genomic copy of A. niger arginase gene was disrupted by homologous integration of the bar marker. The agaA disruptant was subsequently complemented by transforming it with agaA expression vectors. Both citA and trpC promoters were able to drive the expression of arginase cDNA. Such agaA+ transformants displayed arginase expression pattern distinct from that of the parent strain. The results are also consistent with a single catabolic route for arginine in this fungus. A simple yet novel arginine-based selection for filamentous fungal transformation is thus described.

  2. Multi-marker-LD based genetic algorithm for tag SNP selection.

    PubMed

    Mouawad, Amer E; Mansour, Nashat

    2014-12-01

    Despite the advances in genotyping technologies which have led to large reduction in genotyping cost, the Tag SNP Selection problem remains an important problem for computational biologists and geneticists. Selecting the smallest subset of tag SNPs that can predict the other SNPs would considerably minimize the complexity of genome-wide or block-based SNP-disease association studies. These studies would lead to better diagnosis and treatment of diseases. In this work, we propose three variations of a genetic algorithm based on two-marker linkage disequilibrium, multi-marker linkage disequilibrium, and a third measure that we denote by prediction power. The performance of the three algorithms are compared with those of a recognized tag SNP selection algorithm using three different real data sets from the HapMap project. The results indicate that the multi-marker linkage disequilibrium based genetic algorithm yields better prediction accuracy.

  3. Opportunities of marker-assisted selection for rice fragrance through marker-trait association analysis of microsatellites and gene-based markers.

    PubMed

    Golestan Hashemi, F S; Rafii, M Y; Razi Ismail, M; Mohamed, M T M; Rahim, H A; Latif, M A; Aslani, F

    2015-09-01

    Developing fragrant rice through marker-assisted/aided selection (MAS) is an economical and profitable approach worldwide for the enrichment of an elite genetic background with a pleasant aroma. The PCR-based DNA markers that distinguish the alleles of major fragrance genes in rice have been synthesised to develop rice scent biofortification through MAS. Thus, the present study examined the aroma biofortification potential of these co-dominant markers in a germplasm panel of 189 F2 progeny developed from crosses between a non-aromatic variety (MR84) and a highly aromatic but low-yielding variety (MRQ74) to determine the most influential diagnostic markers for fragrance biofortification. The SSRs and functional DNA markers RM5633 (on chromosome 4), RM515, RM223, L06, NKSbad2, FMbadh2-E7, BADEX7-5, Aro7 and SCU015RM (on chromosome 8) were highly associated with the 2AP (2-acetyl-1-pyrroline) content across the population. The alleles traced via these markers were also in high linkage disequilibrium (R(2) > 0.70) and explained approximately 12.1, 27.05, 27.05, 27.05, 25.42, 25.42, 20.53, 20.43 and 20.18% of the total phenotypic variation observed for these biomarkers, respectively. F2 plants harbouring the favourable alleles of these effective markers produced higher levels of fragrance. Hence, these rice plants can be used as donor parents to increase the development of fragrance-biofortified tropical rice varieties adapted to growing conditions and consumer preferences, thus contributing to the global rice market. © 2015 German Botanical Society and The Royal Botanical Society of the Netherlands.

  4. Space-based visual attention: a marker of immature selective attention in toddlers?

    PubMed

    Rivière, James; Brisson, Julie

    2014-11-01

    Various studies suggested that attentional difficulties cause toddlers' failure in some spatial search tasks. However, attention is not a unitary construct and this study investigated two attentional mechanisms: location selection (space-based attention) and object selection (object-based attention). We investigated how toddlers' attention is distributed in the visual field during a manual search task for objects moving out of sight, namely the moving boxes task. Results show that 2.5-year-olds who failed this task allocated more attention to the location of the relevant object than to the object itself. These findings suggest that in some manual search tasks the primacy of space-based attention over object-based attention could be a marker of immature selective attention in toddlers.

  5. Counter-selectable marker for bacterial-based interaction trap systems.

    PubMed

    Meng, Xiangdong; Smith, Robin M; Giesecke, Astrid V; Joung, J Keith; Wolfe, Scot A

    2006-02-01

    Counter-selectable markers can be used in two-hybrid systems to search libraries for a protein or compound that interferes with a macromolecular interaction or to identify macromolecules from a population that cannot mediate a particular interaction. In this report, we describe the adaptation of the yeast URA3/5-FOA counter-selection system for use in bacterial interaction trap experiments. Two different URA3 reporter systems were developed that allow robust counter-selection: (i) a single copy F' episome reporter and (ii) a co-cistronic HIS3-URA3 reporter vector. The HIS3-URA3 reporter can be used for either positive or negative selections in appropriate bacterial strains. These reagents extend the utility of the bacterial two-hybrid system as an alternative to its yeast-based counterpart.

  6. A Site-Specific Recombinase-Based Method to Produce Antibiotic Selectable Marker Free Transgenic Cattle

    PubMed Central

    Tong, Qi; Liu, Xu; Su, Feng; Quan, Fusheng; Guo, Zekun; Zhang, Yong

    2013-01-01

    Antibiotic selectable marker genes have been widely used to generate transgenic animals. Once transgenic animals have been obtained, the selectable marker is no longer necessary but raises public concerns regarding biological safety. The aim of this study was to prepare competent antibiotic selectable marker free transgenic cells for somatic cell nuclear transfer (SCNT). PhiC31 intergrase was used to insert a transgene cassette into a “safe harbor” in the bovine genome. Then, Cre recombinase was employed to excise the selectable marker under the monitoring of a fluorescent double reporter. By visually tracking the phenotypic switch from red to green fluorescence, antibiotic selectable marker free cells were easily detected and sorted by fluorescence-activated cell sorting. For safety, we used phiC31 mRNA and cell-permeant Cre protein in this study. When used as donor nuclei for SCNT, these safe harbor integrated marker-free transgenic cells supported a similar developmental competence of SCNT embryos compared with that of non-transgenic cells. After embryo transfer, antibiotic selectable marker free transgenic cattle were generated and anti-bacterial recombinant human β-defensin-3 in milk was detected during their lactation period. Thus, this approach offers a rapid and safe alternative to produce antibiotic selectable marker free transgenic farm animals, thereby making it a valuable tool to promote the healthy development and welfare of transgenic farm animals. PMID:23658729

  7. A site-specific recombinase-based method to produce antibiotic selectable marker free transgenic cattle.

    PubMed

    Yu, Yuan; Wang, Yongsheng; Tong, Qi; Liu, Xu; Su, Feng; Quan, Fusheng; Guo, Zekun; Zhang, Yong

    2013-01-01

    Antibiotic selectable marker genes have been widely used to generate transgenic animals. Once transgenic animals have been obtained, the selectable marker is no longer necessary but raises public concerns regarding biological safety. The aim of this study was to prepare competent antibiotic selectable marker free transgenic cells for somatic cell nuclear transfer (SCNT). PhiC31 intergrase was used to insert a transgene cassette into a "safe harbor" in the bovine genome. Then, Cre recombinase was employed to excise the selectable marker under the monitoring of a fluorescent double reporter. By visually tracking the phenotypic switch from red to green fluorescence, antibiotic selectable marker free cells were easily detected and sorted by fluorescence-activated cell sorting. For safety, we used phiC31 mRNA and cell-permeant Cre protein in this study. When used as donor nuclei for SCNT, these safe harbor integrated marker-free transgenic cells supported a similar developmental competence of SCNT embryos compared with that of non-transgenic cells. After embryo transfer, antibiotic selectable marker free transgenic cattle were generated and anti-bacterial recombinant human β-defensin-3 in milk was detected during their lactation period. Thus, this approach offers a rapid and safe alternative to produce antibiotic selectable marker free transgenic farm animals, thereby making it a valuable tool to promote the healthy development and welfare of transgenic farm animals.

  8. Construction of a Genetic Linkage Map Based on Amplified Fragment Length Polymorphism Markers and Development of Sequence-Tagged Site Markers for Marker-Assisted Selection of the Sporeless Trait in the Oyster Mushroom (Pleurotus eryngii)

    PubMed Central

    Ueda, Jun; Obatake, Yasushi; Murakami, Shigeyuki; Fukumasa, Yukitaka; Matsumoto, Teruyuki

    2012-01-01

    A large number of spores from fruiting bodies can lead to allergic reactions and other problems during the cultivation of edible mushrooms, including Pleurotus eryngii (DC.) Quél. A cultivar harboring a sporulation-deficient (sporeless) mutation would be useful for preventing these problems, but traditional breeding requires extensive time and labor. In this study, using a sporeless P. eryngii strain, we constructed a genetic linkage map to introduce a molecular breeding program like marker-assisted selection. Based on the segregation of 294 amplified fragment length polymorphism markers, two mating type factors, and the sporeless trait, the linkage map consisted of 11 linkage groups with a total length of 837.2 centimorgans (cM). The gene region responsible for the sporeless trait was located in linkage group IX with 32 amplified fragment length polymorphism markers and the B mating type factor. We also identified eight markers closely linked (within 1.2 cM) to the sporeless locus using bulked-segregant analysis-based amplified fragment length polymorphism. One such amplified fragment length polymorphism marker was converted into two sequence-tagged site markers, SD488-I and SD488-II. Using 14 wild isolates, sequence-tagged site analysis indicated the potential usefulness of the combination of two sequence-tagged site markers in cross-breeding of the sporeless strain. It also suggested that a map constructed for P. eryngii has adequate accuracy for marker-assisted selection. PMID:22210222

  9. Genetic relationships between selected Turkish mulberry genotypes (Morus spp) based on RAPD markers.

    PubMed

    Orhan, E; Ercisli, S

    2010-11-03

    Mulberry (Morus spp, Moraceae) is an important horticultural crop in Turkey, which is one of the main world producers of mulberry fruit. We evaluated the genetic relationships among 26 mulberry genotypes selected for agronomic characteristics, using RAPD markers. A total of 367 DNA markers were generated with 34 random primers. The highest genetic similarity (0.80) was observed between Oltu58 (M. nigra) and Olur90 (M. nigra) genotypes. The genotypes Oltu3 (M. alba) and Oltu18 (M. rubra) were the most distant (0.36). We found that the RAPD technique is a useful tool to discriminate mulberry genotypes at both the intra- and interspecific level. This type of information will aid in accurate identification of useful genotypes for breeding programs.

  10. A Marker-Based Approach for the Automated Selection of a Single Segmentation from a Hierarchical Set of Image Segmentations

    NASA Technical Reports Server (NTRS)

    Tarabalka, Y.; Tilton, J. C.; Benediktsson, J. A.; Chanussot, J.

    2012-01-01

    The Hierarchical SEGmentation (HSEG) algorithm, which combines region object finding with region object clustering, has given good performances for multi- and hyperspectral image analysis. This technique produces at its output a hierarchical set of image segmentations. The automated selection of a single segmentation level is often necessary. We propose and investigate the use of automatically selected markers for this purpose. In this paper, a novel Marker-based HSEG (M-HSEG) method for spectral-spatial classification of hyperspectral images is proposed. Two classification-based approaches for automatic marker selection are adapted and compared for this purpose. Then, a novel constrained marker-based HSEG algorithm is applied, resulting in a spectral-spatial classification map. Three different implementations of the M-HSEG method are proposed and their performances in terms of classification accuracies are compared. The experimental results, presented for three hyperspectral airborne images, demonstrate that the proposed approach yields accurate segmentation and classification maps, and thus is attractive for remote sensing image analysis.

  11. Selection of high heterozygosity popcorn varieties in Brazil based on SSR markers.

    PubMed

    Eloi, I B O; Mangolin, C A; Scapim, C A; Gonçalves, C S; Machado, M F P S

    2012-07-19

    We analyzed genetic structure and diversity among eight populations of popcorn, using SSR loci as genetic markers. Our objectives were to select SSR loci that could be used to estimate genetic diversity within popcorn populations, and to analyze the genetic structure of promising populations with high levels of heterozygosity that could be used in breeding programs. Fifty-seven alleles (3.7 alleles per locus) were detected; the highest effective number of alleles (4.21) and the highest gene diversity (0.763) were found for the Umc2226 locus. A very high level of population differentiation was found (F(ST) = 0.3664), with F(ST) for each locus ranging from 0.1029 (Umc1664) to 0.6010 (Umc2350). This analysis allowed us to identify SSR loci with high levels of heterozygosity and heterozygous varieties, which could be selected for production of inbred lines and for developing new cultivars.

  12. Oral cancer prognosis based on clinicopathologic and genomic markers using a hybrid of feature selection and machine learning methods

    PubMed Central

    2013-01-01

    Background Machine learning techniques are becoming useful as an alternative approach to conventional medical diagnosis or prognosis as they are good for handling noisy and incomplete data, and significant results can be attained despite a small sample size. Traditionally, clinicians make prognostic decisions based on clinicopathologic markers. However, it is not easy for the most skilful clinician to come out with an accurate prognosis by using these markers alone. Thus, there is a need to use genomic markers to improve the accuracy of prognosis. The main aim of this research is to apply a hybrid of feature selection and machine learning methods in oral cancer prognosis based on the parameters of the correlation of clinicopathologic and genomic markers. Results In the first stage of this research, five feature selection methods have been proposed and experimented on the oral cancer prognosis dataset. In the second stage, the model with the features selected from each feature selection methods are tested on the proposed classifiers. Four types of classifiers are chosen; these are namely, ANFIS, artificial neural network, support vector machine and logistic regression. A k-fold cross-validation is implemented on all types of classifiers due to the small sample size. The hybrid model of ReliefF-GA-ANFIS with 3-input features of drink, invasion and p63 achieved the best accuracy (accuracy = 93.81%; AUC = 0.90) for the oral cancer prognosis. Conclusions The results revealed that the prognosis is superior with the presence of both clinicopathologic and genomic markers. The selected features can be investigated further to validate the potential of becoming as significant prognostic signature in the oral cancer studies. PMID:23725313

  13. [Artificial selection for cattle based on high-density SNP markers].

    PubMed

    Liu, Xi-Dong; Wang, Zhi-Peng; Fan, Hui-Zhong; Li, Jun-Ya; Gao, Hui-Jiang

    2012-10-01

    With the implementation of genetic improvement in recent years, artificial selection has greatly improved beef cattle production performance and its genetic basis has been dramatically changed. In this study, based on the Illumina BovineSNP50 (54K) and BovineHD (770K) BeadChip and the FST value, we analyzed the genetic differentiation of cattle and screened the imprints of selection in bovine genome. Finally, we found 47104 OUTLIER SNP loci and 3064 candidate genes, for example, CLIC5, TG, CACNA2D1, and FSHR etc. The biological processes and molecular functions of genes were analyzed through gene annotation.The results of this study established a genome-wide map of selection footprints in beef cattle genome and a clue for in-depth study of artificial selection and understanding of biological evolution.Our results indicate that artificial selection has played an important role in cattle breed genetic improvement.

  14. Dynamic markers based on blood perfusion fluctuations for selecting skin melanocytic lesions for biopsy

    NASA Astrophysics Data System (ADS)

    Lancaster, Gemma; Stefanovska, Aneta; Pesce, Margherita; Marco Vezzoni, Gian; Loggini, Barbara; Pingitore, Raffaele; Ghiara, Fabrizio; Barachini, Paolo; Cervadoro, Gregorio; Romanelli, Marco; Rossi, Marco

    2015-08-01

    Skin malignant melanoma is a highly angiogenic cancer, necessitating early diagnosis for positive prognosis. The current diagnostic standard of biopsy and histological examination inevitably leads to many unnecessary invasive excisions. Here, we propose a non-invasive method of identification of melanoma based on blood flow dynamics. We consider a wide frequency range from 0.005-2 Hz associated with both local vascular regulation and effects of cardiac pulsation. Combining uniquely the power of oscillations associated with individual physiological processes we obtain a marker which distinguishes between melanoma and atypical nevi with sensitivity of 100% and specificity of 90.9%. The method reveals valuable functional information about the melanoma microenvironment. It also provides the means for simple, accurate, in vivo distinction between malignant melanoma and atypical nevi, and may lead to a substantial reduction in the number of biopsies currently undertaken.

  15. Selection of bioavailability markers for herbal extracts based on in silico descriptors and their correlation to in vitro permeability.

    PubMed

    Pade, Devendra; Stavchansky, Salomon

    2008-01-01

    Bioavailability data for herbal supplements in humans is not readily available or is difficult to obtain, because of the complexity of the composition and the diversity of the constituents. Potency of an herbal extract is due to the synergistic interactions between several constituents. Thus, the use of in silico methods is an attractive alternative to predict the qualitative intestinal permeability of the active constituents for the selection of appropriate bioavailability markers. Molecular descriptors such as CLogP, minimal cross-sectional area and polar surface area of 37 active components from selected herbal extracts such as milk thistle, kava, ginkgo, ginseng, valerian, black cohosh and garlic were estimated. In vitro permeability of the compounds was determined by SimBioDAS an in vitro epithelial cell permeability assay. Based on the in silico descriptors and their relationship with the in vitro permeability, the qualitative intestinal permeability of the active compounds was predicted. Bioavailability and bioequivalence markers were predicted for kava, Ginkgo biloba and milk thistle. Choosing a compound which has the least intestinal permeability as a marker is the most conservative approach toward ensuring the bioavailability of the entire extract.

  16. Identifying Predictive Markers for Personalized Treatment Selection

    PubMed Central

    Shen, Yuanyuan

    2017-01-01

    Summary It is now well recognized that the effectiveness and potential risk of a treatment often vary by patient subgroups. Although trial-and-error and one-size-fits-all approaches to treatment selection remains a common practice, much recent focus has been placed on individualized treatment selection based on patient information (La Thangue and Kerr, 2011; Ong et al., 2012). Genetic and molecular markers are becoming increasingly available to guide treatment selection for various diseases including HIV and breast cancer (Mallal et al., 2008; Zujewski and Kamin, 2008). In recent years, many statistical procedures for developing individualized treatment rules (ITRs) have been proposed. However, less focus has been given to efficient selection of predictive biomarkers for treatment selection. The standard Wald test for interactions between treatment and the set of markers of interest may not work well when the marker effects are non-linear. Furthermore, interaction based test is scale dependent and may fail to capture markers useful for predicting individualized treatment differences. In this paper, we propose to overcome these difficulties by developing a kernel machine (KM) score test that can efficiently identify markers predictive of treatment difference. Simulation studies show that our proposed KM based score test is more powerful than the Wald test when there is non-linear effect among the predictors and when the outcome is binary with non-linear link functions. Furthermore, when there is high-correlation among predictors and when the number of predictors is not small, our method also over-performs Wald test. The proposed method is illustrated with two randomized clinical trials. PMID:26999054

  17. Evaluating markers for selecting a patient's treatment.

    PubMed

    Song, Xiao; Pepe, Margaret Sullivan

    2004-12-01

    Selecting the best treatment for a patient's disease may be facilitated by evaluating clinical characteristics or biomarker measurements at diagnosis. We consider how to evaluate the potential impact of such measurements on treatment selection algorithms. For example, magnetic resonance neurographic imaging is potentially useful for deciding whether a patient should be treated surgically for Carpal Tunnel Syndrome or should receive less-invasive conservative therapy. We propose a graphical display, the selection impact (SI) curve that shows the population response rate as a function of treatment selection criteria based on the marker. The curve can be useful for choosing a treatment policy that incorporates information on the patient's marker value exceeding a threshold. The SI curve can be estimated using data from a comparative randomized trial conducted in the population as long as treatment assignment in the trial is independent of the predictive marker. Estimating the SI curve is therefore part of a post hoc analysis to determine whether the marker identifies patients that are more likely to benefit from one treatment over another. Nonparametric and parametric estimates of the SI curve are proposed in this article. Asymptotic distribution theory is used to evaluate the relative efficiencies of the estimators. Simulation studies show that inference is straightforward with realistic sample sizes. We illustrate the SI curve and statistical inference for it with data motivated by an ongoing trial of surgery versus conservative therapy for Carpal Tunnel Syndrome.

  18. Genome-wide association study and marker-based selection for seed weight in soybean (Glycine max)

    USDA-ARS?s Scientific Manuscript database

    Seed weight (SW) is important for the yield and quality in food/vegetable uses of soybean (Glycine max), a worldwide major crop. However, our knowledge of genes controlling SW remains limited. To better understand the molecular mechanism of the trait and identify DNA markers usable in marker-based b...

  19. Improved site-specific recombinase-based method to produce selectable marker- and vector-backbone-free transgenic cells

    NASA Astrophysics Data System (ADS)

    Yu, Yuan; Tong, Qi; Li, Zhongxia; Tian, Jinhai; Wang, Yizhi; Su, Feng; Wang, Yongsheng; Liu, Jun; Zhang, Yong

    2014-02-01

    PhiC31 integrase-mediated gene delivery has been extensively used in gene therapy and animal transgenesis. However, random integration events are observed in phiC31-mediated integration in different types of mammalian cells; as a result, the efficiencies of pseudo attP site integration and evaluation of site-specific integration are compromised. To improve this system, we used an attB-TK fusion gene as a negative selection marker, thereby eliminating random integration during phiC31-mediated transfection. We also excised the selection system and plasmid bacterial backbone by using two other site-specific recombinases, Cre and Dre. Thus, we generated clean transgenic bovine fetal fibroblast cells free of selectable marker and plasmid bacterial backbone. These clean cells were used as donor nuclei for somatic cell nuclear transfer (SCNT), indicating a similar developmental competence of SCNT embryos to that of non-transgenic cells. Therefore, the present gene delivery system facilitated the development of gene therapy and agricultural biotechnology.

  20. Multiplex PCR-based simultaneous amplification of selectable marker and reporter genes for the screening of genetically modified crops.

    PubMed

    Randhawa, Gurinder Jit; Chhabra, Rashmi; Singh, Monika

    2009-06-24

    The development and commercialization of genetically modified (GM) crops with enhanced insect and herbicide resistance, abiotic stress tolerance, and improved nutritional quality has expanded dramatically. Notwithstanding the huge potential benefits of GM crops, the perceived environmental risks associated with these crops need to be addressed in proper perspective. One critical concern is the adventitious presence or unintentional mixing of GM seed in non-GM seed lots, which can seriously affect the global seed market. It would therefore be necessary though a challenging task to develop reliable, efficient, and economical assays for GM detection, identification, and quantification in non-GM seed lots. This can be systematically undertaken by preliminary screening for control elements and selectable or scorable (reporter) marker genes. In this study, simplex and multiplex polymerase chain reaction (PCR) assays individually as well as simultaneously amplifying the commonly used selectable marker genes, i.e., aadA, bar, hpt, nptII, pat encoding, respectively, for aminoglycoside-3'-adenyltransferase, Streptococcus viridochromogenes phosphinothricin-N-acetyltransferase, hygromycin phosphotransferase, neomycin phosphotransferase, Streptococcus hygroscopicus phosphinothricin-N-acetyltransferase, and a reporter gene uidA encoding beta-d-glucuronidase, were developed as a reliable tool for qualitative screening of GM crops. The efficiency of the assays was also standardized in the test samples prepared by artificial mixing of transgenic seed samples in different proportions. The developed multiplex PCR assays will be useful in verifying the GM status of a sample irrespective of the crop and GM trait.

  1. Marker-assisted backcross selection in an interspecific Cucumis population broadens the genetic base of cucumber (Cucumis sativus L.)

    USDA-ARS?s Scientific Manuscript database

    Cucumber (Cucumis sativus L.) is a major cucurbit vegetable species whose genetic base has been drastically reduced during its domestication. The crop's narrow genetic base (3-12% DNA polymorphism) has resulted from the use of limited genetic material and intense selection during plant improvement....

  2. Screening for recombinants of Crambe abyssynica after transformation by the pMF1 marker-free vector based on chemical selection and meristematic regeneration.

    PubMed

    Qi, Weicong; Tinnenbroek-Capel, Iris E M; Salentijn, Elma M J; Schaart, Jan G; Cheng, Jihua; Denneboom, Christel; Zhang, Zhao; Zhang, Xiaolin; Zhao, Han; Visser, Richard G F; Huang, Bangquan; Van Loo, Eibertus N; Krens, Frans A

    2015-09-11

    The T-DNA region of pMF1 vector of marker-free system developed by Wageningen UR, has Recombinase R-LBD gene fusion and nptII and codA gene fusion between two recombination sites. After transformation applying dexamethasone (DEX) can activate the recombinase to remove the T-DNA fragment between recombination sites. The recombinant ought to be selected on 5-fluorocytocine (5-FC) because of codA converting 5-FC into 5-fluorouracil the toxic. A PMF1 vector was transformed into hexaploid species Crambe abyssinica. Two independent transformants were chosen for DEX-induced recombination and later 5-FC selection. In contrast to earlier pMF1 experiments, the strategy of stepwise selection based on meristematic regeneration was engaged. After a long period of 5-FC selection, recombinants were obtained successfully, but most of the survivors were wildtype and non-recombinant. The results revealed when applying the PMF1 marker-free system on C. abyssinica, 1) Increasing in the DEX concentration did not correspondingly enhance the success of recombination; 2) both of the DEX-induced recombination and 5-FC negative selection were apparently insufficient which was leading to the extremely high frequency in chimerism occurring for recombinant and non-recombinant cells in tissues; 3) the strategy of stepwise selection based on meristem tissue regeneration was crucial for successfully isolating the recombinant germplasm from the chimera.

  3. Screening for recombinants of Crambe abyssynica after transformation by the pMF1 marker-free vector based on chemical selection and meristematic regeneration

    PubMed Central

    Qi, Weicong; Tinnenbroek-Capel, Iris E. M.; Salentijn, Elma M. J.; Schaart, Jan G.; Cheng, Jihua; Denneboom, Christel; Zhang, Zhao; Zhang, Xiaolin; Zhao, Han; Visser, Richard G. F.; Huang, Bangquan; Van Loo, Eibertus N.; Krens, Frans A.

    2015-01-01

    The T-DNA region of pMF1 vector of marker-free system developed by Wageningen UR, has Recombinase R-LBD gene fusion and nptII and codA gene fusion between two recombination sites. After transformation applying dexamethasone (DEX) can activate the recombinase to remove the T-DNA fragment between recombination sites. The recombinant ought to be selected on 5-fluorocytocine (5-FC) because of codA converting 5-FC into 5-fluorouracil the toxic. A PMF1 vector was transformed into hexaploid species Crambe abyssinica. Two independent transformants were chosen for DEX-induced recombination and later 5-FC selection. In contrast to earlier pMF1 experiments, the strategy of stepwise selection based on meristematic regeneration was engaged. After a long period of 5-FC selection, recombinants were obtained successfully, but most of the survivors were wildtype and non-recombinant. The results revealed when applying the PMF1 marker-free system on C. abyssinica, 1) Increasing in the DEX concentration did not correspondingly enhance the success of recombination; 2) both of the DEX-induced recombination and 5-FC negative selection were apparently insufficient which was leading to the extremely high frequency in chimerism occurring for recombinant and non-recombinant cells in tissues; 3) the strategy of stepwise selection based on meristem tissue regeneration was crucial for successfully isolating the recombinant germplasm from the chimera. PMID:26358007

  4. Applying antibiotic selection markers for nematode genetics.

    PubMed

    Cornes, Eric; Quéré, Cécile A L; Giordano-Santini, Rosina; Dupuy, Denis

    2014-08-01

    Antibiotic selection markers have been recently developed in the multicellular model organism Caenorhabditis elegans and other related nematode species, opening great opportunities in the field of nematode transgenesis. Here we describe how these antibiotic selection systems can be easily combined with many well-established genetic approaches to study gene function, improving time- and cost-effectiveness of the nematode genetic toolbox. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Potential of marker selection to increase prediction accuracy of genomic selection in soybean (Glycine max L.).

    PubMed

    Ma, Yansong; Reif, Jochen C; Jiang, Yong; Wen, Zixiang; Wang, Dechun; Liu, Zhangxiong; Guo, Yong; Wei, Shuhong; Wang, Shuming; Yang, Chunming; Wang, Huicai; Yang, Chunyan; Lu, Weiguo; Xu, Ran; Zhou, Rong; Wang, Ruizhen; Sun, Zudong; Chen, Huaizhu; Zhang, Wanhai; Wu, Jian; Hu, Guohua; Liu, Chunyan; Luan, Xiaoyan; Fu, Yashu; Guo, Tai; Han, Tianfu; Zhang, Mengchen; Sun, Bincheng; Zhang, Lei; Chen, Weiyuan; Wu, Cunxiang; Sun, Shi; Yuan, Baojun; Zhou, Xinan; Han, Dezhi; Yan, Hongrui; Li, Wenbin; Qiu, Lijuan

    Genomic selection is a promising molecular breeding strategy enhancing genetic gain per unit time. The objectives of our study were to (1) explore the prediction accuracy of genomic selection for plant height and yield per plant in soybean [Glycine max (L.) Merr.], (2) discuss the relationship between prediction accuracy and numbers of markers, and (3) evaluate the effect of marker preselection based on different methods on the prediction accuracy. Our study is based on a population of 235 soybean varieties which were evaluated for plant height and yield per plant at multiple locations and genotyped by 5361 single nucleotide polymorphism markers. We applied ridge regression best linear unbiased prediction coupled with fivefold cross-validations and evaluated three strategies of marker preselection. For plant height, marker density and marker preselection procedure impacted prediction accuracy only marginally. In contrast, for grain yield, prediction accuracy based on markers selected with a haplotype block analyses-based approach increased by approximately 4 % compared with random or equidistant marker sampling. Thus, applying marker preselection based on haplotype blocks is an interesting option for a cost-efficient implementation of genomic selection for grain yield in soybean breeding.

  6. A high-density SNP genotyping array for Brassica napus and its ancestral diploid species based on optimised selection of single-locus markers in the allotetraploid genome.

    PubMed

    Clarke, Wayne E; Higgins, Erin E; Plieske, Joerg; Wieseke, Ralf; Sidebottom, Christine; Khedikar, Yogendra; Batley, Jacqueline; Edwards, Dave; Meng, Jinling; Li, Ruiyuan; Lawley, Cynthia Taylor; Pauquet, Jérôme; Laga, Benjamin; Cheung, Wing; Iniguez-Luy, Federico; Dyrszka, Emmanuelle; Rae, Stephen; Stich, Benjamin; Snowdon, Rod J; Sharpe, Andrew G; Ganal, Martin W; Parkin, Isobel A P

    2016-10-01

    The Brassica napus Illumina array provides genome-wide markers linked to the available genome sequence, a significant tool for genetic analyses of the allotetraploid B. napus and its progenitor diploid genomes. A high-density single nucleotide polymorphism (SNP) Illumina Infinium array, containing 52,157 markers, was developed for the allotetraploid Brassica napus. A stringent selection process employing the short probe sequence for each SNP assay was used to limit the majority of the selected markers to those represented a minimum number of times across the highly replicated genome. As a result approximately 60 % of the SNP assays display genome-specificity, resolving as three clearly separated clusters (AA, AB, and BB) when tested with a diverse range of B. napus material. This genome specificity was supported by the analysis of the diploid ancestors of B. napus, whereby 26,504 and 29,720 markers were scorable in B. oleracea and B. rapa, respectively. Forty-four percent of the assayed loci on the array were genetically mapped in a single doubled-haploid B. napus population allowing alignment of their physical and genetic coordinates. Although strong conservation of the two positions was shown, at least 3 % of the loci were genetically mapped to a homoeologous position compared to their presumed physical position in the respective genome, underlying the importance of genetic corroboration of locus identity. In addition, the alignments identified multiple rearrangements between the diploid and tetraploid Brassica genomes. Although mostly attributed to genome assembly errors, some are likely evidence of rearrangements that occurred since the hybridisation of the progenitor genomes in the B. napus nucleus. Based on estimates for linkage disequilibrium decay, the array is a valuable tool for genetic fine mapping and genome-wide association studies in B. napus and its progenitor genomes.

  7. A pyramid breeding of eight grain-yield related quantitative trait loci based on marker-assistant and phenotype selection in rice (Oryza sativa L.).

    PubMed

    Zong, Guo; Wang, Ahong; Wang, Lu; Liang, Guohua; Gu, Minghong; Sang, Tao; Han, Bin

    2012-07-20

    1000-Grain weight and spikelet number per panicle are two important components for rice grain yield. In our previous study, eight quantitative trait loci (QTLs) conferring spikelet number per panicle and 1000-grain weight were mapped through sequencing-based genotyping of 150 rice recombinant inbred lines (RILs). In this study, we validated the effects of four QTLs from Nipponbare using chromosome segment substitution lines (CSSLs), and pyramided eight grain yield related QTLs. The new lines containing the eight QTLs with positive effects showed increased panicle and spikelet size as compared with the parent variety 93-11. We further proposed a novel pyramid breeding scheme based on marker-assistant and phenotype selection (MAPS). This scheme allowed pyramiding of as many as 24 QTLs at a single hybridization without massive cross work. This study provided insights into the molecular basis of rice grain yield for direct wealth for high-yielding rice breeding. Copyright © 2012. Published by Elsevier Ltd.

  8. Application of a high-speed breeding technology to apple (Malus × domestica) based on transgenic early flowering plants and marker-assisted selection.

    PubMed

    Flachowsky, Henryk; Le Roux, Pierre-Marie; Peil, Andreas; Patocchi, Andrea; Richter, Klaus; Hanke, Magda-Viola

    2011-10-01

    Breeding of apple (Malus × domestica) remains a slow process because of protracted generation cycles. Shortening the juvenile phase to achieve the introgression of traits from wild species into prebreeding material within a reasonable time frame is a great challenge. In this study, we evaluated early flowering transgenic apple lines overexpressing the BpMADS4 gene of silver birch with regard to tree morphology in glasshouse conditions. Based on the results obtained, line T1190 was selected for further analysis and application to fast breeding. The DNA sequences flanking the T-DNA were isolated and the T-DNA integration site was mapped on linkage group 4. The inheritance and correctness of the T-DNA integration were confirmed after meiosis. A crossbred breeding programme was initiated by crossing T1190 with the fire blight-resistant wild species Malus fusca. Transgenic early flowering F(1) seedlings were selected and backcrossed with 'Regia' and 98/6-10 in order to introgress the apple scab Rvi2, Rvi4 and powdery mildew Pl-1, Pl-2 resistance genes and the fire blight resistance quantitative trait locus FB-F7 present in 'Regia'. Three transgenic BC'1 seedlings pyramiding Rvi2, Rvi4 and FB-F7, as well as three other BC'1 seedlings combining Pl-1 and Pl-2, were identified. Thus, the first transgenic early flowering-based apple breeding programme combined with marker-assisted selection was established. © 2011 The Authors. New Phytologist © 2011 New Phytologist Trust.

  9. Alanine Racemase Mutants of Burkholderia pseudomallei and Burkholderia mallei and Use of Alanine Racemase as a Non-Antibiotic-Based Selectable Marker

    PubMed Central

    Zajdowicz, Sheryl L. W.; Jones-Carson, Jessica; Vazquez-Torres, Andres; Jobling, Michael G.; Gill, Ronald E.; Holmes, Randall K.

    2011-01-01

    Burkholderia pseudomallei and Burkholderia mallei are category B select agents and must be studied under BSL3 containment in the United States. They are typically resistant to multiple antibiotics, and the antibiotics used to treat B. pseudomallei or B. mallei infections may not be used as selective agents with the corresponding Burkholderia species. Here, we investigated alanine racemase deficient mutants of B. pseudomallei and B. mallei for development of non-antibiotic-based genetic selection methods and for attenuation of virulence. The genome of B. pseudomallei K96243 has two annotated alanine racemase genes (bpsl2179 and bpss0711), and B. mallei ATCC 23344 has one (bma1575). Each of these genes encodes a functional enzyme that can complement the alanine racemase deficiency of Escherichia coli strain ALA1. Herein, we show that B. pseudomallei with in-frame deletions in both bpsl2179 and bpss0711, or B. mallei with an in-frame deletion in bma1575, requires exogenous d-alanine for growth. Introduction of bpsl2179 on a multicopy plasmid into alanine racemase deficient variants of either Burkholderia species eliminated the requirement for d-alanine. During log phase growth without d-alanine, the viable counts of alanine racemase deficient mutants of B. pseudomallei and B. mallei decreased within 2 hours by about 1000-fold and 10-fold, respectively, and no viable bacteria were present at 24 hours. We constructed several genetic tools with bpsl2179 as a selectable genetic marker, and we used them without any antibiotic selection to construct an in-frame ΔflgK mutant in the alanine racemase deficient variant of B. pseudomallei K96243. In murine peritoneal macrophages, wild type B. mallei ATCC 23344 was killed much more rapidly than wild type B. pseudomallei K96243. In addition, the alanine racemase deficient mutant of B. pseudomallei K96243 exhibited attenuation versus its isogenic parental strain with respect to growth and survival in murine peritoneal macrophages

  10. Alanine racemase mutants of Burkholderia pseudomallei and Burkholderia mallei and use of alanine racemase as a non-antibiotic-based selectable marker.

    PubMed

    Zajdowicz, Sheryl L W; Jones-Carson, Jessica; Vazquez-Torres, Andres; Jobling, Michael G; Gill, Ronald E; Holmes, Randall K

    2011-01-01

    Burkholderia pseudomallei and Burkholderia mallei are category B select agents and must be studied under BSL3 containment in the United States. They are typically resistant to multiple antibiotics, and the antibiotics used to treat B. pseudomallei or B. mallei infections may not be used as selective agents with the corresponding Burkholderia species. Here, we investigated alanine racemase deficient mutants of B. pseudomallei and B. mallei for development of non-antibiotic-based genetic selection methods and for attenuation of virulence. The genome of B. pseudomallei K96243 has two annotated alanine racemase genes (bpsl2179 and bpss0711), and B. mallei ATCC 23344 has one (bma1575). Each of these genes encodes a functional enzyme that can complement the alanine racemase deficiency of Escherichia coli strain ALA1. Herein, we show that B. pseudomallei with in-frame deletions in both bpsl2179 and bpss0711, or B. mallei with an in-frame deletion in bma1575, requires exogenous D-alanine for growth. Introduction of bpsl2179 on a multicopy plasmid into alanine racemase deficient variants of either Burkholderia species eliminated the requirement for D-alanine. During log phase growth without D-alanine, the viable counts of alanine racemase deficient mutants of B. pseudomallei and B. mallei decreased within 2 hours by about 1000-fold and 10-fold, respectively, and no viable bacteria were present at 24 hours. We constructed several genetic tools with bpsl2179 as a selectable genetic marker, and we used them without any antibiotic selection to construct an in-frame ΔflgK mutant in the alanine racemase deficient variant of B. pseudomallei K96243. In murine peritoneal macrophages, wild type B. mallei ATCC 23344 was killed much more rapidly than wild type B. pseudomallei K96243. In addition, the alanine racemase deficient mutant of B. pseudomallei K96243 exhibited attenuation versus its isogenic parental strain with respect to growth and survival in murine peritoneal macrophages.

  11. Imputation of unordered markers and the impact on genomic selection accuracy

    USDA-ARS?s Scientific Manuscript database

    Genomic selection, a breeding method that promises to accelerate rates of genetic gain, requires dense, genome-wide marker data. Sequence-based genotyping methods can generate de novo large numbers of markers. However, without a reference genome, these markers are unordered and typically have a lar...

  12. Development of a codominant PCR-based marker for allelic selection of the pink trait in onions (Allium cepa), based on the insertion mutation in the promoter of the anthocyanidin synthase gene.

    PubMed

    Kim, Sunggil; Yoo, Kil-Sun; Pike, Leonard M

    2005-02-01

    Bulb color in onions (Allium cepa) is an important trait and is inherited in a complex manner. However, the mechanism of color inheritance is poorly understood at the molecular level. A previous study showed that pink bulb color in onions is inherited as a single recessive trait. This trait is attributable to a significantly reduced transcription of the anthocyanidin synthase (ANS) gene. In this study, we developed a PCR-based marker for an allelic selection of the ANS gene to avoid the laborious progeny tests traditionally employed. To identify polymorphisms between pink and red alleles of the ANS gene, promoter sequences of both alleles were isolated. There was 97% nucleotide sequence identity between the promoter sequences of the two alleles. A 390-bp insertion was identified 632 bp upstream from the putative transcription start site in the pink allele. A pair of primers was designed on the flanking sequences of the inserted region and utilized as a PCR-based marker for allelic selection of the ANS gene. The reliability of the marker was tested using parents, F1 hybrids, and F3 lines whose genotypes had been identified by progeny tests. The marker was also used to evaluate the distribution of the pink allele in white and yellow breeding lines. The results indicated that a majority of the breeding lines tested were homozygous recessive.

  13. Development of a co-dominant, PCR-based marker for allelic selection of the pink trait in onions (Allium cepa), based on the insertion mutation in the promoter of the anthocyanidin synthase gene.

    PubMed

    Kim, Sunggil; Yoo, Kil-Sun; Pike, Leonard M

    2005-02-01

    Bulb color in onions (Allium cepa) is an important trait and is inherited in a complex manner. However, the mechanism of color inheritance is poorly understood at the molecular level. A previous study showed that pink bulb color in onions is inherited as a single recessive trait. This trait is attributable to a significantly reduced transcription of the anthocyanidin synthase (ANS) gene. In this study, we developed a PCR-based marker for an allelic selection of the ANS gene to avoid the laborious progeny tests traditionally employed. To identify polymorphisms between pink and red alleles of the ANS gene, promoter sequences of both alleles were isolated. There was 97% nucleotide sequence identity between the promoter sequences of the two alleles. A 390-bp insertion was identified 632 bp upstream from the putative transcription start site in the pink allele. A pair of primers was designed on the flanking sequences of the inserted region and utilized as a PCR-based marker for allelic selection of the ANS gene. The reliability of the marker was tested using parents, F1 hybrids, and F3 lines whose genotypes had been identified by progeny tests. The marker was also used to evaluate the distribution of the pink allele in white and yellow breeding lines. The results indicated that a majority of the breeding lines tested were homozygous recessive.

  14. Swapped green algal promoters: aphVIII-based gene constructs with Chlamydomonas flanking sequences work as dominant selectable markers in Volvox and vice versa.

    PubMed

    Hallmann, A; Wodniok, S

    2006-06-01

    Production of transgenic organisms is a well-established, versatile course of action in molecular biology. Genetic engineering often requires heterologous, dominant antibiotic resistance genes that have been used as selectable markers in many species. However, as heterologous 5' and 3' flanking sequences often result in very low expression rates, endogenous flanking sequences, especially promoters, are mostly required and are easily obtained in model organisms, but it is much more complicated and time-consuming to get appropriate sequences from less common organisms. In this paper, we show that aminoglycoside 3'-phosphotransferase gene (aphVIII) based constructs with 3' and 5' untranslated flanking sequences (including promoters) from the multicellular green alga Volvox work in the unicellular green alga Chlamydomonas and flanking sequences from Chlamydomonas work in Volvox, at least if a low expression rate is compensated by an enforced high gene dosage. This strategy might be useful for all investigators that intend to transform species in which genomic sequences are not available, but sequences from related organisms exist.

  15. Considerations for marker-assisted selection in peanut

    USDA-ARS?s Scientific Manuscript database

    Marker-assisted selection (MAS) offers considerable promise but requires careful planning. Among the first DNA markers used for peanut improvement were wild species-derived alleles for nematode resistance, now being combined with the high-oleic trait. These are screened as qualitative traits. Thes...

  16. Considerations for Marker-assisted selection in peanut

    USDA-ARS?s Scientific Manuscript database

    Marker-assisted selection (MAS) offers considerable promise but requires careful planning. Among the first DNA markers used for peanut improvement were wild species-derived alleles for nematode resistance, now being combined with the high-oleic trait. These are screened as qualitative traits. These ...

  17. Consensus mapping and identification of markers for marker-assisted selection of Wsm2 in wheat

    USDA-ARS?s Scientific Manuscript database

    A recently identified Wheat streak mosaic virus (WSMV) resistance gene Wsm2 confers a high level of resistance. Objective of this study was to identify closely linked DNA markers for Wsm2 for use in marker-assisted selection (MAS) in wheat. Two segregating populations (CO960293-2/’TAM 111’ and CO96...

  18. Selectable marker genes and unintended changes to the plant transcriptome.

    PubMed

    Miki, Brian; Abdeen, Ashraf; Manabe, Yuzuki; MacDonald, Phil

    2009-04-01

    The intended effect of a selectable marker gene is to confer a novel trait that allows for the selection and recovery of transgenic plants. Unintended effects may also occur as a result of interactions between the selectable marker gene or its regulatory elements and genetic elements at the site of insertion. These are called position effects. Other unintended effects may occur if the selectable marker gene has a range of pleiotropic effects related to the functional and regulatory domains within the coding region or the regulatory elements used to drive expression. Both pleiotropic and position effects may generate unpredictable events depending on the process used for transgenesis and the state of knowledge associated with the selectable marker gene. Although some selectable marker genes, such as the neomycin phosphotransferase type II gene (nptII), have no pleiotropic effects on the transcriptomes of transgenic plants, others, such as the bialaphos resistance gene (bar), have pleiotropic effects. These must be clearly understood and accounted for when evaluating the expression patterns conferred by other co-transforming transgenes under study. The number and kinds of selectable marker genes are large. A detailed understanding of their unintended effects is needed to develop transgenic strategies that will minimize or eliminate unintended and unpredictable changes to plants with newly inserted genes.

  19. Algorithms for selecting informative marker panels for population assignment.

    PubMed

    Rosenberg, Noah A

    2005-11-01

    Given a set of potential source populations, genotypes of an individual of unknown origin at a collection of markers can be used to predict the correct source population of the individual. For improved efficiency, informative markers can be chosen from a larger set of markers to maximize the accuracy of this prediction. However, selecting the loci that are individually most informative does not necessarily produce the optimal panel. Here, using genotypes from eight species--carp, cat, chicken, dog, fly, grayling, human, and maize--this univariate accumulation procedure is compared to new multivariate "greedy" and "maximin" algorithms for choosing marker panels. The procedures generally suggest similar panels, although the greedy method often recommends inclusion of loci that are not chosen by the other algorithms. In seven of the eight species, when applied to five or more markers, all methods achieve at least 94% assignment accuracy on simulated individuals, with one species--dog--producing this level of accuracy with only three markers, and the eighth species--human--requiring approximately 13-16 markers. The new algorithms produce substantial improvements over use of randomly selected markers; where differences among the methods are noticeable, the greedy algorithm leads to slightly higher probabilities of correct assignment. Although none of the approaches necessarily chooses the panel with optimal performance, the algorithms all likely select panels with performance near enough to the maximum that they all are suitable for practical use.

  20. Food selection based on total antioxidant capacity can modify antioxidant intake, systemic inflammation, and liver function without altering markers of oxidative stress.

    PubMed

    Valtueña, Silvia; Pellegrini, Nicoletta; Franzini, Laura; Bianchi, Marta A; Ardigò, Diego; Del Rio, Daniele; Piatti, PierMarco; Scazzina, Francesca; Zavaroni, Ivana; Brighenti, Furio

    2008-05-01

    It is unknown whether diets with a high dietary total antioxidant capacity (TAC) can modify oxidative stress, low-grade inflammation, or liver dysfunction, all of which are risk factors for type 2 diabetes and cardiovascular disease. We studied the effect of high- and low-TAC (HT and LT, respectively) diets on markers of antioxidant status, systemic inflammation, and liver dysfunction. In a crossover intervention, 33 healthy adults (19 men, 14 women) received the HT and LT diets for 2 wk each. Dietary habits were checked with a 3-d food record during both diet periods and the washout period. Fruit and vegetable, macronutrient, dietary fiber, and alcohol intakes did not differ significantly between the 2 diets, whereas dietary TAC, alpha-tocopherol, and ascorbic acid were significantly (P < 0.001) higher during the HT diet. Plasma alpha-tocopherol rose during the HT and decreased during the LT diet (P < 0.02 for difference) without changes in markers of oxidative stress except plasma malondialdehyde, which decreased unexpectedly during the LT diet (P < 0.05). Plasma high-sensitivity C-reactive protein, alanine aminotransferase, gamma-glutamyltranspeptidase, and alkaline phosphatase concentrations decreased during the HT compared with the LT diet (mean +/- SEM for pre-post changes: -0.72 +/- 0.37 compared with 1.05 +/- 0.60 mg/L, P < 0.01; -1.73 +/- 1.02 compared with 2.33 +/- 2.58 U/L, P < 0.01; -2.12 +/- 1.45 compared with 5.15 +/- 2.98 U/L, P < 0.05; and 1.36 +/- 1.34 compared with 5.06 +/- 2.00 U/L, P < 0.01, respectively). Selecting foods according to their TAC markedly affects antioxidant intake and modulates hepatic contribution to systemic inflammation without affecting traditional markers of antioxidant status.

  1. Plant phosphomannose isomerase as a selectable marker for rice transformation

    PubMed Central

    Hu, Lei; Li, Hao; Qin, Ruiying; Xu, Rongfang; Li, Juan; Li, Li; Wei, Pengcheng; Yang, Jianbo

    2016-01-01

    The E. coli phosphomannose isomerase (EcPMI) gene is widely used as a selectable marker gene (SMG) in mannose (Man) selection-based plant transformation. Although some plant species exhibit significant PMI activity and active PMIs were even identified in Man-sensitive plants, whether plant PMIs can be used as SMGs remains unclear. In this study, we isolated four novel PMI genes from Chlorella variabilis and Oryza sativa. Their isoenzymatic activities were examined in vitro and compared with that of EcPMI. The active plant PMIs were separately constructed into binary vectors as SMGs and then transformed into rice via Agrobacterium. In both Indica and Japonica subspecies, our results indicated that the plant PMIs could select and produce transgenic plants in a pattern similar to that of EcPMI. The transgenic plants exhibited an accumulation of plant PMI transcripts and enhancement of the in vivo PMI activity. Furthermore, a gene of interest was successfully transformed into rice using the plant PMIs as SMGs. Thus, novel SMGs for Man selection were isolated from plants, and our analysis suggested that PMIs encoding active enzymes might be common in plants and could potentially be used as appropriate genetic elements in cisgenesis engineering. PMID:27174847

  2. A Tet-on and Cre-loxP Based Genetic Engineering System for Convenient Recycling of Selection Markers in Penicillium oxalicum.

    PubMed

    Jiang, Baojie; Zhang, Ruiqin; Feng, Dan; Wang, Fangzhong; Liu, Kuimei; Jiang, Yi; Niu, Kangle; Yuan, Quanquan; Wang, Mingyu; Wang, Hailong; Zhang, Youming; Fang, Xu

    2016-01-01

    The lack of selective markers has been a key problem preventing multistep genetic engineering in filamentous fungi, particularly for industrial species such as the lignocellulose degrading Penicillium oxalicum JUA10-1(formerly named as Penicillium decumbens). To resolve this problem, we constructed a genetic manipulation system taking advantage of two established genetic systems: the Cre-loxP system and Tet-on system in P. oxalicum JUA10-1. This system is efficient and convenient. The expression of Cre recombinase was activated by doxycycline since it was controlled by Tet-on system. Using this system, two genes, ligD and bglI, were sequentially disrupted by loxP flanked ptrA. The successful application of this procedure will provide a useful tool for genetic engineering in filamentous fungi. This system will also play an important role in improving the productivity of interesting products and minimizing by-product when fermented by filamentous fungi.

  3. A Tet-on and Cre-loxP Based Genetic Engineering System for Convenient Recycling of Selection Markers in Penicillium oxalicum

    PubMed Central

    Jiang, Baojie; Zhang, Ruiqin; Feng, Dan; Wang, Fangzhong; Liu, Kuimei; Jiang, Yi; Niu, Kangle; Yuan, Quanquan; Wang, Mingyu; Wang, Hailong; Zhang, Youming; Fang, Xu

    2016-01-01

    The lack of selective markers has been a key problem preventing multistep genetic engineering in filamentous fungi, particularly for industrial species such as the lignocellulose degrading Penicillium oxalicum JUA10-1(formerly named as Penicillium decumbens). To resolve this problem, we constructed a genetic manipulation system taking advantage of two established genetic systems: the Cre-loxP system and Tet-on system in P. oxalicum JUA10-1. This system is efficient and convenient. The expression of Cre recombinase was activated by doxycycline since it was controlled by Tet-on system. Using this system, two genes, ligD and bglI, were sequentially disrupted by loxP flanked ptrA. The successful application of this procedure will provide a useful tool for genetic engineering in filamentous fungi. This system will also play an important role in improving the productivity of interesting products and minimizing by-product when fermented by filamentous fungi. PMID:27148179

  4. The accuracy of marker-assisted selection for quantitative traits within populations in linkage equilibrium.

    PubMed Central

    Ollivier, L

    1998-01-01

    Using the concept of conditional coancestry, given observed markers, an explicit expression of the accuracy of marker-based selection is derived in situations of linkage equilibrium between markers and quantitative trait loci (QTL), for the general case of full-sib families nested within half-sib families. Such a selection scheme is rather inaccurate for moderate values of family sizes and QTL variance, and the accuracies predicted for linkage disequilibrium can never be reached. The result is used to predict the accuracy of marker-assisted combined selection (MACS) and is shown to agree with previous MACS results obtained by simulation of a best linear unbiased prediction animal model. Low gains in accuracy are generally to be expected compared to standard combined selection. The maximum gain, assuming infinite family size and all QTLs marked, is about 50%. PMID:9539449

  5. Marker selection by Akaike information criterion and Bayesian information criterion.

    PubMed

    Li, W; Nyholt, D R

    2001-01-01

    We carried out a discriminant analysis with identity by descent (IBD) at each marker as inputs, and the sib pair type (affected-affected versus affected-unaffected) as the output. Using simple logistic regression for this discriminant analysis, we illustrate the importance of comparing models with different number of parameters. Such model comparisons are best carried out using either the Akaike information criterion (AIC) or the Bayesian information criterion (BIC). When AIC (or BIC) stepwise variable selection was applied to the German Asthma data set, a group of markers were selected which provide the best fit to the data (assuming an additive effect). Interestingly, these 25-26 markers were not identical to those with the highest (in magnitude) single-locus lod scores.

  6. Acceleration of peanut breeding programs by molecular marker assisted selection

    USDA-ARS?s Scientific Manuscript database

    Peanut breeding has played a significant role in yield increases and disease control. Conventional breeding focuses on field selection and phenotypic analysis and it typically takes 12-15 years before a new cultivar can be released. Molecular markers developed from sequencing data can be of great ...

  7. Polymorphism among EST-based markers in tomato

    USDA-ARS?s Scientific Manuscript database

    Cultivated tomato (Lycopersicon esculentum Mill.) has a narrow genetic base. This is in part due to population genetic processes such as founder events, genetic bottlenecks, and natural and artificial selection during domestication. We characterize the nucleotide polymorphism in 26 EST-based markers...

  8. Within-family marker-assisted selection for aquaculture species

    PubMed Central

    Sonesson, Anna K

    2007-01-01

    A within-family marker-assisted selection scheme was designed for typical aquaculture breeding schemes, where most traits are recorded on sibs of the candidates. Here, sibs of candidates were tested for the trait and genotyped to establish genetic marker effects on the trait. BLUP breeding values were calculated, including information of the markers (MAS) or not (NONMAS). These breeding values were identical for all family members in the NONMAS schemes, but differed between family members in the MAS schemes, making within-family selection possible. MAS had up to twice the total genetic gain of the corresponding NONMAS scheme. MAS was somewhat less effective when heritability increased from 0.06 to 0.12 or when the frequency of the positive allele was < 0.5. The relative efficiency of MAS was higher for schemes with more candidates, because of larger fullsib family sizes. MAS was also more efficient when male:female mating ratio changed from 1:1 to 1:5 or when the QTL explained more of the total genetic variation. Four instead of two markers linked to the QTL increased genetic gain somewhat. There was no significant difference in polygenic genetic gain between MAS and NONMAS for most schemes. The rates of inbreeding were lower for MAS than NON-MAS schemes, because fewer full-sibs were selected by MAS. PMID:17433243

  9. Metabolic Syndrome and Selective Inflammatory Markers in Psoriatic Patients

    PubMed Central

    Vachatova, Simona; Andrys, Ctirad; Salavec, Miloslav; Ettler, Karel; Rehacek, Vit; Cermakova, Eva; Malkova, Andrea

    2016-01-01

    The presented article studies the role of selected inflammatory and anti-inflammatory serum markers of psoriatic patients in the pathogenesis of metabolic syndrome (MS) and psoriasis. The study is based on the comparison between the group of psoriatic patients (74) and the control group (65). We found significantly higher BMI (p < 0.05) and diastolic blood pressure (p < 0.05) in the psoriatic patients. The values of waist circumference and BMI were significantly higher (p < 0.05) in the male patients compared to the men in the control group. The analysis revealed significantly higher CRP (p < 0.001), Lp-PLA2 (p < 0.001), leptin (p < 0.01), and resistin (p < 0.01) levels in the psoriatic patients. Significantly higher levels of CRP (p < 0.01), Lp-PLA2 (p < 0.001), leptin (p < 0.01), and resistin (p < 0.05) were found in the patients with MS compared to the controls with MS. The level of adiponectin was significantly lower (p < 0.01) in the patients with MS. Finally, we found significantly higher level of Lp-PLA2 (p < 0.001) in the group of patients without MS compared to the controls without MS. In conclusion, observed inflammatory and anti-inflammatory markers (CRP, adiponectin, leptin, resistin, and Lp-PLA2) are involved in both pathogenesis of MS and pathogenesis of psoriasis. The level of Lp-PLA2 indicates the presence of subclinical atherosclerosis (cardiovascular risk) in psoriatic patients. PMID:28097156

  10. Marker-Based Estimates Reveal Significant Nonadditive Effects in Clonally Propagated Cassava (Manihot esculenta): Implications for the Prediction of Total Genetic Value and the Selection of Varieties

    PubMed Central

    Wolfe, Marnin D.; Kulakow, Peter; Rabbi, Ismail Y.; Jannink, Jean-Luc

    2016-01-01

    In clonally propagated crops, nonadditive genetic effects can be effectively exploited by the identification of superior genetic individuals as varieties. Cassava (Manihot esculenta Crantz) is a clonally propagated staple food crop that feeds hundreds of millions. We quantified the amount and nature of nonadditive genetic variation for three key traits in a breeding population of cassava from sub-Saharan Africa using additive and nonadditive genome-wide marker-based relationship matrices. We then assessed the accuracy of genomic prediction for total (additive plus nonadditive) genetic value. We confirmed previous findings based on diallel crosses that nonadditive genetic variation is significant for key cassava traits. Specifically, we found that dominance is particularly important for root yield and epistasis contributes strongly to variation in cassava mosaic disease (CMD) resistance. Further, we showed that total genetic value predicted observed phenotypes more accurately than additive only models for root yield but not for dry matter content, which is mostly additive or for CMD resistance, which has high narrow-sense heritability. We address the implication of these results for cassava breeding and put our work in the context of previous results in cassava, and other plant and animal species. PMID:27587297

  11. Generation of marker-free plastid transformants using a transiently cointegrated selection gene.

    PubMed

    Klaus, Sebastian M J; Huang, Fong-Chin; Golds, Timothy J; Koop, Hans-Ulrich

    2004-02-01

    Genetic engineering of higher plant plastids typically involves stable introduction of antibiotic resistance genes as selection markers. Even though chloroplast genes are maternally inherited in most crops, the possibility of marker transfer to wild relatives or microorganisms cannot be completely excluded. Furthermore, marker expression can be a substantial metabolic drain. Therefore, efficient methods for complete marker removal from plastid transformants are necessary. One method to remove the selection gene from higher plant plastids is based on loop-out recombination, a process difficult to control because selection of homoplastomic transformants is unpredictable. Another method uses the CRE/lox system, but requires additional retransformation and sexual crossing for introduction and subsequent removal of the CRE recombinase. Here we describe the generation of marker-free chloroplast transformants in tobacco using the reconstitution of wild-type pigmentation in combination with plastid transformation vectors, which prevent stable integration of the kanamycin selection marker. One benefit of a procedure using mutants is that marker-free plastid transformants can be produced directly in the first generation (T0) without retransformation or crossing.

  12. Selection of Taste Markers Related to Lactic Acid Bacteria Microflora Metabolism for Chinese Traditional Paocai: A Gas Chromatography-Mass Spectrometry-Based Metabolomics Approach.

    PubMed

    Zhao, Nan; Zhang, Chuchu; Yang, Qin; Guo, Zhuang; Yang, Bo; Lu, Wenwei; Li, Dongyao; Tian, Fengwei; Liu, Xiaoming; Zhang, Hao; Chen, Wei

    2016-03-23

    Traditional paocai brine (PB) is continuously propagated by back-slopping and contains numerous lactic acid bacteria (LAB) strains. Although PB is important for the quality of paocai (Chinese sauerkraut), the taste features, taste-related compounds of PB-paocai and the effects of LAB communities from PB on the taste compounds remain unclear. An electronic tongue was used to evaluate the taste features of 13 PB-paocai samples. Umami, saltiness, bitterness, sweetness, and aftertaste astringency were the main taste features of PB-paocai. A total of 14 compounds were identified as discriminant taste markers for PB-paocai via gas chromatography-mass spectrometry (GC-MS)-based multimarker profiling. A LAB co-culture (Lactobacillus plantarum, Lactobacillus buchneri, and Pediococcus ethanoliduran) from PB could significantly increase glutamic acid (umami), sucrose (sweetness), glycine (sweetness), lactic acid (sourness), and γ-aminobutyric acid in PB-paocai, which would endow it with important flavor features. Such features could then facilitate starter screening and fermentation optimization to produce paocai-related foods with better nutritional and sensory qualities.

  13. TETX: a novel nuclear selection marker for Chlamydomonas reinhardtii transformation.

    PubMed

    Garcia-Echauri, Sergio A; Cardineau, Guy A

    2015-01-01

    Transformation of microalgae to obtain recombinant proteins, lipids or metabolites of economic value is of growing interest due to low costs associated with culture growth and scaling up. At present there are only three stable nuclear selection markers for the transformation of Chlamydomonas reinhardtii, which is the most commonly transformed microalgae, specifically: the aminoglycoside phosphotransferaseses aph7and aphVIII and the phleomycin resistance ble gene. As several microalgae are resistant to some of the antibiotics associated with the mentioned resistance genes, we have developed another alternative, tetX, a NADP-requiring Oxidoreductase that hydroxylates tetracycline substrates. We provide evidence that tetX can be used to obtain nuclear transformants of Chlamydomonas reinhardtii. We obtained nuclear transformants harbouring the tetX gene under the control of beta 2 tubulin or HSP70ARBCS2 promoters at an efficiency of transformation of 3.28 and 6.18 colony forming units/μg DNA respectively. This is the first report of a eukaryotic cell transformed using tetracycline as a selectable marker. We developed a protocol for the nuclear transformation of Chlamydomonas reinhardtii using tetX as a selectable marker that confers stable resistance to tetracycline up to 100 μg/mL. We believe tetX can be used to transform Chlamydomonas reinhardtii chloroplasts, related microalgae and other aerobic organisms sensitive to any tetracycline antibiotic.

  14. Modeling of genetic gain for single traits from marker-assisted seedling selection in clonally propagated crops

    PubMed Central

    Ru, Sushan; Hardner, Craig; Carter, Patrick A; Evans, Kate; Main, Dorrie; Peace, Cameron

    2016-01-01

    Seedling selection identifies superior seedlings as candidate cultivars based on predicted genetic potential for traits of interest. Traditionally, genetic potential is determined by phenotypic evaluation. With the availability of DNA tests for some agronomically important traits, breeders have the opportunity to include DNA information in their seedling selection operations—known as marker-assisted seedling selection. A major challenge in deploying marker-assisted seedling selection in clonally propagated crops is a lack of knowledge in genetic gain achievable from alternative strategies. Existing models based on additive effects considering seed-propagated crops are not directly relevant for seedling selection of clonally propagated crops, as clonal propagation captures all genetic effects, not just additive. This study modeled genetic gain from traditional and various marker-based seedling selection strategies on a single trait basis through analytical derivation and stochastic simulation, based on a generalized seedling selection scheme of clonally propagated crops. Various trait-test scenarios with a range of broad-sense heritability and proportion of genotypic variance explained by DNA markers were simulated for two populations with different segregation patterns. Both derived and simulated results indicated that marker-based strategies tended to achieve higher genetic gain than phenotypic seedling selection for a trait where the proportion of genotypic variance explained by marker information was greater than the broad-sense heritability. Results from this study provides guidance in optimizing genetic gain from seedling selection for single traits where DNA tests providing marker information are available. PMID:27148453

  15. The effect of using approximate gametic variance covariance matrices on marker assisted selection by BLUP.

    PubMed

    Totir, Liviu R; Fernando, Rohan L; Dekkers, Jack C M; Fernández, Soledad A; Guldbrandtsen, Bernt

    2004-01-01

    Under additive inheritance, the Henderson mixed model equations (HMME) provide an efficient approach to obtaining genetic evaluations by marker assisted best linear unbiased prediction (MABLUP) given pedigree relationships, trait and marker data. For large pedigrees with many missing markers, however, it is not feasible to calculate the exact gametic variance covariance matrix required to construct HMME. The objective of this study was to investigate the consequences of using approximate gametic variance covariance matrices on response to selection by MABLUP. Two methods were used to generate approximate variance covariance matrices. The first method (Method A) completely discards the marker information for individuals with an unknown linkage phase between two flanking markers. The second method (Method B) makes use of the marker information at only the most polymorphic marker locus for individuals with an unknown linkage phase. Data sets were simulated with and without missing marker data for flanking markers with 2, 4, 6, 8 or 12 alleles. Several missing marker data patterns were considered. The genetic variability explained by marked quantitative trait loci (MQTL) was modeled with one or two MQTL of equal effect. Response to selection by MABLUP using Method A or Method B were compared with that obtained by MABLUP using the exact genetic variance covariance matrix, which was estimated using 15,000 samples from the conditional distribution of genotypic values given the observed marker data. For the simulated conditions, the superiority of MABLUP over BLUP based only on pedigree relationships and trait data varied between 0.1% and 13.5% for Method A, between 1.7% and 23.8% for Method B, and between 7.6% and 28.9% for the exact method. The relative performance of the methods under investigation was not affected by the number of MQTL in the model.

  16. Selection of chemical markers for the quality control of medicinal plants of the genus Cecropia.

    PubMed

    Rivera-Mondragón, Andrés; Ortíz, Orlando O; Bijttebier, Sebastiaan; Vlietinck, Arnold; Apers, Sandra; Pieters, Luc; Caballero-George, Catherina

    2017-12-01

    Several Cecropia (Cecropiaceae) species are traditionally used in Latin America for the treatment of a variety of diseases including diabetes, arterial hypertension, asthma, bronchitis, anxiety, and inflammation. At present, a number of commercial products based on these plants have been introduced into the market with very little information on methods for guaranteeing their quality and safety. This work proposes potential chemical markers for the quality control of the raw materials of Cecropia obtusifolia Bertol., Cecropia peltata L., Cecropia glaziovii Snethl., Cecropia pachystachya Trécul, and Cecropia hololeuca Miq. The Herbal Chemical Marker Ranking System (Herb MaRS) developed by the National Institute of Complementary Medicine (NICM) at the University of Western Sydney was used for selecting chemical markers for the quality control of selected medicinal species of Cecropia. This review covers the period from 1982 to 2016. Chlorogenic acid, flavonoidal glycosides (orientin, isoorientin, vitexin, isovitexin, and rutin), catechin, epicatechin, procyanidins (B2, B5, and C1), steroids (β-sitosterol), and triterpenoids (α-amyrin, pomolic, tormentic and ursolic acids) were selected as chemical markers for the quality control of the leaves. It is necessary to establish comprehensive standards for guaranteeing quality, safety and efficacy of herbal drugs. The selection of adequate chemical markers for quality control purposes requires a good knowledge about the chemical composition of medicinal plants and their associated biological properties. To the best of our knowledge this review article is the first to address the identification and quantitative determination of the chemical markers for the genus Cecropia.

  17. Manipulator-driven selection of semi-active MR-visible markers.

    PubMed

    An, Junmo; Webb, Andrew G; Shah, Dipan J; Chin, Karen; Tsekos, Nikolaos V

    2017-06-29

    A method for the identification of semi-active fiducial magnetic resonance (MR) markers is presented based on selectively optically tuning and detuning them. Four inductively coupled solenoid coils with photoresistors were connected to light sources. A microcontroller timed the optical tuning/detuning of coils and image collection. The markers were tested on an MR manipulator linking the microcontroller to the manipulator control to visibly select the marker subset according to the actuated joint. In closed-loop control, the average and maximum were 0.76° ± 0.41° and 1.18° errors for a rotational joint, and 0.87 mm ± 0.26 mm and 1.13 mm for the prismatic joint. This technique is suitable for MR-compatible actuated devices that use semi-active MR-compatible markers. Copyright © 2017 John Wiley & Sons, Ltd.

  18. Improving selection of markers in nutrition research: evaluation of the criteria proposed by the ILSI Europe Marker Validation Initiative.

    PubMed

    Calder, Philip C; Boobis, Alan; Braun, Deborah; Champ, Claire L; Dye, Louise; Einöther, Suzanne; Greyling, Arno; Matthys, Christophe; Putz, Peter; Wopereis, Suzan; Woodside, Jayne V; Antoine, Jean-Michel

    2017-06-01

    The conduct of high-quality nutrition research requires the selection of appropriate markers as outcomes, for example as indicators of food or nutrient intake, nutritional status, health status or disease risk. Such selection requires detailed knowledge of the markers, and consideration of the factors that may influence their measurement, other than the effects of nutritional change. A framework to guide selection of markers within nutrition research studies would be a valuable tool for researchers. A multidisciplinary Expert Group set out to test criteria designed to aid the evaluation of candidate markers for their usefulness in nutrition research and subsequently to develop a scoring system for markers. The proposed criteria were tested using thirteen markers selected from a broad range of nutrition research fields. The result of this testing was a modified list of criteria and a template for evaluating a potential marker against the criteria. Subsequently, a semi-quantitative system for scoring a marker and an associated template were developed. This system will enable the evaluation and comparison of different candidate markers within the same field of nutrition research in order to identify their relative usefulness. The ranking criteria of proven, strong, medium or low are likely to vary according to research setting, research field and the type of tool used to assess the marker and therefore the considerations for scoring need to be determined in a setting-, field- and tool-specific manner. A database of such markers, their interpretation and range of possible values would be valuable to nutrition researchers.

  19. Marker-assisted selection of Fusarium wilt-resistant and gynoecious melon (Cucumis melo L.).

    PubMed

    Gao, P; Liu, S; Zhu, Q L; Luan, F S

    2015-12-08

    In this study, molecular markers were designed based on the sex determination genes ACS7 (A) and WIP1 (G) and the domain in the Fusarium oxysporum-resistant gene Fom-2 (F) in order to achieve selection of F. oxysporum-resistant gynoecious melon plants. Markers of A and F are cleaved amplified polymorphic sequences that distinguish alleles according to restriction analysis. Twenty F1 and 1863 F2 plants derived from the crosses between the gynoecious line WI998 and the Fusarium wilt-resistant line MR-1 were genotyped based on the markers. The results showed that the polymerase chain reaction and enzyme digestion results could be effectively used to identify plants with the AAggFF genotype in F2 populations. In the F2 population, 35 gynoecious wilt-resistant plants were selected by marker-assisted selection and were confirmed by disease infection assays, demonstrating that these markers can be used in breeding to select F. oxysporum-resistant gynoecious melon plants.

  20. A pseudo-R2 measure for selecting genomic markers with crossing hazards functions

    PubMed Central

    2011-01-01

    Background In genomic medical studies, one of the major objectives is to identify genomic factors with a prognostic impact on time-to-event outcomes so as to provide new insights into the disease process. Selection usually relies on statistical univariate indices based on the Cox model. Such model assumes proportional hazards (PH) which is unlikely to hold for each genomic marker. Methods In this paper, we introduce a novel pseudo-R2 measure derived from a crossing hazards model and designed for the selection of markers with crossing effects. The proposed index is related to the score statistic and quantifies the extent of a genomic factor to separate patients according to their survival times and marker measurements. We also show the importance of considering genomic markers with crossing effects as they potentially reflect the complex interplay between markers belonging to the same pathway. Results Simulations show that our index is not affected by the censoring and the sample size of the study. It also performs better than classical indices under the crossing hazards assumption. The practical use of our index is illustrated in a lung cancer study. The use of the proposed pseudo-R2 allows the identification of cell-cycle dependent genes not identified when relying on the PH assumption. Conclusions The proposed index is a novel and promising tool for selecting markers with crossing hazards effects. PMID:21401964

  1. Advances in Research to Improve Selection for Beef Production and Quality Using Genetic Markers

    USDA-ARS?s Scientific Manuscript database

    Genetic marker technology for selection of beef cattle has changed and continues to change. Some ideas about how this technology would be used and how much of the genetic differences could easily be identified have changed. New approaches based on what we have learned so far and on new genotyping ...

  2. A framework linkage map of perennial ryegrass based on SSR markers

    Treesearch

    G.P. Gill; P.L. Wilcox; D.J. Whittaker; R.A. Winz; P. Bickerstaff; Craig E. Echt; J. Kent; M.O. Humphreys; K.M. Elborough; R.C. Gardner

    2006-01-01

    A moderate-density linkage map for Lolium perenne L. has been constructed based on 376 simple sequence repeat (SSR) markers. Approximately one third ( 124) of the SSR markers were developed from GeneThresher libraries that preferentially select genomic DNA clones from the gene-rich unmethylated portion of the genome. The remaining SSR marker loci...

  3. Ghost marker detection and elimination in marker-based optical tracking systems for real-time tracking in stereotactic body radiotherapy

    SciTech Connect

    Yan, Guanghua Li, Jonathan; Huang, Yin; Mittauer, Kathryn; Lu, Bo; Liu, Chihray

    2014-10-15

    Purpose: To propose a simple model to explain the origin of ghost markers in marker-based optical tracking systems (OTS) and to develop retrospective strategies to detect and eliminate ghost markers. Methods: In marker-based OTS, ghost markers are virtual markers created due to the cross-talk between the two camera sensors, which can lead to system execution failure or inaccuracy in patient tracking. As a result, the users have to limit the number of markers and avoid certain marker configurations to reduce the chances of ghost markers. In this work, the authors propose retrospective strategies to detect and eliminate ghost markers. The two camera sensors were treated as mathematical points in space. The authors identified the coplanar within limit (CWL) condition as the necessary condition for ghost marker occurrence. A simple ghost marker detection method was proposed based on the model. Ghost marker elimination was achieved through pattern matching: a ghost marker-free reference set was matched with the optical marker set observed by the OTS; unmatched optical markers were eliminated as either ghost markers or misplaced markers. The pattern matching problem was formulated as a constraint satisfaction problem (using pairwise distances as constraints) and solved with an iterative backtracking algorithm. Wildcard markers were introduced to address missing or misplaced markers. An experiment was designed to measure the sensor positions and the limit for the CWL condition. The ghost marker detection and elimination algorithms were verified with samples collected from a five-marker jig and a nine-marker anthropomorphic phantom, rotated with the treatment couch from −60° to +60°. The accuracy of the pattern matching algorithm was further validated with marker patterns from 40 patients who underwent stereotactic body radiotherapy (SBRT). For this purpose, a synthetic optical marker pattern was created for each patient by introducing ghost markers, marker position

  4. Comparing genomic selection and marker-assisted selection for Fusarium head blight resistance in wheat (Triticum aestivum L.)

    USDA-ARS?s Scientific Manuscript database

    Genomic selection (GS) and marker-assisted selection (MAS) rely on marker-trait associations and are both routinely used for breeding purposes. Although similar, these two approaches differ in their applications and how markers are used to estimate breeding values. In this study, GS and MAS were com...

  5. Interpretation of variation across marker loci as evidence of selection.

    PubMed Central

    Vitalis, R; Dawson, K; Boursot, P

    2001-01-01

    Population structure and history have similar effects on the genetic diversity at all neutral loci. However, some marker loci may also have been strongly influenced by natural selection. Selection shapes genetic diversity in a locus-specific manner. If we could identify those loci that have responded to selection during the divergence of populations, then we may obtain better estimates of the parameters of population history by excluding these loci. Previous attempts were made to identify outlier loci from the distribution of sample statistics under neutral models of population structure and history. Unfortunately these methods depend on assumptions about population structure and history that usually cannot be verified. In this article, we define new population-specific parameters of population divergence and construct sample statistics that are estimators of these parameters. We then use the joint distribution of these estimators to identify outlier loci that may be subject to selection. We found that outlier loci are easier to recognize when this joint distribution is conditioned on the total number of allelic states represented in the pooled sample at each locus. This is so because the conditional distribution is less sensitive to the values of nuisance parameters. PMID:11514464

  6. a Comparison Study of Different Marker Selection Methods for Spectral-Spatial Classification of Hyperspectral Images

    NASA Astrophysics Data System (ADS)

    Akbari, D.; Safari, A. R.; Homayouni, S.; Khazai, S.

    2015-12-01

    An effective approach based on the Minimum Spanning Forest (MSF), grown from automatically selected markers using Support Vector Machines (SVM), has been proposed for spectral-spatial classification of hyperspectral images by Tarabalka et al. This paper aims at improving this approach by using image segmentation to integrate the spatial information into marker selection process. In this study, the markers are extracted from the classification maps, obtained by both SVM and segmentation algorithms, and then are used to build the MSF. The segmentation algorithms are the watershed, expectation maximization (EM) and hierarchical clustering. These algorithms are used in parallel and independently to segment the image. Moreover, the pixels of each class, with the largest population in the classification map, are kept for each region of the segmentation map. Lastly, the most reliable classified pixels are chosen from among the exiting pixels as markers. Two benchmark urban hyperspectral datasets are used for evaluation: Washington DC Mall and Berlin. The results of our experiments indicate that, compared to the original MSF approach, the marker selection using segmentation algorithms leads in more accurate classification maps.

  7. Biosafety considerations for selectable and scorable markers used in cassava (Manihot esculenta Crantz) biotechnology.

    PubMed

    Petersen, William; Umbeck, Paul; Hokanson, Karen; Halsey, Mark

    2005-01-01

    Cassava is an important subsistence crop grown only in the tropics, and represents a major source of calories for many people in developing countries. Improvements in the areas of resistance to insects and viral diseases, enhanced nutritional qualities, reduced cyanogenic content and modified starch characteristics are urgently needed. Traditional breeding is hampered by the nature of the crop, which has a high degree of heterozygosity, irregular flowering, and poor seed set. Biotechnology has the potential to enhance crop improvement efforts, and genetic engineering techniques for cassava have thus been developed over the past decade. Selectable and scorable markers are critical to efficient transformation technology, and must be evaluated for biosafety, as well as efficiency and cost-effectiveness. In order to facilitate research planning and regulatory submission, the literature on biosafety aspects of the selectable and scorable markers currently used in cassava biotechnology is surveyed. The source, mode of action and current use of each marker gene is described. The potential for toxicity, allergenicity, pleiotropic effects, horizontal gene transfer, and the impact of these on food or feed safety and environmental safety is evaluated. Based on extensive information, the selectable marker genes nptII, hpt, bar/pat, and manA, and the scorable marker gene uidA, all have little risk in terms of biosafety. These appear to represent the safest options for use in cassava biotechnology available at this time.

  8. Selective DNA Pooling for Determination of Linkage between a Molecular Marker and a Quantitative Trait Locus

    PubMed Central

    Darvasi, A.; Soller, M.

    1994-01-01

    Selective genotyping is a method to reduce costs in marker-quantitative trait locus (QTL) linkage determination by genotyping only those individuals with extreme, and hence most informative, quantitative trait values. The DNA pooling strategy (termed: ``selective DNA pooling'') takes this one step further by pooling DNA from the selected individuals at each of the two phenotypic extremes, and basing the test for linkage on marker allele frequencies as estimated from the pooled samples only. This can reduce genotyping costs of marker-QTL linkage determination by up to two orders of magnitude. Theoretical analysis of selective DNA pooling shows that for experiments involving backcross, F(2) and half-sib designs, the power of selective DNA pooling for detecting genes with large effect, can be the same as that obtained by individual selective genotyping. Power for detecting genes with small effect, however, was found to decrease strongly with increase in the technical error of estimating allele frequencies in the pooled samples. The effect of technical error, however, can be markedly reduced by replication of technical procedures. It is also shown that a proportion selected of 0.1 at each tail will be appropriate for a wide range of experimental conditions. PMID:7896115

  9. User-friendly markers linked to Fusarium wilt race 1 resistance Fw gene for marker-assisted selection in pea

    USDA-ARS?s Scientific Manuscript database

    Fusarium wilt is one of the most widespread diseases of pea. Resistance to Fusarium wilt race 1 was reported as a single gene, Fw, located on linkage group III. The previously reported AFLP and RAPD markers linked to Fw have limited usage in marker-assisted selection due to their map distance and l...

  10. Selection of Genetic Markers for Association Analyses, Using Linkage Disequilibrium and Haplotypes

    PubMed Central

    Meng, Zhaoling; Zaykin, Dmitri V.; Xu, Chun-Fang; Wagner, Michael; Ehm, Margaret G.

    2003-01-01

    The genotyping of closely spaced single-nucleotide polymorphism (SNP) markers frequently yields highly correlated data, owing to extensive linkage disequilibrium (LD) between markers. The extent of LD varies widely across the genome and drives the number of frequent haplotypes observed in small regions. Several studies have illustrated the possibility that LD or haplotype data could be used to select a subset of SNPs that optimize the information retained in a genomic region while reducing the genotyping effort and simplifying the analysis. We propose a method based on the spectral decomposition of the matrices of pairwise LD between markers, and we select markers on the basis of their contributions to the total genetic variation. We also modify Clayton’s “haplotype tagging SNP” selection method, which utilizes haplotype information. For both methods, we propose sliding window–based algorithms that allow the methods to be applied to large chromosomal regions. Our procedures require genotype information about a small number of individuals for an initial set of SNPs and selection of an optimum subset of SNPs that could be efficiently genotyped on larger numbers of samples while retaining most of the genetic variation in samples. We identify suitable parameter combinations for the procedures, and we show that a sample size of 50–100 individuals achieves consistent results in studies of simulated data sets in linkage equilibrium and LD. When applied to experimental data sets, both procedures were similarly effective at reducing the genotyping requirement while maintaining the genetic information content throughout the regions. We also show that haplotype-association results that Hosking et al. obtained near CYP2D6 were almost identical before and after marker selection. PMID:12796855

  11. MPR1 as a novel selection marker in Saccharomyces cerevisiae.

    PubMed

    Ogawa-Mitsuhashi, Kaoru; Sagane, Koji; Kuromitsu, Junro; Takagi, Hiroshi; Tsukahara, Kappei

    2009-11-01

    L-Azetidine-2-carboxylic acid (AZC) is a toxic four-membered ring analogue of L-proline that is transported into cells by proline transporters. AZC and L-proline in the cells are competitively incorporated into nascent proteins. When AZC is present in a minimum medium, misfolded proteins are synthesized in the cells, thereby inhibiting cell growth. The MPR1 gene has been isolated from the budding yeast Saccharomyces cerevisiae Sigma1278b as a multicopy suppressor of AZC-induced growth inhibition. MPR1 encodes a novel acetyltransferase that detoxifies AZC via N-acetylation. Since MPR1 is absent in the laboratory strain of S. cerevisiae S288C, it could be a positive selection marker that confers AZC resistance in the S288C background strains. To examine the usefulness of MPR1, we constructed some plasmid vectors that harboured MPR1 under the control of various promoters and introduced them into the S288C-derived strains. The expression of MPR1 conferred AZC resistance that was largely dependent on the expression level of MPR1. In an additional experiment, the galactose-inducible MPR1 and ppr1(+), the fission yeast Schizosaccharomyces pombe homologue of MPR1, were used for gene disruption by homologous recombination, and here AZC-resistant colonies were also successfully selected. We concluded that our MPR1-AZC system provides a powerful tool for yeast transformation. Copyright (c) 2009 John Wiley & Sons, Ltd.

  12. Impact of chronic lead exposure on selected biological markers.

    PubMed

    Jangid, Ambica P; John, P J; Yadav, D; Mishra, Sandhya; Sharma, Praveen

    2012-01-01

    Lead poisoning remains a major problem in India due to the lack of awareness of its ill effects among the clinical community. Blood lead, δ-aminolevulinic acid dehydratase (δ-ALAD) and zinc protoporphyrin (ZPP) concentrations are widely used as biomarkers for lead toxicity The present study was designed to determine the impact of chronic lead exposure on selected biological markers. A total of 250 subjects, of both sexes, ranging in age from 20 to 70 years, were recruited. On the basis of BLLs, the subjects were categorized into four groups: Group A (BLL: 0-10 μg/dl), Group B (BLL: 10-20 μg/dl). Group C (BLL: 20-30 μg/dl) and Group D (BLL: 30-40 μg/dl) having BLLs of 3.60 ± 2.71 μg/dl, 15.21 ± 2.65 μg/dl, 26.82 ± 2.53 μg/dl and 36.38 ± 2.83 μg/dl, respectively. Significant changes in biological markers due to elevated BLLs were noted. The relation of BLL and biological markers to demographic characteristics such as sex, habits, diet and substances abuse (smoking effect) were also studied in the present investigation. Males, urban population, non-vegetarians, and smokers had higher blood lead levels. δ-ALAD activity was found to be significantly lower with increased BLL (P < 0.001), while the ZPP level was significantly higher with increased BLL (P < 0.001). Further, BLL showed a negative correlation with δ-ALAD (r = -0.425, P < 0.001, N = 250) and a positive correlations with ZPP (r = 0.669, P < 0.001, N = 250). Chronic lead exposure affects the prooxidant-antioxidant equilibrium leading to cellular oxidative stress.

  13. Identification of two novel inactive DFR-A alleles responsible for failure to produce anthocyanin and development of a simple PCR-based molecular marker for bulb color selection in onion (Allium cepa L.).

    PubMed

    Kim, Sunggil; Baek, Doohyun; Cho, Dong Youn; Lee, Eul-Tai; Yoon, Moo-Kyoung

    2009-05-01

    Two novel inactive alleles of Dihydroflavonol 4-reductase-A (DFR-A) were identified in yellow onion (Allium cepa L.) cultivars and breeding lines from Korea and Japan. Unlike the previously reported inactive yellow DFR-A allele, designated as DFR-A ( TRN ) , in which the 3' portion of the coding sequences was deleted, an allele containing a premature stop codon, DFR-A ( PS ) , was isolated from the majority of cultivars. Co-segregation of DFR-A ( PS ) and color phenotypes in the F(2) population from a cross between yellow and red parents showed that inactivation of DFR-A was responsible for lack of anthocyanin in these yellow onions. In addition, RT-PCR analysis of F(2) population showed that the transcription level of the DFR-A ( PS ) allele was significantly reduced owing to non-sense-mediated mRNA decay. A 20-bp deletion of a simple sequence repeat in the promoter region of the DFR-A ( PS ) allele was used to develop a simple PCR-based molecular marker for selection of the DFR-A ( PS ) allele. All genotypes of 138 F(2) individuals were clearly distinguished by this molecular marker. In addition to the DFR-A ( PS ) allele, another DFR-A allele, DFR-A ( DEL ) , was identified in some cultivars. In case of the DFR-A ( DEL ) allele, no PCR products were amplified throughout DFR-A sequences including promoter regions, suggesting deletion of the entire DFR-A gene. Co-segregation of the absence of DFR-A and color phenotypes was confirmed in another F(2) population. Furthermore, RT-PCR results showed that no DFR-A transcript was detected in any yellow F(2) individuals.

  14. Production of marker-free disease-resistant potato using isopentenyl transferase gene as a positive selection marker.

    PubMed

    Khan, Raham Sher; Ntui, Valentine Otang; Chin, Dong Poh; Nakamura, Ikuo; Mii, Masahiro

    2011-04-01

    The use of antibiotic or herbicide resistant genes as selection markers for production of transgenic plants and their continuous presence in the final transgenics has been a serious problem for their public acceptance and commercialization. MAT (multi-auto-transformation) vector system has been one of the different strategies to excise the selection marker gene and produce marker-free transgenic plants. In the present study, ipt (isopentenyl transferase) gene was used as a selection marker gene. A chitinase gene, ChiC (isolated from Streptomyces griseus strain HUT 6037) was used as a gene of interest. ChiC gene was cloned from the binary vector, pEKH1 to an ipt-type MAT vector, pMAT21 by gateway cloning and transferred to Agrobacterium tumefaciens strain EHA105. The infected tuber discs of potato were cultured on hormone- and antibiotic-free MS medium. Seven of the 35 explants infected with the pMAT21/ChiC produced shoots. The same antibiotic- and hormones-free MS medium was used in subcultures of the shoots (ipt like and normal shoots). Molecular analyses of genomic DNA from transgenic plants confirmed the integration of gene of interest and excision of the selection marker in 3 of the 7 clones. Expression of ChiC gene was confirmed by Northern blot and western blot analyses. Disease-resistant assay of the marker-free transgenic, in vitro and greenhouse-grown plants exhibited enhanced resistance against Alternaria solani (early blight), Botrytis cinerea (gray mold) and Fusarium oxysporum (Fusarium wilt). From these results it could be concluded that ipt gene can be used as a selection marker to produce marker-free disease-resistant transgenic potato plants on PGR- and antibiotic-free MS medium.

  15. The dsdA gene from Escherichia coli provides a novel selectable marker for plant transformation.

    PubMed

    Erikson, Oskar; Hertzberg, Magnus; Näsholm, Torgny

    2005-02-01

    Plants are sensitive to D-serine, but functional expression of the dsdA gene, encoding D-serine ammonia lyase, from Escherichia coli can alleviate this toxicity. Plants, in contrast to many other organisms, lack the common pathway for oxidative deamination of D-amino acids. This difference in metabolism has major consequences for plant responses to D-amino acids, since several D-amino acids are toxic to plants even at relatively low concentrations. Therefore, introducing an enzyme specific for a phytotoxic D-amino acid should generate a selectable characteristic that can be screened. Here we present the use of the dsdA gene as a selectable marker for transformation of Arabidopsis. D-serine ammonia lyase catalyses the deamination of D-serine into pyruvate, water and ammonium. dsdA transgenic seedlings can be clearly distinguished from wild type, having an unambiguous phenotype immediately following germination when selected on D-serine containing medium. The dsdA marker allows flexibility in application of the selective agent: it can be applied in sterile plates, in foliar sprays or in liquid culture. Selection with D-serine resistance was compared with selection based on kanamycin resistance, and was found to generate similar transformation frequencies but also to be more unambiguous, more rapid and more versatile with respect to the way the selective agent can be supplied.

  16. Development of Gateway Binary Vector Series with Four Different Selection Markers for the Liverwort Marchantia polymorpha.

    PubMed

    Ishizaki, Kimitsune; Nishihama, Ryuichi; Ueda, Minoru; Inoue, Keisuke; Ishida, Sakiko; Nishimura, Yoshiki; Shikanai, Toshiharu; Kohchi, Takayuki

    2015-01-01

    We previously reported Agrobacterium-mediated transformation methods for the liverwort Marchantia polymorpha using the hygromycin phosphotransferase gene as a marker for selection with hygromycin. In this study, we developed three additional markers for M. polymorpha transformation: the gentamicin 3'-acetyltransferase gene for selection with gentamicin; a mutated acetolactate synthase gene for selection with chlorsulfuron; and the neomycin phosphotransferase II gene for selection with G418. Based on these four marker genes, we have constructed a series of Gateway binary vectors designed for transgenic experiments on M. polymorpha. The 35S promoter from cauliflower mosaic virus and endogenous promoters for constitutive and heat-inducible expression were used to create these vectors. The reporters and tags used were Citrine, 3×Citrine, Citrine-NLS, TagRFP, tdTomato, tdTomato-NLS, GR, SRDX, SRDX-GR, GUS, ELuc(PEST), and 3×FLAG. These vectors, designated as the pMpGWB series, will facilitate molecular genetic analyses of the emerging model plant M. polymorpha.

  17. Development of Gateway Binary Vector Series with Four Different Selection Markers for the Liverwort Marchantia polymorpha

    PubMed Central

    Ueda, Minoru; Inoue, Keisuke; Ishida, Sakiko; Nishimura, Yoshiki; Shikanai, Toshiharu; Kohchi, Takayuki

    2015-01-01

    We previously reported Agrobacterium-mediated transformation methods for the liverwort Marchantia polymorpha using the hygromycin phosphotransferase gene as a marker for selection with hygromycin. In this study, we developed three additional markers for M. polymorpha transformation: the gentamicin 3'-acetyltransferase gene for selection with gentamicin; a mutated acetolactate synthase gene for selection with chlorsulfuron; and the neomycin phosphotransferase II gene for selection with G418. Based on these four marker genes, we have constructed a series of Gateway binary vectors designed for transgenic experiments on M. polymorpha. The 35S promoter from cauliflower mosaic virus and endogenous promoters for constitutive and heat-inducible expression were used to create these vectors. The reporters and tags used were Citrine, 3×Citrine, Citrine-NLS, TagRFP, tdTomato, tdTomato-NLS, GR, SRDX, SRDX-GR, GUS, ELuc(PEST), and 3×FLAG. These vectors, designated as the pMpGWB series, will facilitate molecular genetic analyses of the emerging model plant M. polymorpha. PMID:26406247

  18. Association mapping and marker-assisted selection of the lettuce dieback resistance gene Tvr1

    PubMed Central

    2009-01-01

    Background Lettuce (Lactuca saliva L.) is susceptible to dieback, a soilborne disease caused by two viruses from the family Tombusviridae. Susceptibility to dieback is widespread in romaine and leaf-type lettuce, while modern iceberg cultivars are resistant to this disease. Resistance in iceberg cultivars is conferred by Tvr1 - a single, dominant gene that provides durable resistance. This study describes fine mapping of the resistance gene, analysis of nucleotide polymorphism and linkage disequilibrium in the Tvr1 region, and development of molecular markers for marker-assisted selection. Results A combination of classical linkage mapping and association mapping allowed us to pinpoint the location of the Tvr1 resistance gene on chromosomal linkage group 2. Nine molecular markers, based on expressed sequence tags (EST), were closely linked to Tvr1 in the mapping population, developed from crosses between resistant (Salinas and Salinas 88) and susceptible (Valmaine) cultivars. Sequencing of these markers from a set of 68 cultivars revealed a relatively high level of nucleotide polymorphism (θ = 6.7 × 10-3) and extensive linkage disequilibrium (r2 = 0.124 at 8 cM) in this region. However, the extent of linkage disequilibrium was affected by population structure and the values were substantially larger when the analysis was performed only for romaine (r2 = 0.247) and crisphead (r2 = 0.345) accessions. The association mapping approach revealed that one of the nine markers (Cntg10192) in the Tvr1 region matched exactly with resistant and susceptible phenotypes when tested on a set of 200 L. sativa accessions from all horticultural types of lettuce. The marker-trait association was also confirmed on two accessions of Lactuca serriola - a wild relative of cultivated lettuce. The combination of three single-nucleotide polymorphisms (SNPs) at the Cntg10192 marker identified four haplotypes. Three of the haplotypes were associated with resistance and one of them was always

  19. Imputation of unordered markers and the impact on genomic selection accuracy

    USDA-ARS?s Scientific Manuscript database

    Genomic selection, a breeding method that promises to accelerate rates of genetic gain, requires dense, genome-wide marker data. Genotyping-by-sequencing can generate a large number of de novo markers. However, without a reference genome, these markers are unordered and typically have a large propo...

  20. Weibull regression with Bayesian variable selection to identify prognostic tumour markers of breast cancer survival.

    PubMed

    Newcombe, P J; Raza Ali, H; Blows, F M; Provenzano, E; Pharoah, P D; Caldas, C; Richardson, S

    2017-02-01

    As data-rich medical datasets are becoming routinely collected, there is a growing demand for regression methodology that facilitates variable selection over a large number of predictors. Bayesian variable selection algorithms offer an attractive solution, whereby a sparsity inducing prior allows inclusion of sets of predictors simultaneously, leading to adjusted effect estimates and inference of which covariates are most important. We present a new implementation of Bayesian variable selection, based on a Reversible Jump MCMC algorithm, for survival analysis under the Weibull regression model. A realistic simulation study is presented comparing against an alternative LASSO-based variable selection strategy in datasets of up to 20,000 covariates. Across half the scenarios, our new method achieved identical sensitivity and specificity to the LASSO strategy, and a marginal improvement otherwise. Runtimes were comparable for both approaches, taking approximately a day for 20,000 covariates. Subsequently, we present a real data application in which 119 protein-based markers are explored for association with breast cancer survival in a case cohort of 2287 patients with oestrogen receptor-positive disease. Evidence was found for three independent prognostic tumour markers of survival, one of which is novel. Our new approach demonstrated the best specificity.

  1. Comparison of two alternative dominant selectable markers for wine yeast transformation.

    PubMed

    Cebollero, Eduardo; Gonzalez, Ramon

    2004-12-01

    Genetic improvement of industrial yeast strains is restricted by the availability of selectable transformation markers. Antibiotic resistance markers have to be avoided for public health reasons, while auxotrophy markers are generally not useful for wine yeast strain transformation because most industrial Saccharomyces cerevisiae strains are prototrophic. For this work, we performed a comparative study of the usefulness of two alternative dominant selectable markers in both episomic and centromeric plasmids. Even though the selection for sulfite resistance conferred by FZF1-4 resulted in a larger number of transformants for a laboratory strain, the p-fluoro-DL-phenylalanine resistance conferred by ARO4-OFP resulted in a more suitable selection marker for all industrial strains tested. Both episomic and centromeric constructions carrying this marker resulted in transformation frequencies close to or above 10(3) transformants per microg of DNA for the three wine yeast strains tested.

  2. Comparison of Two Alternative Dominant Selectable Markers for Wine Yeast Transformation

    PubMed Central

    Cebollero, Eduardo; Gonzalez, Ramon

    2004-01-01

    Genetic improvement of industrial yeast strains is restricted by the availability of selectable transformation markers. Antibiotic resistance markers have to be avoided for public health reasons, while auxotrophy markers are generally not useful for wine yeast strain transformation because most industrial Saccharomyces cerevisiae strains are prototrophic. For this work, we performed a comparative study of the usefulness of two alternative dominant selectable markers in both episomic and centromeric plasmids. Even though the selection for sulfite resistance conferred by FZF1-4 resulted in a larger number of transformants for a laboratory strain, the p-fluoro-dl-phenylalanine resistance conferred by ARO4-OFP resulted in a more suitable selection marker for all industrial strains tested. Both episomic and centromeric constructions carrying this marker resulted in transformation frequencies close to or above 103 transformants per μg of DNA for the three wine yeast strains tested. PMID:15574895

  3. Network Marker Selection for Untargeted LC-MS Metabolomics Data.

    PubMed

    Cai, Qingpo; Alvarez, Jessica A; Kang, Jian; Yu, Tianwei

    2017-02-17

    Untargeted metabolomics using high-resolution liquid chromatography-mass spectrometry (LC-MS) is becoming one of the major areas of high-throughput biology. Functional analysis, that is, analyzing the data based on metabolic pathways or the genome-scale metabolic network, is critical in feature selection and interpretation of metabolomics data. One of the main challenges in the functional analyses is the lack of the feature identity in the LC-MS data itself. By matching mass-to-charge ratio (m/z) values of the features to theoretical values derived from known metabolites, some features can be matched to one or more known metabolites. When multiple matchings occur, in most cases only one of the matchings can be true. At the same time, some known metabolites are missing in the measurements. Current network/pathway analysis methods ignore the uncertainty in metabolite identification and the missing observations, which could lead to errors in the selection of significant subnetworks/pathways. In this paper, we propose a flexible network feature selection framework that combines metabolomics data with the genome-scale metabolic network. The method adopts a sequential feature screening procedure and machine learning-based criteria to select important subnetworks and identify the optimal feature matching simultaneously. Simulation studies show that the proposed method has a much higher sensitivity than the commonly used maximal matching approach. For demonstration, we apply the method on a cohort of healthy subjects to detect subnetworks associated with the body mass index (BMI). The method identifies several subnetworks that are supported by the current literature, as well as detects some subnetworks with plausible new functional implications. The R code is available at http://web1.sph.emory.edu/users/tyu8/MSS.

  4. Comparison of a retrotransposon-based marker with microsatellite markers for discriminating accessions of Vitis vinifera.

    PubMed

    Sant'Ana, G C; Ferreira, J L; Rocha, H S; Borém, A; Pasqual, M; Cançado, G M A

    2012-05-21

    Identification and knowledge concerning genetic diversity are fundamental for efficient management and use of grapevine germplasm. Recently, new types of molecular markers have been developed, such as retrotransposon-based markers. Because of their multilocus pattern, retrotransposon-based markers might be able to differentiate grapevine accessions with just one pair of primers. In order to evaluate the efficiency of this type of marker, we compared retrotransposon marker Tvv1 with seven microsatellite markers frequently used for genotyping of the genus Vitis (VVMD7, VVMD25, VVMD5, VVMD27, VVMD31, VVS2, and VZAG62). The reference population that we used consisted of 26 accessions of Vitis, including seven European varieties of Vitis vinifera, four North American varieties and hybrids of Vitis labrusca, and 15 rootstock hybrids obtained from crosses of several Vitis species. Individually, the Tvv1 and the group of seven SSR markers were capable of distinguishing all accessions except 'White Niagara' compared to 'Red Niagara'. Using the Structure software, the retrotransposon marker Tvv1 generated two clusters: one with V. vinifera plus North American varieties and the other comprising rootstocks. The seven SSR markers generated five clusters: V. vinifera, the North American varieties, and three groups of rootstock hybrids. The percentages of variation explained by the first two components in the principal coordinate analysis were 65.21 (Tvv1) and 50.42 (SSR markers) while the Mantel correlation between the distance matrixes generated by the two types of markers was 42.5%. We conclude that the Tvv1 marker is useful for DNA fingerprinting, but it lacks efficiency for discrimination of structured groups.

  5. Testing marker-based estimates of heritability in the wild.

    PubMed

    Coltman, David W

    2005-07-01

    Marker-based estimates of heritability are an attractive alternative to pedigree-based methods for estimating quantitative genetic parameters in field studies where it is difficult or impossible to determine relationships and pedigrees. Here I test the ability of the marker-based method to estimate heritability of a suite of traits in a wild population of bighorn sheep (Ovis canadensis) using marker data from 32 microsatellite loci. I compared marker-based estimates with estimates obtained using a pedigree and the animal model. Marker-based estimates of heritability were imprecise and downwardly biased. The high degree of uncertainty in marker-based estimates suggests that the method may be sufficient to detect the presence of genetic variance for highly heritable traits, but not sufficiently reliable to estimate genetic parameters.

  6. A vision-based automated guided vehicle system with marker recognition for indoor use.

    PubMed

    Lee, Jeisung; Hyun, Chang-Ho; Park, Mignon

    2013-08-07

    We propose an intelligent vision-based Automated Guided Vehicle (AGV) system using fiduciary markers. In this paper, we explore a low-cost, efficient vehicle guiding method using a consumer grade web camera and fiduciary markers. In the proposed method, the system uses fiduciary markers with a capital letter or triangle indicating direction in it. The markers are very easy to produce, manipulate, and maintain. The marker information is used to guide a vehicle. We use hue and saturation values in the image to extract marker candidates. When the known size fiduciary marker is detected by using a bird's eye view and Hough transform, the positional relation between the marker and the vehicle can be calculated. To recognize the character in the marker, a distance transform is used. The probability of feature matching was calculated by using a distance transform, and a feature having high probability is selected as a captured marker. Four directional signals and 10 alphabet features are defined and used as markers. A 98.87% recognition rate was achieved in the testing phase. The experimental results with the fiduciary marker show that the proposed method is a solution for an indoor AGV system.

  7. A Vision-Based Automated Guided Vehicle System with Marker Recognition for Indoor Use

    PubMed Central

    Lee, Jeisung; Hyun, Chang-Ho; Park, Mignon

    2013-01-01

    We propose an intelligent vision-based Automated Guided Vehicle (AGV) system using fiduciary markers. In this paper, we explore a low-cost, efficient vehicle guiding method using a consumer grade web camera and fiduciary markers. In the proposed method, the system uses fiduciary markers with a capital letter or triangle indicating direction in it. The markers are very easy to produce, manipulate, and maintain. The marker information is used to guide a vehicle. We use hue and saturation values in the image to extract marker candidates. When the known size fiduciary marker is detected by using a bird's eye view and Hough transform, the positional relation between the marker and the vehicle can be calculated. To recognize the character in the marker, a distance transform is used. The probability of feature matching was calculated by using a distance transform, and a feature having high probability is selected as a captured marker. Four directional signals and 10 alphabet features are defined and used as markers. A 98.87% recognition rate was achieved in the testing phase. The experimental results with the fiduciary marker show that the proposed method is a solution for an indoor AGV system. PMID:23966180

  8. Expanding Possibilities for Intervention against Small Ruminant Lentiviruses through Genetic Marker-Assisted Selective Breeding

    PubMed Central

    White, Stephen N.; Knowles, Donald P.

    2013-01-01

    Small ruminant lentiviruses include members that infect sheep (ovine lentivirus [OvLV]; also known as ovine progressive pneumonia virus/maedi-visna virus) and goats (caprine arthritis encephalitis virus [CAEV]). Breed differences in seroprevalence and proviral concentration of OvLV had suggested a strong genetic component in susceptibility to infection by OvLV in sheep. A genetic marker test for susceptibility to OvLV has been developed recently based on the TMEM154 gene with validation data from over 2,800 sheep representing nine cohorts. While no single genotype has been shown to have complete resistance to OvLV, consistent association in thousands of sheep from multiple breeds and management conditions highlight a new strategy for intervention by selective breeding. This genetic marker-assisted selection (MAS) has the potential to be a useful addition to existing viral control measures. Further, the discovery of multiple additional genomic regions associated with susceptibility to or control of OvLV suggests that additional genetic marker tests may be developed to extend the reach of MAS in the future. This review will cover the strengths and limitations of existing data from host genetics as an intervention and outline additional questions for future genetic research in sheep, goats, small ruminant lentiviruses, and their host-pathogen interactions. PMID:23771240

  9. Recent patents on biosafety strategies of selectable marker genes in genetically modified crops.

    PubMed

    Jiang, Yiming; Hu, Xiaoning; Huang, Haiying

    2014-01-01

    Genetically modified crops (GMCs) have been planted world wide since 1990s, but the potential insecurity of selectable marker genes raises the questions about GMC safety. Therefore, several researches have been conducted on marker gene safety issues and recently several patents have been issued on this subject. There are two main approaches to achieve this goal: seeking the biosafety selectable marker and eliminating these insecure marker genes after transformation. Results show that these two systems are quite effective. Recent patents on the two ways are discussed in this review.

  10. Transformation system for Hypocrea jecorina (Trichoderma reesei) that favors homologous integration and employs reusable bidirectionally selectable markers.

    PubMed

    Steiger, Matthias G; Vitikainen, Marika; Uskonen, Pekka; Brunner, Kurt; Adam, Gerhard; Pakula, Tiina; Penttilä, Merja; Saloheimo, Markku; Mach, Robert L; Mach-Aigner, Astrid R

    2011-01-01

    Hypocrea jecorina is an industrially important filamentous fungus due to its effective production of hydrolytic enzymes. It has received increasing interest because of its ability to convert lignocellulosic biomass to monomeric sugars, which can be converted into biofuels or platform chemicals. Genetic engineering of strains is a highly important means of meeting the requirements of tailor-made applications. Therefore, we report the development of a transformation system that allows highly efficient gene targeting by using a tmus53 (human LIG4 homolog) deletion strain. Moreover, it permits the unlimited reuse of the same marker by employing a Cre/loxP-based excision system. Both marker insertion and marker excision can be positively selected for by combining resistance to hygromycin B and loss of sensitivity to fluoroacetamide. Finally, the marker pyr4, also positively selectable for insertion and loss, can be used to remove the cre gene.

  11. Marker-assisted selection for biotic stress resistance in peanut

    USDA-ARS?s Scientific Manuscript database

    Peanut ranks second to soybean in the world market trade of oilseeds both in area grown and tonnage produced, and is well-suited to contribute significantly to poverty reduction in the developing world. Peanut is a tetrapoid of recent origin, and has shown low levels of molecular marker polymorphis...

  12. Comparative Analysis of Marker-Assisted and Phenotypic Selection for Yield Components in Cucumber

    USDA-ARS?s Scientific Manuscript database

    Theorectical studies suggest that marker-assisted selection (MAS) has case-specific advantages over phenotypic selection (PHE) for selection of quantitative traits. However, few studies have been conducted that empirically compare these selection methods in the context of a plant breeding program. ...

  13. Marker-Based Hierarchical Segmentation and Classification Approach for Hyperspectral Imagery

    NASA Technical Reports Server (NTRS)

    Tarabalka, Yuliya; Tilton, James C.; Benediktsson, Jon Atli; Chanussot, Jocelyn

    2011-01-01

    The Hierarchical SEGmentation (HSEG) algorithm, which is a combination of hierarchical step-wise optimization and spectral clustering, has given good performances for hyperspectral image analysis. This technique produces at its output a hierarchical set of image segmentations. The automated selection of a single segmentation level is often necessary. We propose and investigate the use of automatically selected markers for this purpose. In this paper, a novel Marker-based HSEG (M-HSEG) method for spectral-spatial classification of hyperspectral images is proposed. First, pixelwise classification is performed and the most reliably classified pixels are selected as markers, with the corresponding class labels. Then, a novel constrained marker-based HSEG algorithm is applied, resulting in a spectral-spatial classification map. The experimental results show that the proposed approach yields accurate segmentation and classification maps, and thus is attractive for hyperspectral image analysis.

  14. Imputation of Unordered Markers and the Impact on Genomic Selection Accuracy

    PubMed Central

    Rutkoski, Jessica E.; Poland, Jesse; Jannink, Jean-Luc; Sorrells, Mark E.

    2013-01-01

    Genomic selection, a breeding method that promises to accelerate rates of genetic gain, requires dense, genome-wide marker data. Genotyping-by-sequencing can generate a large number of de novo markers. However, without a reference genome, these markers are unordered and typically have a large proportion of missing data. Because marker imputation algorithms were developed for species with a reference genome, algorithms suited for unordered markers have not been rigorously evaluated. Using four empirical datasets, we evaluate and characterize four such imputation methods, referred to as k-nearest neighbors, singular value decomposition, random forest regression, and expectation maximization imputation, in terms of their imputation accuracies and the factors affecting accuracy. The effect of imputation method on the genomic selection accuracy is assessed in comparison with mean imputation. The effect of excluding markers with a large proportion of missing data on the genomic selection accuracy is also examined. Our results show that imputation of unordered markers can be accurate, especially when linkage disequilibrium between markers is high and genotyped individuals are related. Of the methods evaluated, random forest regression imputation produced superior accuracy. In comparison with mean imputation, all four imputation methods we evaluated led to greater genomic selection accuracies when the level of missing data was high. Including rather than excluding markers with a large proportion of missing data nearly always led to greater GS accuracies. We conclude that high levels of missing data in dense marker sets is not a major obstacle for genomic selection, even when marker order is not known. PMID:23449944

  15. A discrimination index for selecting markers of tumor growth dynamic across multiple cancer studies with a cure fraction.

    PubMed

    Rouam, Sigrid; Broët, Philippe

    2013-08-01

    To identify genomic markers with consistent effect on tumor dynamics across multiple cancer series, discrimination indices based on proportional hazards models can be used since they do not depend heavily on the sample size. However, the underlying assumption of proportionality of the hazards does not always hold, especially when the studied population is a mixture of cured and uncured patients, like in early-stage cancers. We propose a novel index that quantifies the capability of a genomic marker to separate uncured patients, according to their time-to-event outcomes. It allows to identify genomic markers characterizing tumor growth dynamic across multiple studies. Simulation results show that our index performs better than classical indices based on the Cox model. It is neither affected by the sample size nor the cure rate fraction. In a cross-study of early-stage breast cancers, the index allows to select genomic markers with a potential consistent effect on tumor growth dynamics.

  16. Markers

    ERIC Educational Resources Information Center

    Healthy Schools Network, Inc., 2011

    2011-01-01

    Dry erase whiteboards come with toxic dry erase markers and toxic cleaning products. Dry erase markers labeled "nontoxic" are not free of toxic chemicals and can cause health problems. Children are especially vulnerable to environmental health hazards; moreover, schools commonly have problems with indoor air pollution, as they are more densely…

  17. Whole Genome Sequencing of Elite Rice Cultivars as a Comprehensive Information Resource for Marker Assisted Selection

    PubMed Central

    Duitama, Jorge; Silva, Alexander; Sanabria, Yamid; Cruz, Daniel Felipe; Quintero, Constanza; Ballen, Carolina; Lorieux, Mathias; Scheffler, Brian; Farmer, Andrew; Torres, Edgar; Oard, James; Tohme, Joe

    2015-01-01

    Current advances in sequencing technologies and bioinformatics revealed the genomic background of rice, a staple food for the poor people, and provided the basis to develop large genomic variation databases for thousands of cultivars. Proper analysis of this massive resource is expected to give novel insights into the structure, function, and evolution of the rice genome, and to aid the development of rice varieties through marker assisted selection or genomic selection. In this work we present sequencing and bioinformatics analyses of 104 rice varieties belonging to the major subspecies of Oryza sativa. We identified repetitive elements and recurrent copy number variation covering about 200 Mbp of the rice genome. Genotyping of over 18 million polymorphic locations within O. sativa allowed us to reconstruct the individual haplotype patterns shaping the genomic background of elite varieties used by farmers throughout the Americas. Based on a reconstruction of the alleles for the gene GBSSI, we could identify novel genetic markers for selection of varieties with high amylose content. We expect that both the analysis methods and the genomic information described here would be of great use for the rice research community and for other groups carrying on similar sequencing efforts in other crops. PMID:25923345

  18. Charles W. Stuber: Maize geneticist and pioneer of marker-assisted selection

    USDA-ARS?s Scientific Manuscript database

    Charles W. Stuber is considered a pioneer of quantitative genetic mapping and marker-assisted selection in maize. The achievements of his four decade career in research include the development of genetic marker systems used in maize and adapted in many other crops, the first methods and studies to i...

  19. Selection enhanced estimates of marker effects on means and variances of beef tenderness

    USDA-ARS?s Scientific Manuscript database

    Genetic marker associations from surveys of industry cattle populations have low frequencies of rare homozygous animals. Selection for calpain (CAPN1) and calpastatin (CAST) genetic markers was replicated in two cattle populations (Angus and MARC III) at the U.S. Meat Animal Research Center. These...

  20. Impact of marker ascertainment bias on genomic selection accuracy and estimates of genetic diversity

    USDA-ARS?s Scientific Manuscript database

    Genome-wide molecular markers are readily being applied to evaluate genetic diversity in germplasm collections and for making genomic selections in breeding programs. To accurately predict phenotypes and assay genetic diversity, molecular markers should assay a representative sample of the polymorp...

  1. Development and Validation of High-Resolution Melting Markers Derived from Rysto STS Markers for High-Throughput Marker-Assisted Selection of Potato Carrying Rysto.

    PubMed

    Nie, Xianzhou; Sutherland, Darcy; Dickison, Virginia; Singh, Mathuresh; Murphy, Agnes M; De Koeyer, David

    2016-11-01

    Sequence analysis of the chromosome region harboring the sequence-tagged site (STS) markers YES3-3A and YES3-3B for Rysto, a gene responsible for extreme resistance to Potato virus Y (PVY) in potato, was performed in tetraploid potato 'Barbara' (Rrrr) and 'AC Chaleur' (rrrr) as well as their progeny selections. Three and two sequence variants were identified in Barbara resistant (R) selections and AC Chaleur susceptible (S) selections, respectively. Further analysis indicates that the variant with a 21-nucleotide (nt) deletion is likely the chromosome copy harboring the STS markers. Two primer pairs, one targeting the region containing a 20-nt deletion and the other targeting the region anchoring the YES3-3A reverse primer, were designed. As anticipated, pair one produced two visible fragments in Barbara-R bulk and one visible fragment in AC Chaleur-S bulk; pair two produced one visible fragment in all samples. When subjected to high-resolution melting (HRM) analysis, two distinct melting profiles for R and S samples were observed. Analysis of 147 progeny of Barbara × AC Chaleur revealed 72 and 75 progeny with R and S melting profiles, respectively, which was consistent with YES3-3A and YES3-3B assays and phenotyping analysis, thus demonstrating the potential of HRM profiles as novel molecular markers for Rysto. The efficacy of the newly developed HRM markers for high-throughput marker-assisted selection for Rysto-conferred resistance to PVY was validated further with three populations involving Barbara as the R parent.

  2. Evolution of the polymorphism at molecular markers in QTL and non-QTL regions in selected chicken lines (Open Access publication)

    PubMed Central

    Loywyck, Valérie; Bed'hom, Bertrand; Pinard-van der Laan, Marie-Hélène; Pitel, Frédérique; Verrier, Étienne; Bijma, Piter

    2008-01-01

    We investigated the joint evolution of neutral and selected genomic regions in three chicken lines selected for immune response and in one control line. We compared the evolution of polymorphism of 21 supposedly neutral microsatellite markers versus 30 microsatellite markers located in seven quantitative trait loci (QTL) regions. Divergence of lines was observed by factor analysis. Five supposedly neutral markers and 12 markers in theQTL regions showed Fst values greater than 0.15. However, the non-significant difference (P > 0.05) between matrices of genetic distances based on genotypes at supposedly neutral markers on the one hand, and at markers in QTL regions, on the other hand, showed that none of the markers in the QTL regions were influenced by selection. A supposedly neutral marker and a marker located in the QTL region on chromosome 14 showed temporal variations in allele frequencies that could not be explained by drift only. Finally, to confirm thatmarkers located inQTL regions on chromosomes 1, 7 and 14were under the influence of selection, simulations were performed using haplotype dropping along the existing pedigree. In the zone located on chromosome 14, the simulation results confirmed that selection had an effect on the evolution of polymorphism of markers within the zone. PMID:18990356

  3. A Novel Selection Marker for Efficient DNA Cloning and Recombineering in E. coli

    PubMed Central

    Jang, Chuan-Wei; Magnuson, Terry

    2013-01-01

    Production of recombinant DNA in bacterial cells is an essential technique in molecular biology. Plasmids are usually maintained in an E. coli host by antibiotic selection. However, there are only a few antibiotic-resistance markers available in common use. Here we report the adoption of a novel selection marker, mfabI (mutant fabI) for plasmid propagation in E. coli. mfabI expands the limited repertoire of selection markers and allows for more efficient molecular manipulation and plasmid propagation in E. coli. We show that mfabI is not only an efficient plasmid selection marker, but it also possesses unique activity that may facilitate molecular manipulation of unstable sequences. Furthermore, we have incorporated mfabI in the recombineering tool kit for generating mouse gene targeting vectors and demonstrate the advantage of using mfabI-containing recombineering vectors. PMID:23437314

  4. Dominant selectable markers for Penicillium spp. transformation and gene function studies

    USDA-ARS?s Scientific Manuscript database

    Penicillium spp. has been genetically manipulated and gene function studies have utilized single gene deletion strains for phenotypic analysis. Fungal transformation experiments have relied on hygromycin and hygromycin phosphotransferase (hph) as the main dominant selectable marker (DSM) system in P...

  5. Neural and cognitive face-selective markers: An integrative review.

    PubMed

    Yovel, Galit

    2016-03-01

    Faces elicit robust and selective neural responses in the primate brain. These neural responses have been investigated with functional MRI and EEG in numerous studies, which have reported face-selective activations in the occipital-temporal cortex and an electrophysiological face-selective response that peaks 170 ms after stimulus onset at occipital-temporal sites. Evidence for face-selective processes has also been consistently reported in cognitive studies, which investigated the face inversion effect, the composite face effect and the left visual field (LVF) superiority. These cognitive effects indicate that the perceptual representation that we generate for faces differs from the representation that is generated for inverted faces or non-face objects. In this review, I will show that the fMRI and ERP face-selective responses are strongly associated with these three well-established behavioral face-selective measures. I will further review studies that examined the relationship between fMRI and EEG face-selective measures suggesting that they are strongly linked. Taken together these studies imply that a holistic representation of a face is generated at 170 ms after stimulus onset over the right hemisphere. These findings, which reveal a strong link between the various and complementary cognitive and neural measures of face processing, allow to characterize where, when and how faces are represented during the first 200 ms of face processing. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. One Cycle of Phenotypic Selection Combined with Marker Assisted Selection for Improving Yield and Quality in Cucumber

    USDA-ARS?s Scientific Manuscript database

    Estimation of response to marker-assisted selection (MAS) is critical to breeding programs. To investigate MAS for line development, two cucumber recombinant inbred lines (RILs) (7026B76 and 7022C8) differing in plant habit were mated, and F4 and F5 progeny was MAS selected to improve both yield an...

  7. Determination of Genetic Diversity Using 15 Simple Sequence Repeats Markers in Long Term Selected Japanese Quail Lines

    PubMed Central

    Karabağ, Kemal; Balcıoğlu, Murat Soner; Karlı, Taki; Alkan, Sezai

    2016-01-01

    Japanese quail is still used as a model for poultry research because of their usefulness as laying, meat, and laboratory animals. Microsatellite markers are the most widely used molecular markers, due to their relative ease of scoring and high levels of polymorphism. The objective of the research was to determine genetic diversity and population genetic structures of selected Japanese quail lines (high body weight 1 [HBW1], HBW2, low body weight [LBW], and layer [L]) throughout 15th generations and an unselected control (C). A total of 69 individuals from five quail lines were genotyped by fifteen microsatellite markers. When analyzed profiles of the markers the observed (Ho) and expected (He) heterozygosity ranged from 0.04 (GUJ0027) to 0.64 (GUJ0087) and 0.21 (GUJ0027) to 0.84 (GUJ0037), respectively. Also, Ho and He were separated from 0.30 (L and LBW) to 0.33 (C and HBW2) and from 0.52 (HBW2) to 0.58 (L and LBW), respectively. The mean polymorphic information content (PIC) ranged from 0.46 (HBW2) to 0.52 (L). Approximately half of the markers were informative (PIC≥0.50). Genetic distances were calculated from 0.09 (HBW1 and HBW2) to 0.33 (C and L). Phylogenetic dendrogram showed that the quail lines were clearly defined by the microsatellite markers used here. Bayesian model-based clustering supported the results from the phylogenetic tree. These results reflect that the set of studied markers can be used effectively to capture the magnitude of genetic variability in selected Japanese quail lines. Also, to identify markers and alleles which are specific to the divergence lines, further generations of selection are required. PMID:27165027

  8. Empirical Selection of Informative Microsatellite Markers within Co-ancestry Pig Populations Is Required for Improving the Individual Assignment Efficiency.

    PubMed

    Li, Y H; Chu, H P; Jiang, Y N; Lin, C Y; Li, S H; Li, K T; Weng, G J; Cheng, C C; Lu, D J; Ju, Y T

    2014-05-01

    The Lanyu is a miniature pig breed indigenous to Lanyu Island, Taiwan. It is distantly related to Asian and European pig breeds. It has been inbred to generate two breeds and crossed with Landrace and Duroc to produce two hybrids for laboratory use. Selecting sets of informative genetic markers to track the genetic qualities of laboratory animals and stud stock is an important function of genetic databases. For more than two decades, Lanyu derived breeds of common ancestry and crossbreeds have been used to examine the effectiveness of genetic marker selection and optimal approaches for individual assignment. In this paper, these pigs and the following breeds: Berkshire, Duroc, Landrace and Yorkshire, Meishan and Taoyuan, TLRI Black Pig No. 1, and Kaohsiung Animal Propagation Station Black pig are studied to build a genetic reference database. Nineteen microsatellite markers (loci) provide information on genetic variation and differentiation among studied breeds. High differentiation index (FST) and Cavalli-Sforza chord distances give genetic differentiation among breeds, including Lanyu's inbred populations. Inbreeding values (FIS) show that Lanyu and its derived inbred breeds have significant loss of heterozygosity. Individual assignment testing of 352 animals was done with different numbers of microsatellite markers in this study. The testing assigned 99% of the animals successfully into their correct reference populations based on 9 to 14 markers ranking D-scores, allelic number, expected heterozygosity (HE) or FST, respectively. All miss-assigned individuals came from close lineage Lanyu breeds. To improve individual assignment among close lineage breeds, microsatellite markers selected from Lanyu populations with high polymorphic, heterozygosity, FST and D-scores were used. Only 6 to 8 markers ranking HE, FST or allelic number were required to obtain 99% assignment accuracy. This result suggests empirical examination of assignment-error rates is required if

  9. A CAPS marker to assist selection of tomato spotted wilt virus (TSWV) resistance in pepper.

    PubMed

    Moury, B; Pflieger, S; Blattes, A; Lefebvre, V; Palloix, A

    2000-02-01

    The hypersensitive resistance to tomato spotted wilt virus (TSWV) in pepper is determined by a single dominant gene (resistant allele: Tsw) in several Capsicum chinense genotypes. In order to facilitate the selection for this resistance, four RAPD (among 250 10-mer primers tested) were found linked to the Tsw locus using the bulked segregant analysis and 153 F2 individuals. A close RAPD marker was converted into a codominant cleaved amplified polymorphic sequence (CAPS) using specific PCR primers and restriction enzymes. This CAPS marker is tightly linked to Tsw (0.9 +/- 0.6 cM) and is helpful for marker-assisted selection in a wide range of genetic intercrosses.

  10. ISSR marker-assisted genetic diversity analysis of Dioscorea hispida and selection of the best variety for sustainable production.

    PubMed

    Nudin, Nur Fatihah Hasan; Ali, Abdul Manaf; Ngah, Norhayati; Mazlan, Nor Zuhailah; Mat, Nashriyah; Ghani, Mohd Noor Abd; Alias, Nadiawati; Zakaria, Abd Jamil; Jahan, Md Sarwar

    2017-09-06

    Plant breeding is a way of selection of a particular individual for the production of the progeny by separating or combining desired characteristics. The objective of this study was to justify different characteristics of Dioscorea hispida (Ubi gadong) varieties using molecular techniques to select the best variety for sustainable production at the farmer's level. A total of 160 germplasms of Ubi gadong were collected from different locations at the Terengganu and Kelantan states of Malaysia. Forty eight (48) out of 160 germplasms were selected as "primary" selection based on yield and other qualitative characters. Selected collections were then grown and maintained for ISSR marker-assisted genetic diversity analysis. Overall plant growth and yield of tubers were also determined. A total of 12 ISSR markers were tested to justify the characteristics of Ubi gadong varieties among which three markers showed polymorphic bands and on average 57.3% polymorphism were observed representing the highest variation among germplasms. The ISSR marker based on UPGMA cluster analysis grouped all 48 D. hispida into 10 vital groups that proved a vast genetic variation among germplasm collections. Therefore, hybridization should be made between two distant populations. The D. hispida is already proved as the highest starch content tuber crops and very rich in vitamins with both micro and macro minerals. Considering all these criteria and results from marker-assisted diversity analysis, accessions that are far apart based on their genetic coefficient (like DH27 and DH71; DH30 and DH70; DH43 and DH62; DH45 and DH61; DH77 and DH61; DH78 and DH57) could be selected as parents for further breeding programs. This will bring about greater diversity, which will lead to high productive index in terms of increase in yield and overall quality and for the ultimate target of sustainable Ubi gadong production. Copyright © 2017 Académie des sciences. Published by Elsevier Masson SAS. All rights

  11. Plasmid selection in Escherichia coli using an endogenous essential gene marker

    PubMed Central

    Goh, Shan; Good, Liam

    2008-01-01

    Background Antibiotic resistance genes are widely used for selection of recombinant bacteria, but their use risks contributing to the spread of antibiotic resistance. In particular, the practice is inappropriate for some intrinsically resistant bacteria and in vaccine production, and costly for industrial scale production. Non-antibiotic systems are available, but require mutant host strains, defined media or expensive reagents. An unexplored concept is over-expression of a host essential gene to enable selection in the presence of a chemical inhibitor of the gene product. To test this idea in E. coli, we used the growth essential target gene fabI as the plasmid-borne marker and the biocide triclosan as the selective agent. Results The new cloning vector, pFab, enabled selection by triclosan at 1 μM. Interestingly, pFab out-performed the parent pUC19-ampicillin system in cell growth, plasmid stability and plasmid yield. Also, pFab was toxic to host cells in a way that was reversed by triclosan. Therefore, pFab and triclosan are toxic when used alone but in combination they enhance growth and plasmid production through a gene-inhibitor interaction. Conclusion The fabI-triclosan model system provides an alternative plasmid selection method based on essential gene over-expression, without the use of antibiotic-resistance genes and conventional antibiotics. PMID:18694482

  12. Evaluating gene flow using selected markers: a case study.

    PubMed Central

    Lenormand, T; Guillemaud, T; Bourguet, D; Raymond, M

    1998-01-01

    The extent to which an organism is locally adapted in an environmental pocket depends on the selection intensities inside and outside the pocket, on migration, and on the size of the pocket. When two or more loci are involved in this local adaptation, measuring their frequency gradients and their linkage disequilbria allows one to disentangle the forces-migration and selection-acting on the system. We apply this method to the case of a local adaptation to organophosphate insecticides in the mosquito Culex pipiens pipiens in southern France. The study of two different resistance loci allowed us to estimate with support limits gene flow as well as selection pressure on insecticide resistance and the fitness costs associated with each locus. These estimates permit us to pinpoint the conditions for the maintenance of this pocket of adaptation as well as the effect of the interaction between the two resistance loci. PMID:9649528

  13. Dinitrochlorobenzene contact hypersensitivity as a marker trait for selection to improve disease resistance in calves.

    PubMed

    Burton, J L; Kennedy, B W; Burnside, E B; Wilkie, B N; Burton, J H

    1989-09-01

    Measurements of double skin fold thickness (mm) were used to assess the feasibility of use of cutaneous delayed-type hypersensitivity response as a marker trait to select for genetically improved disease resistance. Data were from a sample of 149 7-wk-old Holstein calves, from 15 sires, that previously had been sensitized to the contact allergen dinitrochlorobenzene. Response was monitored 24 and 48 h postchallenge. Analysis was by least squares according to a model that included fixed managerial effects, disease prevalence, and severity categories and sire of calf. Double skin fold thickness increased about 50% by 24 h, followed by a 4% decrease between 24 and 48 h. Season of birth was the only consistently significant fixed effect. Calves born in fall showed larger skin changes at each stage of response than calves born in spring. Restricted maximum likelihood estimates of paternal half-sib heritability were moderate to high for all stages of response, and phenotypic and genetic correlations between stages were significant and positive. Although the prevalence and severity of naturally occurring pneumonia and diarrhea did not significantly affect quantity of response, there is need to study further this relationship for intracellular pathogens. Delayed-type hypersensitivity response to dinitrochlorobenzene can meet requirements for a successful marker on which to base sire selection for disease resistance, if it is related to economically important, intracellularly characterized, pathogenic bovine diseases.

  14. FastTagger: an efficient algorithm for genome-wide tag SNP selection using multi-marker linkage disequilibrium

    PubMed Central

    2010-01-01

    Background Human genome contains millions of common single nucleotide polymorphisms (SNPs) and these SNPs play an important role in understanding the association between genetic variations and human diseases. Many SNPs show correlated genotypes, or linkage disequilibrium (LD), thus it is not necessary to genotype all SNPs for association study. Many algorithms have been developed to find a small subset of SNPs called tag SNPs that are sufficient to infer all the other SNPs. Algorithms based on the r2 LD statistic have gained popularity because r2 is directly related to statistical power to detect disease associations. Most of existing r2 based algorithms use pairwise LD. Recent studies show that multi-marker LD can help further reduce the number of tag SNPs. However, existing tag SNP selection algorithms based on multi-marker LD are both time-consuming and memory-consuming. They cannot work on chromosomes containing more than 100 k SNPs using length-3 tagging rules. Results We propose an efficient algorithm called FastTagger to calculate multi-marker tagging rules and select tag SNPs based on multi-marker LD. FastTagger uses several techniques to reduce running time and memory consumption. Our experiment results show that FastTagger is several times faster than existing multi-marker based tag SNP selection algorithms, and it consumes much less memory at the same time. As a result, FastTagger can work on chromosomes containing more than 100 k SNPs using length-3 tagging rules. FastTagger also produces smaller sets of tag SNPs than existing multi-marker based algorithms, and the reduction ratio ranges from 3%-9% when length-3 tagging rules are used. The generated tagging rules can also be used for genotype imputation. We studied the prediction accuracy of individual rules, and the average accuracy is above 96% when r2 ≥ 0.9. Conclusions Generating multi-marker tagging rules is a computation intensive task, and it is the bottleneck of existing multi-marker based tag

  15. The use of marker-assisted selection in animal breeding and biotechnology.

    PubMed

    Williams, J L

    2005-04-01

    Improvement of livestock has focused on the selective breeding of individuals with superior phenotypes. With the development of increasingly advanced statistical methods that maximise selection for genetic gain, this simple approach has been extremely successful in increasing the quantity of agricultural output. However, information now available on the organisation and functioning of the genome could be used in breeding programmes to improve a range of traits. Many traits are under the control of several genetic loci, each of which contribute to the variation in the trait and hence are called quantitative trait loci (QTL). While genetic markers for QTL that are linked to the trait gene could be used to choose animals for selective breeding programmes, the most effective markers are the functional mutations within the trait genes. Strategies to identify markers for traits and the application of these markers are described by reference to examples of loci that control a range of different traits.

  16. Comparing the predictive abilities of phenotypic and marker-assisted selection methods in a biparental lettuce population

    USDA-ARS?s Scientific Manuscript database

    Breeding and selection for the traits with polygenic inheritance is a challenging task that can be done by phenotypic selection, by marker-assisted selection or by genome wide selection. We tested predictive ability of four selection models in a biparental population genotyped with 95 SNP markers an...

  17. Puromycin-N-acetyltransferase as a selectable marker for use in Plasmodium falciparum.

    PubMed

    de Koning-Ward, T F; Waters, A P; Crabb, B S

    2001-10-01

    The limited number of selectable markers available for malaria transfection has hindered extensive manipulation of the Plasmodium falciparum genome and subsequently thorough genetic analysis of this organism. In this paper, we demonstrate that P. falciparum is highly sensitive to the drug puromycin, but that transgenic expression of the puromycin-N-acetyltransferase (PAC) gene from Streptomyces alboninger confers resistance to this drug with the IC(50) and IC(90) values increasing approximately 3- and 7-fold, respectively in PAC-expressing parasites. Despite this relatively low level of resistance, parasite populations transfected with the PAC selectable marker and selected directly on puromycin emerged at the same rate post-transfection as human dihydrofolate reductase (hDHFR)-expressing parasites, selected independently with the anti-folate drug WR99210. Transfected parasites generally maintained the PAC expression plasmid episomally at between two and six copies per parasite. We also demonstrate by cycling transfected parasites in the presence and absence of puromycin for several weeks, that the PAC selectable marker can be used for gene-targeting. Since the mode of action of puromycin is distinct from other drugs currently used for the stable transfection of P. falciparum, the PAC selectable marker should also have applicability for use in conjunction with other positive selectable markers, thereby increasing the possibilities for more complex functional studies of this organism.

  18. Heliocides as markers in selection of insect resistant cotton varieties

    USDA-ARS?s Scientific Manuscript database

    Development of insect resistance cotton varieities is an important goal in cotton genetics and in making selections in breeding programs. Presently, the use of genetically modified cottons containing Bt genes derived from the soil bacterium Bacillus thurengensis, are extensively utilized. An alter...

  19. A system for multi-locus chromosomal integration and transformation-free selection marker rescue

    PubMed Central

    Siddiqui, Michael S.; Choksi, Atri; Smolke, Christina D.

    2014-01-01

    Yeast integrating plasmids (YIPs) are a versatile tool for stable integration in Saccharomyces cerevisiae. However, current YIP systems necessitate time- and labor-intensive methods for cloning and selection marker rescue. Here we describe the design, construction, and validation of a new YIP system capable of accelerating the stable integration of multiple expression constructs into different loci in the yeast S. cerevisiae. These “directed pop-out” plasmids enable a simple, two-step integration protocol that results in a scarless integration alongside a complete rescue of the selection marker. These plasmids combine three key features: a dedicated “YIPout” fragment directs a recombination event that rescues the selection marker while avoiding undesired excision of the target DNA sequence, a multi-fragment modular DNA assembly system simplifies cloning, and a new set of counterselectable markers enables serial integration followed by a transformation-free marker rescue event. We constructed and tested directed pop-out YIPs for integration of fluorescent reporter genes into four yeast loci. We validated our new YIP design by integrating three reporter genes into three different loci with transformation-free rescue of selection markers. These new YIP designs will facilitate the construction of yeast strains that express complex heterologous metabolic pathways. PMID:25226817

  20. Financial feasibility of marker-aided selection in Douglas-fir.

    Treesearch

    G.R. Johnson; N.C. Wheeler; S.H. Strauss

    2000-01-01

    The land area required for a marker-aided selection (MAS) program to break-even (i.e., have equal costs and benefits) was estimated using computer simulation for coastal Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) in the Pacific Northwestern United States. We compared the selection efficiency obtained when using an index that included the...

  1. Construction of two selectable markers for integrative/conjugative plasmids in Flavobacterium columnare

    NASA Astrophysics Data System (ADS)

    Zhang, Jin; Zou, Hong; Wang, Liangfa; Huang, Bei; Li, Nan; Wang, Guitang; Nie, Pin

    2012-03-01

    Flavobacterium columnare, the etiological agent of columnaris disease, is one of the most important and widespread bacterial pathogens of freshwater fish. In this study, we constructed two artificial selectable markers (chloramphenicol and spectinomycin resistance) for gene transfer in F. columnare. These two new artificial selectable markers, which were created by placing the chloramphenicol or spectinomycin resistance gene under the control of the native acs regulatory region of F. columnare, were functional in both F. columnare and Escherichia coli. The integrative/conjugative plasmids constructed by using these markers were introduced into F. columnare G4 via electroporation or conjugation. The integrated plasmid DNA was confirmed by Southern blotting and PCR analysis. These two markers can be employed in future investigations into gene deletion and the pathogenicity of virulence factors in F. columnare.

  2. A method for selection of restriction enzymes for sdCAPS marker construction

    USDA-ARS?s Scientific Manuscript database

    Development of PCR-based markers for SNP detection is prerequisite for various genetic analyses. The use of restriction enzymes following PCR amplification is a common and relatively low cost method for SNP detection. Simple and cost-effective methodologies for SNP marker development that would en...

  3. Use of the pyrG gene as a food-grade selection marker in Monascus.

    PubMed

    Wang, Bo-hua; Xu, Yang; Li, Yan-ping

    2010-11-01

    Ma-pyrG was cloned from Monascus aurantiacus AS3.4384 using degenerate PCR with primers designed with an algorithm called CODEHOP, and its complete sequence was obtained by a PCR-based strategy for screening a Monascus fosmid library. Ma-pyrG encodes orotidine-5'-phosphate decarboxylase (OMPdecase), a 283-aminoacid protein with 81% sequence identity to that from Aspergillus flavus NRRL 3357. A pyrG mutant strain from M. aurantiacus AS3.4384, named UM28, was isolated by resistance to 5-fluoroorotic acid after UV mutagenesis. Sequence analysis of this mutated gene revealed that it contained a point mutation at nucleotide position +220. Plasmid pGFP-pyrG, bearing the green fluorescent protein gene (GFP) as a model gene and Ma-pyrG as a selection marker, were constructed. pGFP-pyrG were successfully transformed into UM28 by using the PEG method.

  4. Efficient transformation of wheat by using a mutated rice acetolactate synthase gene as a selectable marker.

    PubMed

    Ogawa, Taiichi; Kawahigashi, Hiroyuki; Toki, Seiichi; Handa, Hirokazu

    2008-08-01

    Acetolactate synthase (ALS) is a target enzyme for many herbicides, including sulfonylurea and imidazolinone. We investigated the usefulness of a mutated ALS gene of rice, which had double point mutations and encoded an herbicide-resistant form of the enzyme, as a selectable marker for wheat transformation. After the genomic DNA fragment from rice containing the mutated ALS gene was introduced into immature embryos by means of particle bombardment, transgenic plants were efficiently selected with the herbicide bispyribac sodium (BS). Southern blot analysis confirmed that transgenic plants had one to more than ten copies of the transgene in their chromosomes. Adjustment of the BS concentration combined with repeated selection effectively prevented nontransgenic plants from escaping herbicide selection. Measurement of ALS activity indicated that transgenic plants produced an herbicide-resistant form of ALS and therefore had acquired the resistance to BS. This report is the first to describe a selection system for wheat transformation that uses a selectable marker gene of plant origin.

  5. Rational optimization of tolC as a powerful dual selectable marker for genome engineering

    PubMed Central

    Gregg, Christopher J.; Lajoie, Marc J.; Napolitano, Michael G.; Mosberg, Joshua A.; Goodman, Daniel B.; Aach, John; Isaacs, Farren J.; Church, George M.

    2014-01-01

    Selection has been invaluable for genetic manipulation, although counter-selection has historically exhibited limited robustness and convenience. TolC, an outer membrane pore involved in transmembrane transport in E. coli, has been implemented as a selectable/counter-selectable marker, but counter-selection escape frequency using colicin E1 precludes using tolC for inefficient genetic manipulations and/or with large libraries. Here, we leveraged unbiased deep sequencing of 96 independent lineages exhibiting counter-selection escape to identify loss-of-function mutations, which offered mechanistic insight and guided strain engineering to reduce counter-selection escape frequency by ∼40-fold. We fundamentally improved the tolC counter-selection by supplementing a second agent, vancomycin, which reduces counter-selection escape by 425-fold, compared colicin E1 alone. Combining these improvements in a mismatch repair proficient strain reduced counter-selection escape frequency by 1.3E6-fold in total, making tolC counter-selection as effective as most selectable markers, and adding a valuable tool to the genome editing toolbox. These improvements permitted us to perform stable and continuous rounds of selection/counter-selection using tolC, enabling replacement of 10 alleles without requiring genotypic screening for the first time. Finally, we combined these advances to create an optimized E. coli strain for genome engineering that is ∼10-fold more efficient at achieving allelic diversity than previous best practices. PMID:24452804

  6. Cellulose-lanthanum hydroxide nanocomposite as a selective marker for detection of toxic copper

    PubMed Central

    2014-01-01

    In this current report, a simple, reliable, and rapid method based on modifying the cellulose surface by doping it with different percentages of lanthanum hydroxide (i.e., 1% La(OH)3-cellulose (LC), 5% La(OH)3-cellulose (LC2), and 10% La(OH)3-cellulose (LC3)) was proposed as a selective marker for detection of copper (Cu(II)) in aqueous medium. Surface properties of the newly modified cellulose phases were confirmed by Fourier transform infrared spectroscopy, field emission scanning electron microscope, energy dispersive X-ray spectroscopy, X-ray diffraction, and X-ray photoelectron spectroscopic analysis. The effect of pH on the adsorption of modified cellulose phases for Cu(II) was evaluated, and LC3 was found to be the most selective for Cu(II) at pH 6.0. Other parameters, influencing the maximum uptake of Cu(II) on LC3, were also investigated for a deeper mechanistic understanding of the adsorption phenomena. Results showed that the adsorption capacity for Cu(II) was improved by 211% on the LC3 phase as compared to diethylaminoethyl cellulose phase after only 2 h contact time. Adsorption isotherm data established that the adsorption process nature was monolayer with a homogeneous adsorbent surface. Results displayed that the adsorption of Cu(II) onto the LC3 phase obeyed a pseudo-second-order kinetic model. Selectivity studies toward eight metal ions, i.e., Cd(II), Co(II), Cr(III), Cr(VI), Cu(II), Fe(III), Ni(II), and Zn(II), were further performed at the optimized pH value. Based on the selectivity study, it was found that Cu(II) is highly selective toward the LC3 phase. Moreover, the efficiency of the proposed method was supported by implementing it to real environmental water samples with adequate results. PMID:25258599

  7. Cellulose-lanthanum hydroxide nanocomposite as a selective marker for detection of toxic copper

    NASA Astrophysics Data System (ADS)

    Marwani, Hadi M.; Lodhi, Mazhar Ullah; Khan, Sher Bahadar; Asiri, Abdullah M.

    2014-09-01

    In this current report, a simple, reliable, and rapid method based on modifying the cellulose surface by doping it with different percentages of lanthanum hydroxide (i.e., 1% La(OH)3-cellulose (LC), 5% La(OH)3-cellulose (LC2), and 10% La(OH)3-cellulose (LC3)) was proposed as a selective marker for detection of copper (Cu(II)) in aqueous medium. Surface properties of the newly modified cellulose phases were confirmed by Fourier transform infrared spectroscopy, field emission scanning electron microscope, energy dispersive X-ray spectroscopy, X-ray diffraction, and X-ray photoelectron spectroscopic analysis. The effect of pH on the adsorption of modified cellulose phases for Cu(II) was evaluated, and LC3 was found to be the most selective for Cu(II) at pH 6.0. Other parameters, influencing the maximum uptake of Cu(II) on LC3, were also investigated for a deeper mechanistic understanding of the adsorption phenomena. Results showed that the adsorption capacity for Cu(II) was improved by 211% on the LC3 phase as compared to diethylaminoethyl cellulose phase after only 2 h contact time. Adsorption isotherm data established that the adsorption process nature was monolayer with a homogeneous adsorbent surface. Results displayed that the adsorption of Cu(II) onto the LC3 phase obeyed a pseudo-second-order kinetic model. Selectivity studies toward eight metal ions, i.e., Cd(II), Co(II), Cr(III), Cr(VI), Cu(II), Fe(III), Ni(II), and Zn(II), were further performed at the optimized pH value. Based on the selectivity study, it was found that Cu(II) is highly selective toward the LC3 phase. Moreover, the efficiency of the proposed method was supported by implementing it to real environmental water samples with adequate results.

  8. Cereal Crop Proteomics: Systemic Analysis of Crop Drought Stress Responses Towards Marker-Assisted Selection Breeding

    PubMed Central

    Ghatak, Arindam; Chaturvedi, Palak; Weckwerth, Wolfram

    2017-01-01

    Sustainable crop production is the major challenge in the current global climate change scenario. Drought stress is one of the most critical abiotic factors which negatively impact crop productivity. In recent years, knowledge about molecular regulation has been generated to understand drought stress responses. For example, information obtained by transcriptome analysis has enhanced our knowledge and facilitated the identification of candidate genes which can be utilized for plant breeding. On the other hand, it becomes more and more evident that the translational and post-translational machinery plays a major role in stress adaptation, especially for immediate molecular processes during stress adaptation. Therefore, it is essential to measure protein levels and post-translational protein modifications to reveal information about stress inducible signal perception and transduction, translational activity and induced protein levels. This information cannot be revealed by genomic or transcriptomic analysis. Eventually, these processes will provide more direct insight into stress perception then genetic markers and might build a complementary basis for future marker-assisted selection of drought resistance. In this review, we survey the role of proteomic studies to illustrate their applications in crop stress adaptation analysis with respect to productivity. Cereal crops such as wheat, rice, maize, barley, sorghum and pearl millet are discussed in detail. We provide a comprehensive and comparative overview of all detected protein changes involved in drought stress in these crops and have summarized existing knowledge into a proposed scheme of drought response. Based on a recent proteome study of pearl millet under drought stress we compare our findings with wheat proteomes and another recent study which defined genetic marker in pearl millet. PMID:28626463

  9. Selection of highly informative SNP markers for population affiliation of major US populations.

    PubMed

    Zeng, Xiangpei; Chakraborty, Ranajit; King, Jonathan L; LaRue, Bobby; Moura-Neto, Rodrigo S; Budowle, Bruce

    2016-03-01

    Ancestry informative markers (AIMs) can be used to detect and adjust for population stratification and predict the ancestry of the source of an evidence sample. Autosomal single nucleotide polymorphisms (SNPs) are the best candidates for AIMs. It is essential to identify the most informative AIM SNPs across relevant populations. Several informativeness measures for ancestry estimation have been used for AIMs selection: absolute allele frequency differences (δ), F statistics (F ST), and informativeness for assignment measure (In). However, their efficacy has not been compared objectively, particularly for determining affiliations of major US populations. In this study, these three measures were directly compared for AIMs selection among four major US populations, i.e., African American, Caucasian, East Asian, and Hispanic American. The results showed that the F ST panel performed slightly better for population resolution based on principal component analysis (PCA) clustering than did the δ panel and both performed better than the In panel. Therefore, the 23 AIMs selected by the F ST measure were used to characterize the four major American populations. Genotype data of nine sample populations were used to evaluate the efficiency of the 23-AIMs panel. The results indicated that individuals could be correctly assigned to the major population categories. Our AIMs panel could contribute to the candidate pool of AIMs for potential forensic identification purposes.

  10. Glyphosate-tolerant CP4 and GOX genes as a selectable marker in wheat transformation.

    PubMed

    Zhou, H; Arrowsmith, J W; Fromm, M E; Hironaka, C M; Taylor, M L; Rodriguez, D; Pajeau, M E; Brown, S M; Santino, C G; Fry, J E

    1995-12-01

    The lack of alternative selectable markers in crop transformation has been a substantial barrier for commercial application of agricultural biotechnology. We have developed an efficient selection system for wheat transformation using glyphosate-tolerant CP4 and GOX genes as a selectable marker. Immature embryos of the wheat cultivar Bobwhite were bombarded with two separate plasmids harboring the CP4/GOX and GUS genes. After a 1 week delay, the bombarded embryos were transferred to a selection medium containing 2 mM glyphosate. Embryo-derived calli were subcultured onto the same selection medium every 3 weeks consecutively for 9-12 weeks, and were then regenerated and rooted on selection media with lower glyphosate concentrations. Transgenic plants tolerant to glyphosate were recovered. ELISA assay confirmed expression of the CP4 and GOX genes in R0 plants. Southern blot analysis demonstrated that the transgenes were integrated into the wheat genomes and transmitted to the following generation. The use of CP4 and GOX genes as a selectable marker provides an efficient, effective, and alternative transformation selection system for wheat.

  11. A diphtheria toxin resistance marker for in vitro and in vivo selection of stably transduced human cells.

    PubMed

    Picco, Gabriele; Petti, Consalvo; Trusolino, Livio; Bertotti, Andrea; Medico, Enzo

    2015-09-30

    We developed a selectable marker rendering human cells resistant to Diphtheria Toxin (DT). The marker (DT(R)) consists of a primary microRNA sequence engineered to downregulate the ubiquitous DPH2 gene, a key enzyme for the biosynthesis of the DT target diphthamide. DT(R) expression in human cells invariably rendered them resistant to DT in vitro, without altering basal cell growth. DT(R)-based selection efficiency and stability were comparable to those of established drug-resistance markers. As mice are insensitive to DT, DT(R)-based selection can be also applied in vivo. Direct injection of a GFP-DT(R) lentiviral vector into human cancer cell-line xenografts and patient-derived tumorgrafts implanted in mice, followed by systemic DT administration, yielded tumors entirely composed of permanently transduced cells and detectable by imaging systems. This approach enabled high-efficiency in vivo selection of xenografted human tumor tissues expressing ectopic transgenes, a hitherto unmet need for functional and morphological studies in laboratory animals.

  12. A diphtheria toxin resistance marker for in vitro and in vivo selection of stably transduced human cells

    PubMed Central

    Picco, Gabriele; Petti, Consalvo; Trusolino, Livio; Bertotti, Andrea; Medico, Enzo

    2015-01-01

    We developed a selectable marker rendering human cells resistant to Diphtheria Toxin (DT). The marker (DTR) consists of a primary microRNA sequence engineered to downregulate the ubiquitous DPH2 gene, a key enzyme for the biosynthesis of the DT target diphthamide. DTR expression in human cells invariably rendered them resistant to DT in vitro, without altering basal cell growth. DTR-based selection efficiency and stability were comparable to those of established drug-resistance markers. As mice are insensitive to DT, DTR-based selection can be also applied in vivo. Direct injection of a GFP-DTR lentiviral vector into human cancer cell-line xenografts and patient-derived tumorgrafts implanted in mice, followed by systemic DT administration, yielded tumors entirely composed of permanently transduced cells and detectable by imaging systems. This approach enabled high-efficiency in vivo selection of xenografted human tumor tissues expressing ectopic transgenes, a hitherto unmet need for functional and morphological studies in laboratory animals. PMID:26420058

  13. Optimum location of external markers using feature selection algorithms for real-time tumor tracking in external-beam radiotherapy: a virtual phantom study.

    PubMed

    Nankali, Saber; Torshabi, Ahmad Esmaili; Miandoab, Payam Samadi; Baghizadeh, Amin

    2016-01-08

    In external-beam radiotherapy, using external markers is one of the most reliable tools to predict tumor position, in clinical applications. The main challenge in this approach is tumor motion tracking with highest accuracy that depends heavily on external markers location, and this issue is the objective of this study. Four commercially available feature selection algorithms entitled 1) Correlation-based Feature Selection, 2) Classifier, 3) Principal Components, and 4) Relief were proposed to find optimum location of external markers in combination with two "Genetic" and "Ranker" searching procedures. The performance of these algorithms has been evaluated using four-dimensional extended cardiac-torso anthropomorphic phantom. Six tumors in lung, three tumors in liver, and 49 points on the thorax surface were taken into account to simulate internal and external motions, respectively. The root mean square error of an adaptive neuro-fuzzy inference system (ANFIS) as prediction model was considered as metric for quantitatively evaluating the performance of proposed feature selection algorithms. To do this, the thorax surface region was divided into nine smaller segments and predefined tumors motion was predicted by ANFIS using external motion data of given markers at each small segment, separately. Our comparative results showed that all feature selection algorithms can reasonably select specific external markers from those segments where the root mean square error of the ANFIS model is minimum. Moreover, the performance accuracy of proposed feature selection algorithms was compared, separately. For this, each tumor motion was predicted using motion data of those external markers selected by each feature selection algorithm. Duncan statistical test, followed by F-test, on final results reflected that all proposed feature selection algorithms have the same performance accuracy for lung tumors. But for liver tumors, a correlation-based feature selection algorithm, in

  14. Marker-Based Estimates Reveal Significant Non-additive Effects in Clonally Propagated Cassava (Manihot esculenta): Implications for the Prediction of Total Genetic Value and the Selection of Varieties.

    PubMed

    Wolfe, Marnin D; Kulakow, Peter; Rabbi, Ismail Y; Jannink, Jean-Luc

    2016-08-31

    In clonally propagated crops, non-additive genetic effects can be effectively exploited by the identification of superior genetic individuals as varieties. Cassava (Manihot esculenta Crantz) is a clonally propagated staple food crop that feeds hundreds of millions. We quantified the amount and nature of non-additive genetic variation for three key traits in a breeding population of cassava from sub-Saharan Africa using additive and non-additive genome-wide marker-based relationship matrices. We then assessed the accuracy of genomic prediction for total (additive plus non-additive) genetic value. We confirmed previous findings based on diallel populations, that non-additive genetic variation is significant for key cassava traits. Specifically, we found that dominance is particularly important for root yield and epistasis contributes strongly to variation in CMD resistance. Further, we showed that total genetic value predicted observed phenotypes more accurately than additive only models for root yield but not for dry matter content, which is mostly additive or for CMD resistance, which has high narrow-sense heritability. We address the implication of these results for cassava breeding and put our work in the context of previous results in cassava, and other plant and animal species.

  15. A molecular marker based linkage map of Vitis.

    PubMed

    Lodhi, M A; Daly, M J; Ye, G N; Weeden, N F; Reisch, B I

    1995-08-01

    Genetic linkage maps of Vitis (2n = 38) have been constructed from a single interspecific hybrid grape population (60 seedlings) of 'Cayuga White' X 'Aurore'. The maps were primarily based on 422 RAPD markers but also included 16 RFLP and isozyme markers. These maps had an average distance of 6.1 cM between markers and were developed using a double-pseudotestcross strategy. The 'Cayuga White' map had 214 markers covering 1196 cM and that of 'Aurore' spanned over 1477 cM with 225 markers. The 'Cayuga White' map consisted of 20 linkage groups, whereas 22 linkage groups comprised the 'Aurore' map. The number of groups reduced to 19, as in some instances two or more groups from one parent showed homology with a single group from the other parent on the basis of markers heterozygous in both parents. Each linkage group ranged in size from 14 to 135 cM in 'Aurore' and from 14 to 124 cM in 'Cayuga White'. These maps provide enough coverage of the genome to allow quantitative trait locus analysis and map-based gene cloning.

  16. A genetic linkage map of quinoa ( Chenopodium quinoa) based on AFLP, RAPD, and SSR markers.

    PubMed

    Maughan, P J; Bonifacio, A; Jellen, E N; Stevens, M R; Coleman, C E; Ricks, M; Mason, S L; Jarvis, D E; Gardunia, B W; Fairbanks, D J

    2004-10-01

    Quinoa ( Chenopodium quinoa Willd.) is an important seed crop for human consumption in the Andean region of South America. It is the primary staple in areas too arid or saline for the major cereal crops. The objective of this project was to build the first genetic linkage map of quinoa. Selection of the mapping population was based on a preliminary genetic similarity analysis of four potential mapping parents. Breeding lines 'Ku-2' and '0654', a Chilean lowland type and a Peruvian Altiplano type, respectively, showed a low similarity coefficient of 0.31 and were selected to form an F(2) mapping population. The genetic map is based on 80 F(2) individuals from this population and consists of 230 amplified length polymorphism (AFLP), 19 simple-sequence repeat (SSR), and six randomly amplified polymorphic DNA markers. The map spans 1,020 cM and contains 35 linkage groups with an average marker density of 4.0 cM per marker. Clustering of AFLP markers was not observed. Additionally, we report the primer sequences and map locations for 19 SSR markers that will be valuable tools for future quinoa genome analysis. This map provides a key starting point for genetic dissection of agronomically important characteristics of quinoa, including seed saponin content, grain yield, maturity, and resistance to disease, frost, and drought. Current efforts are geared towards the generation of more than 200 mapped SSR markers and the development of several recombinant-inbred mapping populations.

  17. A rapid and robust selection procedure for generating drug-selectable marker-free recombinant malaria parasites

    PubMed Central

    Manzoni, Giulia; Briquet, Sylvie; Risco-Castillo, Veronica; Gaultier, Charlotte; Topçu, Selma; Ivănescu, Maria Larisa; Franetich, Jean-François; Hoareau-Coudert, Bénédicte; Mazier, Dominique; Silvie, Olivier

    2014-01-01

    Experimental genetics have been widely used to explore the biology of the malaria parasites. The rodent parasites Plasmodium berghei and less frequently P. yoelii are commonly utilised, as their complete life cycle can be reproduced in the laboratory and because they are genetically tractable via homologous recombination. However, due to the limited number of drug-selectable markers, multiple modifications of the parasite genome are difficult to achieve and require large numbers of mice. Here we describe a novel strategy that combines positive-negative drug selection and flow cytometry-assisted sorting of fluorescent parasites for the rapid generation of drug-selectable marker-free P. berghei and P. yoelii mutant parasites expressing a GFP or a GFP-luciferase cassette, using minimal numbers of mice. We further illustrate how this new strategy facilitates phenotypic analysis of genetically modified parasites by fluorescence and bioluminescence imaging of P. berghei mutants arrested during liver stage development. PMID:24755823

  18. A rapid and robust selection procedure for generating drug-selectable marker-free recombinant malaria parasites.

    PubMed

    Manzoni, Giulia; Briquet, Sylvie; Risco-Castillo, Veronica; Gaultier, Charlotte; Topçu, Selma; Ivănescu, Maria Larisa; Franetich, Jean-François; Hoareau-Coudert, Bénédicte; Mazier, Dominique; Silvie, Olivier

    2014-04-23

    Experimental genetics have been widely used to explore the biology of the malaria parasites. The rodent parasites Plasmodium berghei and less frequently P. yoelii are commonly utilised, as their complete life cycle can be reproduced in the laboratory and because they are genetically tractable via homologous recombination. However, due to the limited number of drug-selectable markers, multiple modifications of the parasite genome are difficult to achieve and require large numbers of mice. Here we describe a novel strategy that combines positive-negative drug selection and flow cytometry-assisted sorting of fluorescent parasites for the rapid generation of drug-selectable marker-free P. berghei and P. yoelii mutant parasites expressing a GFP or a GFP-luciferase cassette, using minimal numbers of mice. We further illustrate how this new strategy facilitates phenotypic analysis of genetically modified parasites by fluorescence and bioluminescence imaging of P. berghei mutants arrested during liver stage development.

  19. Genetic Structure, Linkage Disequilibrium and Signature of Selection in Sorghum: Lessons from Physically Anchored DArT Markers

    PubMed Central

    Bouchet, Sophie; Pot, David; Deu, Monique; Rami, Jean-François; Billot, Claire; Perrier, Xavier; Rivallan, Ronan; Gardes, Laëtitia; Xia, Ling; Wenzl, Peter; Kilian, Andrzej; Glaszmann, Jean-Christophe

    2012-01-01

    Population structure, extent of linkage disequilibrium (LD) as well as signatures of selection were investigated in sorghum using a core sample representative of worldwide diversity. A total of 177 accessions were genotyped with 1122 informative physically anchored DArT markers. The properties of DArTs to describe sorghum genetic structure were compared to those of SSRs and of previously published RFLP markers. Model-based (STRUCTURE software) and Neighbor-Joining diversity analyses led to the identification of 6 groups and confirmed previous evolutionary hypotheses. Results were globally consistent between the different marker systems. However, DArTs appeared more robust in terms of data resolution and bayesian group assignment. Whole genome linkage disequilibrium as measured by mean r2 decreased from 0.18 (between 0 to 10 kb) to 0.03 (between 100 kb to 1 Mb), stabilizing at 0.03 after 1 Mb. Effects on LD estimations of sample size and genetic structure were tested using i. random sampling, ii. the Maximum Length SubTree algorithm (MLST), and iii. structure groups. Optimizing population composition by the MLST reduced the biases in small samples and seemed to be an efficient way of selecting samples to make the best use of LD as a genome mapping approach in structured populations. These results also suggested that more than 100,000 markers may be required to perform genome-wide association studies in collections covering worldwide sorghum diversity. Analysis of DArT markers differentiation between the identified genetic groups pointed out outlier loci potentially linked to genes controlling traits of interest, including disease resistance genes for which evidence of selection had already been reported. In addition, evidence of selection near a homologous locus of FAR1 concurred with sorghum phenotypic diversity for sensitivity to photoperiod. PMID:22428056

  20. The bacterial paromomycin resistance gene, aphH, as a dominant selectable marker in Volvox carteri.

    PubMed

    Jakobiak, Thomas; Mages, Wolfgang; Scharf, Birgit; Babinger, Patrick; Stark, Klaus; Schmitt, Rüdiger

    2004-12-01

    The aminoglycoside antibiotic paromomycin that is highly toxic to the green alga Volvox carteri is efficiently inactivated by aminoglycoside 3'-phosphotransferase from Streptomyces rimosus. Therefore, we made constructs in which the bacterial aphH gene encoding this enzyme was combined with Volvox cis-regulatory elements in an attempt to develop a new dominant selectable marker--paromomycin resistance (PmR)--for use in Volvox nuclear transformation. The construct that provided the most efficient transformation was one in which aphH was placed between a chimeric promoter that was generated by fusing the Volvox hsp70 and rbcS3 promoters and the 3' UTR of the Volvox rbcS3 gene. When this plasmid was used in combination with a high-impact biolistic device, the frequency of stable PmR transformants ranged about 15 per 106 target cells. Due to rapid and sharp selection, PmR transformants were readily isolated after six days, which is half the time required for previously used markers. Co-transformation of an unselected marker ranged about 30%. The chimeric aphH gene was stably integrated into the Volvox genome, frequently as tandem multiple copies, and was expressed at a level that made selection of PmR transformants simple and unambiguous. This makes the engineered bacterial aphH gene an efficient dominant selection marker for the transformation and co-transformation of a broad range of V. carteri strains without the recurring need for using auxotrophic recipient strains.

  1. Marker-Assisted Selection for FHB at the Eastern Regional Small Grains Genotyping Lab

    USDA-ARS?s Scientific Manuscript database

    Head scab of wheat, caused by Fusarium graminearum, is a disease that affects wheat production in the Eastern soft wheat growing region of the U.S.A.. Genotyping and marker-assisted selection (MAS) are being applied in the Eastern wheat growing region to develop resistant wheat varieties and to char...

  2. A next-generation marker genotyping platform (AmpSeq) in heterozygous crops: a case study for marker assisted selection in grapevine

    USDA-ARS?s Scientific Manuscript database

    Marker assisted selection (MAS) is often employed in crop breeding programs to accelerate and enhance cultivar development, via selection during the juvenile phase and parental selection prior to crossing. Next generation sequencing (NGS) and its derivative technologies have been used for genome-wid...

  3. A next-generation marker genotyping platform (AmpSeq) in heterozygous crops: A case study for marker assisted selection in grapevine

    USDA-ARS?s Scientific Manuscript database

    Marker assisted selection (MAS) has become widely used in perennial crop breeding programs to accelerate and enhance cultivar development via selection during the juvenile phase and parental selection prior to crossing. Next generation sequencing (NGS) has been widely used for whole genome molecular...

  4. Selective targeting of liver cancer with the endothelial marker CD146

    PubMed Central

    Thomann, Stefan; Longerich, Thomas; Bazhin, Alexandr V.; Mier, Walter; Schemmer, Peter; Ryschich, Eduard

    2014-01-01

    Hepatocellular carcinomas are well-vascularized tumors; the endothelial cells in these tumors have a specific phenotype. Our aim was to develop a new approach for tumor-specific drug delivery with monoclonal antibody targeting of endothelial ligands. CD146, a molecule expressed on the endothelial surface of hepatocellular carcinoma, was identified as a promising candidate for targeting. In the present study, endothelial cells immediately captured circulating anti-CD146 (ME-9F1) antibody, while antibody binding in tumors was significantly higher than in hepatic endothelium. Macroscopically, after intravenous injection, there were no differences in the mean accumulation of anti-CD146 antibody in tumor compared to liver tissue, due to a compensating higher blood vessel density in the liver tissue. Additional blockade of nontumoral epitopes and intra-arterial administration, improved selective antibody capture in the tumor microvasculature and largely prevented antibody distribution in the lung and liver. The potential practical use of this approach was demonstrated by imaging of radionuclide-labeled ME-9F1 antibody, which showed excellent tumor-selective uptake. Our results provide a promising principle for the use of endothelial markers for intratumoral drug delivery. Tumor endothelium–based access might offer new opportunities for the imaging and therapy of hepatocellular carcinoma and other liver malignancies. PMID:25238265

  5. CombiROC: an interactive web tool for selecting accurate marker combinations of omics data.

    PubMed

    Mazzara, Saveria; Rossi, Riccardo L; Grifantini, Renata; Donizetti, Simone; Abrignani, Sergio; Bombaci, Mauro

    2017-03-30

    Diagnostic accuracy can be improved considerably by combining multiple markers, whose performance in identifying diseased subjects is usually assessed via receiver operating characteristic (ROC) curves. The selection of multimarker signatures is a complicated process that requires integration of data signatures with sophisticated statistical methods. We developed a user-friendly tool, called CombiROC, to help researchers accurately determine optimal markers combinations from diverse omics methods. With CombiROC data from different domains, such as proteomics and transcriptomics, can be analyzed using sensitivity/specificity filters: the number of candidate marker panels rising from combinatorial analysis is easily optimized bypassing limitations imposed by the nature of different experimental approaches. Leaving to the user full control on initial selection stringency, CombiROC computes sensitivity and specificity for all markers combinations, performances of best combinations and ROC curves for automatic comparisons, all visualized in a graphic interface. CombiROC was designed without hard-coded thresholds, allowing a custom fit to each specific data: this dramatically reduces the computational burden and lowers the false negative rates given by fixed thresholds. The application was validated with published data, confirming the marker combination already originally described or even finding new ones. CombiROC is a novel tool for the scientific community freely available at http://CombiROC.eu.

  6. CombiROC: an interactive web tool for selecting accurate marker combinations of omics data

    PubMed Central

    Mazzara, Saveria; Rossi, Riccardo L.; Grifantini, Renata; Donizetti, Simone; Abrignani, Sergio; Bombaci, Mauro

    2017-01-01

    Diagnostic accuracy can be improved considerably by combining multiple markers, whose performance in identifying diseased subjects is usually assessed via receiver operating characteristic (ROC) curves. The selection of multimarker signatures is a complicated process that requires integration of data signatures with sophisticated statistical methods. We developed a user-friendly tool, called CombiROC, to help researchers accurately determine optimal markers combinations from diverse omics methods. With CombiROC data from different domains, such as proteomics and transcriptomics, can be analyzed using sensitivity/specificity filters: the number of candidate marker panels rising from combinatorial analysis is easily optimized bypassing limitations imposed by the nature of different experimental approaches. Leaving to the user full control on initial selection stringency, CombiROC computes sensitivity and specificity for all markers combinations, performances of best combinations and ROC curves for automatic comparisons, all visualized in a graphic interface. CombiROC was designed without hard-coded thresholds, allowing a custom fit to each specific data: this dramatically reduces the computational burden and lowers the false negative rates given by fixed thresholds. The application was validated with published data, confirming the marker combination already originally described or even finding new ones. CombiROC is a novel tool for the scientific community freely available at http://CombiROC.eu. PMID:28358118

  7. Empirical Selection of Informative Microsatellite Markers within Co-ancestry Pig Populations Is Required for Improving the Individual Assignment Efficiency

    PubMed Central

    Li, Y. H.; Chu, H. P.; Jiang, Y. N.; Lin, C. Y.; Li, S. H.; Li, K. T.; Weng, G. J.; Cheng, C. C.; Lu, D. J.; Ju, Y. T.

    2014-01-01

    The Lanyu is a miniature pig breed indigenous to Lanyu Island, Taiwan. It is distantly related to Asian and European pig breeds. It has been inbred to generate two breeds and crossed with Landrace and Duroc to produce two hybrids for laboratory use. Selecting sets of informative genetic markers to track the genetic qualities of laboratory animals and stud stock is an important function of genetic databases. For more than two decades, Lanyu derived breeds of common ancestry and crossbreeds have been used to examine the effectiveness of genetic marker selection and optimal approaches for individual assignment. In this paper, these pigs and the following breeds: Berkshire, Duroc, Landrace and Yorkshire, Meishan and Taoyuan, TLRI Black Pig No. 1, and Kaohsiung Animal Propagation Station Black pig are studied to build a genetic reference database. Nineteen microsatellite markers (loci) provide information on genetic variation and differentiation among studied breeds. High differentiation index (FST) and Cavalli-Sforza chord distances give genetic differentiation among breeds, including Lanyu’s inbred populations. Inbreeding values (FIS) show that Lanyu and its derived inbred breeds have significant loss of heterozygosity. Individual assignment testing of 352 animals was done with different numbers of microsatellite markers in this study. The testing assigned 99% of the animals successfully into their correct reference populations based on 9 to 14 markers ranking D-scores, allelic number, expected heterozygosity (HE) or FST, respectively. All miss-assigned individuals came from close lineage Lanyu breeds. To improve individual assignment among close lineage breeds, microsatellite markers selected from Lanyu populations with high polymorphic, heterozygosity, FST and D-scores were used. Only 6 to 8 markers ranking HE, FST or allelic number were required to obtain 99% assignment accuracy. This result suggests empirical examination of assignment-error rates is required if

  8. Development of molecular markers linked to the 'Fiesta' linkage group 7 major QTL for fire blight resistance and their application for marker-assisted selection.

    PubMed

    Khan, Muhammad A; Durel, Charles-Eric; Duffy, Brion; Drouet, Damien; Kellerhals, Markus; Gessler, Cesare; Patocchi, Andrea

    2007-06-01

    A fire blight resistance QTL explaining 34.3%-46.6% of the phenotypic variation was recently identified on linkage group 7 of apple cultivar 'Fiesta' (F7). However, markers flanking this QTL were AFLP and RAPD markers unsuitable for marker-assisted selection (MAS). Two RAPD markers bracketing the QTL have been transformed into SCAR (sequence-characterized amplified region) markers, and an SSR marker specific for the region was developed. Pedigree analysis of 'Fiesta' with these markers enabled tracking of the F7 QTL allele back to 'Cox's Orange Pippin'. Stability of the effect of this QTL allele in different backgrounds was analyzed by inoculating progeny plants of a cross between 'Milwa', a susceptible cultivar, and '1217', a moderately resistant cultivar, and a set of cultivars that carry or lack the allele conferring increased fire blight resistance. Progenies and cultivars that carried both markers were significantly more resistant than those that did not carry both markers, indicating high stability of the F7 QTL allele in different backgrounds. This stability and the availability of reproducible markers bracketing the QTL make this locus promising for use in MAS.

  9. Bulk development and stringent selection of microsatellite markers in the western flower thrips Frankliniella occidentalis

    PubMed Central

    Cao, Li-Jun; Li, Ze-Min; Wang, Ze-Hua; Zhu, Liang; Gong, Ya-Jun; Chen, Min; Wei, Shu-Jun

    2016-01-01

    Recent improvements in next-generation sequencing technologies have enabled investigation of microsatellites on a genome-wide scale. Faced with a huge amount of candidates, the use of appropriate marker selection criteria is crucial. Here, we used the western flower thrips Frankliniella occidentalis for an empirical microsatellite survey and validation; 132,251 candidate microsatellites were identified, 92,102 of which were perfect. Dinucleotides were the most abundant category, while (AG)n was the most abundant motif. Sixty primer pairs were designed and validated in two natural populations, of which 30 loci were polymorphic, stable, and repeatable, but not all in Hardy–Weinberg equilibrium (HWE) and linkage equilibrium. Four marker panels were constructed to understand effect of marker selection on population genetic analyses: (i) only accept loci with single nucleotide insertions (SNI); (ii) only accept the most polymorphic loci (MP); (iii) only accept loci that did not deviate from HWE, did not show SNIs, and had unambiguous peaks (SS) and (iv) all developed markers (ALL). Although the MP panel resulted in microsatellites of highest genetic diversity followed by the SNI, the SS performed best in individual assignment. Our study proposes stringent criteria for selection of microsatellites from a large-scale number of genomic candidates for population genetic studies. PMID:27197749

  10. Selecting soybean resistant to the cyst nematode Heterodera glycines using simple sequence repeat (microssatellite) markers.

    PubMed

    Espindola, S M C G; Hamawaki, O T; Oliveira, A P; Hamawaki, C D L; Hamawaki, R L; Takahashi, L M

    2016-03-11

    The soybean cyst nematode (SCN) is a major cause of soybean yield reduction. The objective of this study was to evaluate the efficiency of marker-assisted selection to identify genotypes resistant to SCN race 3 infection, using Sat_168 and Sat-141 resistance quantitative trait loci. The experiment was carried out under greenhouse conditions, using soybean populations originated from crosses between susceptible and resistant parent stock: CD-201 (susceptible) and Foster IAC (resistant), Conquista (susceptible) and S83-30 (resistant), La-Suprema (susceptible) and S57-11 (resistant), and Parecis (susceptible) and S65-50 (resistant). Plants were inoculated with SCN and evaluated according to the female index (FI), those with FI < 10% were classified as resistant to nematode infection. Plants were genotyped for SCN resistance using microsatellite markers Sat-141 and Sat_168. Marker selection efficiency was analyzed by a contingency table, taking into account genotypic versus phenotypic evaluations for each line. These markers were shown to be useful tool for selection of SCN race 3.

  11. Genetic variability at neutral markers, quantitative trait land trait in a subdivided population under selection.

    PubMed

    Le Corre, Valérie; Kremer, Antoine

    2003-07-01

    Genetic variability in a subdivided population under stabilizing and diversifying selection was investigated at three levels: neutral markers, QTL coding for a trait, and the trait itself. A quantitative model with additive effects was used to link genotypes to phenotypes. No physical linkage was introduced. Using an analytical approach, we compared the diversity within deme (H(S)) and the differentiation (F(ST)) at the QTL with the genetic variance within deme (V(W)) and the differentiation (Q(ST)) for the trait. The difference between F(ST) and Q(ST) was shown to depend on the relative amounts of covariance between QTL within and between demes. Simulations were used to study the effect of selection intensity, variance of optima among demes, and migration rate for an allogamous and predominantly selfing species. Contrasting dynamics of the genetic variability at markers, QTL, and trait were observed as a function of the level of gene flow and diversifying selection. The highest discrepancy among the three levels occurred under highly diversifying selection and high gene flow. Furthermore, diversifying selection might cause substantial heterogeneity among QTL, only a few of them showing allelic differentiation, while the others behave as neutral markers.

  12. Detecting the footprints of divergent selection in oaks with linked markers

    PubMed Central

    Goicoechea, P G; Petit, R J; Kremer, A

    2012-01-01

    Genome scans are increasingly used to study ecological speciation, providing a useful genome-wide perspective on divergent selection in the presence of gene flow. Here, we compare current approaches to detect footprints of divergent selection in closely related species. We analyzed 192 individuals from two interfertile European temperate oak species using 30 nuclear microsatellites from eight linkage groups. These markers present little intraspecific differentiation and can be used in combination to assign individual genotypes to species. We first show that different outlier detection tests give somewhat different results, possibly due to model constraints. Second, using linkage information for these markers, we further characterize the signature of divergent selection in the presence of gene flow. In particular, we show that recombination estimates for regions with outlier markers are lower than those for a control region, in line with a prediction from ecological speciation theory. Most importantly, we show that analyses at the haplotype level can distinguish between truly divergent (bi-directional) selection and positive selection in one of the two species, offering a new and improved method for characterizing the speciation process. PMID:22990311

  13. Genetic transformation of Nannochloropsis oculata with a bacterial phleomycin resistance gene as dominant selective marker

    NASA Astrophysics Data System (ADS)

    Ma, Xiaolei; Pan, Kehou; Zhang, Lin; Zhu, Baohua; Yang, Guanpin; Zhang, Xiangyang

    2016-04-01

    The gene ble from Streptoalloteichus hindustanus is widely used as a selective antibiotic marker. It can control the phleomycin resistance, and significantly increase the tolerance of hosts to zeocin. The unicellular marine microalga Nannochloropsis oculata is extremely sensitive to zeocin. We selected ble as the selective marker for the genetic transformation of N. oculata. After the algal cells at a density of 2×107 cells mL-1 was digested with 4% hemicellulase and 2% driselase for 1 h, the protoplasts accounted for 90% of the total. The ble was placed at the downstream of promoter HSP70A-RUBS2 isolated from Chlamydomonas reinhardtii, yielding a recombinant expression construct pMS188. The construct was transferred into the protoplasts through electroporation (1 kV, 15 μS). The transformed protoplasts were cultured in fresh f/2 liquid medium, and selected on solid f/2 medium supplemented with 500 ng mL-1 zeocin. The PCR result proved that ble existed in the transformants. Three transformants had been cultured for at least 5 generations without losing ble. Southern blotting analysis showed that the ble has been integrated into the genome of N. oculata. The ble will serve as a new dominant selective marker in genetic engineering N. oculata.

  14. A method to infer positive selection from marker dynamics in an asexual population.

    PubMed

    Illingworth, Christopher J R; Mustonen, Ville

    2012-03-15

    The observation of positive selection acting on a mutant indicates that the corresponding mutation has some form of functional relevance. Determining the fitness effects of mutations thus has relevance to many interesting biological questions. One means of identifying beneficial mutations in an asexual population is to observe changes in the frequency of marked subsets of the population. We here describe a method to estimate the establishment times and fitnesses of beneficial mutations from neutral marker frequency data. The method accurately reproduces complex marker frequency trajectories. In simulations for which positive selection is close to 5% per generation, we obtain correlations upwards of 0.91 between correct and inferred haplotype establishment times. Where mutation selection coefficients are exponentially distributed, the inferred distribution of haplotype fitnesses is close to being correct. Applied to data from a bacterial evolution experiment, our method reproduces an observed correlation between evolvability and initial fitness defect.

  15. MRJP microsatellite markers in Africanized Apis mellifera colonies selected on the basis of royal jelly production.

    PubMed

    Parpinelli, R S; Ruvolo-Takasusuki, M C C; Toledo, V A A

    2014-08-28

    It is important to select the best honeybees that produce royal jelly to identify important molecular markers, such as major royal jelly proteins (MRJPs), and hence contribute to the development of new breeding strategies to improve the production of this substance. Therefore, this study focused on evaluating the genetic variability of mrjp3, mrjp5, and mrjp8 and associated allele maintenance during the process of selective reproduction in Africanized Apis mellifera individuals, which were chosen based on royal jelly production. The three loci analyzed were polymorphic, and produced a total of 16 alleles, with 4 new alleles, which were identified at mrjp5. The effective number of alleles at mrjp3 was 3.81. The observed average heterozygosity was 0.4905, indicating a high degree of genetic variability at these loci. The elevated FIS values for mrjp3, mrjp5, and mrjp8 (0.4188, 0.1077, and 0.2847, respectively) indicate an excess of homozygotes. The selection of Africanized A. mellifera queens for royal jelly production has maintained the mrjp3 C, D, and E alleles; although, the C allele occurred at a low frequency. The heterozygosity and FIS values show that the genetic variability of the queens is decreasing at the analyzed loci, generating an excess of homozygotes. However, the large numbers of drones that fertilize the queens make it difficult to develop homozygotes at mrjp3. Mating through instrumental insemination using the drones of known genotypes is required to increase the efficiency of Africanized A. mellifera-breeding programs, and to improve the quality and efficiency of commercial royal jelly apiaries.

  16. Development of PCR-based codominant markers flanking the Alt3 gene in rye.

    PubMed

    Miftahudin; Scoles, G J; Gustafson, J P

    2004-04-01

    Aluminum (Al) toxicity is considered to be a major problem for crop growth and production on acid soils. The ability of crops to overcome Al toxicity varies among crop species and cultivars. Rye (Secale cereale L.) is the most Al-tolerant species among the Triticeae. Our previous study showed that Al tolerance in a rye F6 recombinant inbred line (RIL) population was controlled by a single gene designated as the aluminum tolerance (Alt3) gene on chromosome 4RL. Based on the DNA sequence of a rice (Oryza sativa L.) BAC clone suspected to be syntenic to the Alt3 gene region, we developed two PCR-based codominant markers flanking the gene. These two markers, a sequence-tagged site (STS) marker and a cleaved amplified polymorphic sequence (CAPS) marker, each flanked the Alt3 gene at an approximate distance of 0.4 cM and can be used to facilitate high-resolution mapping of the gene. The markers might also be used for marker-assisted selection in rye or wheat (Triticum aestivum L.) breeding programs to obtain Al-tolerant lines and (or) cultivars.

  17. Novel recombinant binary vectors harbouring Basta (bar) gene as a plant selectable marker for genetic transformation of plants.

    PubMed

    Nada, Reham M

    2016-04-01

    Genetic transformation is one of the most widely used technique in crop improvement. However, most of the binary vectors used in this technique, especially cloning based, contain antibiotic genes as selection marker that raise serious consumer and environmental concerns; moreover, they could be transferred to non-target hosts with deleterious effects. Therefore, the goal of this study was reconstruction of the widely used pBI121 binary vector by substituting the harmful antibiotic selection marker gene with a less-harmful selection marker, Basta (herbicide resistance gene). The generated vectors were designated as pBI121NB and pBI121CB, in which Basta gene was expressed under the control of Nos or CaMV 35S promoter, respectively. The successful integration of the new inserts into both the vectors was confirmed by PCR, restriction digestion and sequencing. Both these vectors were used in transforming Arabidopsis, Egyptian wheat and barley varieties using LBA4404 and GV3101 Agrobacterium strains. The surfactant Tween-20 resulted in an efficient transformation and the number of Arabidopsis transformants was about 6-9 %. Soaked seeds of wheat and barley were transformed with Agrobacterium to introduce the bacteria to the growing shoot apices. The percentage of transgenic lines was around 16-17 and 14-15 % for wheat and barley, respectively. The quantitative studies presented in this work showed that both LBA4404 and GV3101 strains were suitable for transforming Egyptian wheat and barley.

  18. Effect of outlier removal on gene marker selection using support vector machines.

    PubMed

    Moffitt, Richard; Phan, John; Hemby, Scott; Wang, May

    2005-01-01

    Biological markers are useful tools for the diagnosis and prognosis of disease. Many different methods are currently used to extract markers from multiple data sources, including gene expression microarrays. This paper investigates the effect of outlier removal on the performance of one such biomarker selection method, Support Vector Machines (SVM). A simple method of outlier removal is employed as a preprocessing step before the data is used for SVM analysis. Both linear and radial basis kernels are used as well as four different normalization techniques. Results show that outlier removal increases the number of highly predictive genes as well as the number of poorly predicting genes. This result thus supports the use of outlier removal prior to biological marker identification via SVM analysis.

  19. Organophosphorus hydrolase: a multifaceted plant genetic marker which is selectable, scorable, and quantifiable in whole seed.

    PubMed

    Pinkerton, T Scott; Wild, James R; Howard, John A

    2012-01-01

    Organophosphorus hydrolase (OPH, EC 3.1.8.1) provides a novel function as an alternative genetic marker system for use in many types of plant transformations. OPH is a high-capacity hydrolase with multiple organophosphorus substrates, many of which are neurotoxins and thus used extensively as pesticides. This spectrum of organophosphates includes compounds that are phytotoxic as well as those that are hydrolyzed to products that are easily detected visually without significant disruption of plant health. This dichotomy gives OPH the features of both a selectable marker as well as that of a scorable marker system, and these characteristics have been tested at several stages during the plant transformation and regeneration process. Finally, it is possible to quantify hydrolytic activity in the seed without interfering with its subsequent growth and regeneration.

  20. Marker-assisted selection: an approach for precision plant breeding in the twenty-first century.

    PubMed

    Collard, Bertrand C Y; Mackill, David J

    2008-02-12

    DNA markers have enormous potential to improve the efficiency and precision of conventional plant breeding via marker-assisted selection (MAS). The large number of quantitative trait loci (QTLs) mapping studies for diverse crops species have provided an abundance of DNA marker-trait associations. In this review, we present an overview of the advantages of MAS and its most widely used applications in plant breeding, providing examples from cereal crops. We also consider reasons why MAS has had only a small impact on plant breeding so far and suggest ways in which the potential of MAS can be realized. Finally, we discuss reasons why the greater adoption of MAS in the future is inevitable, although the extent of its use will depend on available resources, especially for orphan crops, and may be delayed in less-developed countries. Achieving a substantial impact on crop improvement by MAS represents the great challenge for agricultural scientists in the next few decades.

  1. Genome-wide association analysis of bacterial cold water disease resistance in rainbow trout reveals the potential of a hybrid approach between genomic selection and marker assisted selection

    USDA-ARS?s Scientific Manuscript database

    Genomic selection (GS) simultaneously incorporates dense SNP marker genotypes with phenotypic data from related animals to predict animal-specific genomic breeding value (GEBV), which circumvents the need to measure the disease phenotype in potential breeders. Marker assisted selection (MAS) involv...

  2. A and MdMYB1 allele-specific markers controlling apple (Malus x domestica Borkh.) skin color and suitability for marker-assisted selection.

    PubMed

    Zhang, X J; Wang, L X; Chen, X X; Liu, Y L; Meng, R; Wang, Y J; Zhao, Z Y

    2014-10-31

    Pre-selection for fruit skin color at the seedling stage would be highly advantageous, with marker-assisted selection offering a potential method for apple pre-selection. A and MdMYB1 alleles are allele-specific DNA markers that are potentially associated with apple skin color, and co-segregate with the Rf and Rni loci, respectively. Here, we assessed the potential application of these 2 alleles for marker-assisted breeding across 30 diverse cultivars and 2 apple seedling progenies. The red skin color phenotype was usually associated with the MdMYB1-1 allele and A(1) allele, respectively, while the 2 molecular markers provided approximately 91% predictability in the 'Fuji' x 'Cripps Pink' and 'Fuji' x 'Gala' progenies. The results obtained from the 30 cultivars and 2 progenies were consistent for the 2 molecular markers. Hence, the results supported that Rf and Rni could be located in a gene cluster, or even correspond to alleles of the same gene. Our results are consistent with the hypothesis that red/yellow dimorphism is controlled by a monogenic system, with the presence of the red anthocyanin pigmentation being dominant. In addition, our results supported that the practical utilization of the 2 function markers to efficiently and accurately select red-skinned apple cultivars in apple scion breeding programs.

  3. Evaluation of ensemble Monte Carlo variable selection for identification of metabolite markers on NMR data.

    PubMed

    Esquerre, C A; Gowen, A A; O'Gorman, A; Downey, G; O'Donnell, C P

    2017-04-29

    The aim of this study was to investigate the potential of the recently developed ensemble Monte Carlo Variable Selection (EMCVS) method to identify the relevant portions of high resolution (1)H NMR spectra as a metabolite fingerprinting tool and compare to a widely used method (Variable importance on projection (VIP)) and recently proposed variable selected methods i.e. selectivity ratio (SR) and significance multivariate correlation (sMC). As case studies two quantitative publicly available datasets: wine samples, urine samples of rats, and an experiment on mushroom (Agaricus bisporus) were examined. EMCVS outperformed the three other variable selection methods in most cases, selecting fewer chemical shifts and leading to improved classification of mushrooms and prediction of onion by-products intake and wine components. These fewer chemical shift regions facilitate the interpretation of the NMR spectra, fingerprinting and identification of metabolite markers. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Selectable marker elimination in the T0 generation by Agrobacterium-mediated co-transformation involving Mungbean yellow mosaic virus TrAP as a non-conditional negative selectable marker and bar for transient positive selection.

    PubMed

    RamanaRao, Mangu Venkata; Veluthambi, Karuppannan

    2010-05-01

    Transient selection involving the bar gene and non-conditional negative selection against stable T-DNA integration through the use of the Mungbean yellow mosaic virus (MYMV) transcriptional activator protein gene (TrAP) were used in a novel co-transformation strategy to generate selectable marker gene (SMG)-eliminated transgenic tobacco plants in the T(0) generation itself. Two compatible binary plasmids, pCam-bar-TrAP-gus harbouring bar as an SMG and the MYMV TrAP gene as a non-conditional negative selectable marker, and pGA472 with the nptII gene as an unselected experimental gene of interest (GOI) were placed in the Agrobacterium tumefaciens strain EHA105 and used for co-transformation. Transient selection with 5 mg l(-1) phosphinothricin (PPT) for 2-4 weeks and subsequent establishment in a PPT-minus medium yielded 114 plants from 200 leaf discs. The unselected nptII gene was detected by Southern blot analysis in 13 plants, revealing a co-transformation efficiency of 11.5%. Five of these plants harboured only the nptII gene (GOI) and not the bar gene (SMG). Thus, SMG elimination was achieved in the T(0) generation itself in 4.4% (5/114) of plants, which were transiently selected for 2-4 weeks on PPT. MYMV TrAP, a non-conditional negative selectable marker, effectively reduced the recovery of plants with stable integration of the SMG (bar).

  5. Development of microsatellite markers based on expressed sequence tags in Asparagus cochinchinensis (Asparagaceae)1

    PubMed Central

    Kim, Bo-Yun; Park, Han-Sol; Lee, Jung-Hoon; Kwak, Myounghai; Kim, Young-Dong

    2017-01-01

    Premise of the study: Transcriptome-derived simple sequence repeat (SSR) markers were developed in Asparagus cochinchinensis (Asparagaceae). Due to its application in traditional medicine, its wild populations are threatened by over-collection even in protected areas, requiring immediate conservation efforts. Methods and Results: Based on transcriptome data of A. cochinchinensis, 96 primer pairs with two to seven alleles per locus were selected for initial validation; of those, 27 primer pairs amplified across all samples, resulting in 15 polymorphic and 12 monomorphic microsatellite markers. The usefulness of these markers was assessed in 60 individuals representing three populations of A. cochinchinensis. Observed and expected heterozygosity values ranged from 0.050 to 0.950 and 0.049 to 0.626, respectively. Cross-species amplification of the 27 markers was tested in the related species A. rigidulus and A. schoberioides. Conclusions: These polymorphic, transcriptome-derived SSR markers can be used as molecular markers to study population genetics and ecological conservation in A. cochinchinensis and related taxa. PMID:28439480

  6. A family of LRR sequences in the vicinity of the Co-2 locus for anthracnose resistance in Phaseolus vulgaris and its potential use in marker-assisted selection.

    PubMed

    Geffroy, V; Creusot, F; Falquet, J; Sévignac, M; Adam-Blondon, A F; Bannerot, H; Gepts, P; Dron, M

    1998-03-01

    Molecular markers offer new opportunities for breeding for disease resistance. Resistance gene pyramiding in a single cultivar, as a strategy for durable resistance, can be facilitated by marker-assisted selection (MAS). A RAPD marker, ROH20(450), linked to the Mesoamerican Co-2 anthracnose resistance gene, was previously transformed into a SCAR marker, SCH20. In the present paper we have further characterized the relevance of the SCH20 SCAR marker in different genetic backgrounds. Since this SCAR marker was found to be useful mainly in the Andean gene pool, we identified a new PCR-based marker (SCAreoli) for indirect scoring of the presence of the Co-2 gene. The SCAreoli SCAR marker is polymorphic in the Mesoamerican as well as in the Andean gene pool and should be useful in MAS. We also report that PvH20, the cloned sequence corresponding to the 450-bp RAPD marker ROH20(450), contains six imperfect leucine-rich repeats, and reveals a family of related sequences in the vicinity of the Co-2 locus. These results are discussed in the context of the recent cloning of some plant resistance genes.

  7. Non-homologous end joining-mediated functional marker selection for DNA cloning in the yeast Kluyveromyces marxianus.

    PubMed

    Hoshida, Hisashi; Murakami, Nobutada; Suzuki, Ayako; Tamura, Ryoko; Asakawa, Jun; Abdel-Banat, Babiker M A; Nonklang, Sanom; Nakamura, Mikiko; Akada, Rinji

    2014-01-01

    The cloning of DNA fragments into vectors or host genomes has traditionally been performed using Escherichia coli with restriction enzymes and DNA ligase or homologous recombination-based reactions. We report here a novel DNA cloning method that does not require DNA end processing or homologous recombination, but that ensures highly accurate cloning. The method exploits the efficient non-homologous end-joining (NHEJ) activity of the yeast Kluyveromyces marxianus and consists of a novel functional marker selection system. First, to demonstrate the applicability of NHEJ to DNA cloning, a C-terminal-truncated non-functional ura3 selection marker and the truncated region were PCR-amplified separately, mixed and directly used for the transformation. URA3(+) transformants appeared on the selection plates, indicating that the two DNA fragments were correctly joined by NHEJ to generate a functional URA3 gene that had inserted into the yeast chromosome. To develop the cloning system, the shortest URA3 C-terminal encoding sequence that could restore the function of a truncated non-functional ura3 was determined by deletion analysis, and was included in the primers to amplify target DNAs for cloning. Transformation with PCR-amplified target DNAs and C-terminal truncated ura3 produced numerous transformant colonies, in which a functional URA3 gene was generated and was integrated into the chromosome with the target DNAs. Several K. marxianus circular plasmids with different selection markers were also developed for NHEJ-based cloning and recombinant DNA construction. The one-step DNA cloning method developed here is a relatively simple and reliable procedure among the DNA cloning systems developed to date.

  8. [Cloning goat producing human lactoferrin with genetically modified donor cells selected by single or dual markers].

    PubMed

    An, Liyou; Yuan, Yuguo; Yu, Baoli; Yang, Tingjia; Cheng, Yong

    2012-12-01

    We compared the efficiency of cloning goat using human lactoferrin (hLF) with genetically modified donor cells marked by single (Neo(r)) or double (Neo(r)/GFP) markers. Single marker expression vector (pBLC14) or dual markers expression vector (pAPLM) was delivered to goat fetal fibroblasts (GFF), and then the transgenic GFF was used as donor cells to produce transgenic goats. Respectively, 58.8% (20/34) and 86.7% (26/30) resistant cell lines confirmed the transgenic integration by PCR. Moreover, pAPLM cells lines were subcultured with several passages, only 20% (6/30) cell lines was observed fluorescence from each cell during the cell passage. Somatic cell nuclear transfer using the donor cells harbouring pBLC14 or pAPLM construct, resulting in a total of 806 reconstructed embryos, a pregnancy rate at 35 d (53.8%, 39.1%) and 60 d (26.9%, 21.7%), and an offspring birth rate (1.9%, 1.4%) with 5 and 7 newborn cloned goats, respectively. Transgene was confirmed by PCR and southern-blot in all cloned offspring. There were no significant differences at the reconstructed embryo fusion rates, pregnancy rates and the birth rate (P > 0.05) between single and double markers groups. The Neo(r)/GFP double markers could improve the reliability for accurately and efficiently selecting the genetically modified donor cells. No adverse effect was observed on the efficiency of transgenic goat production by SCNT using somatic cells transfected with double (Neo(r)/GFP) markers vector.

  9. Comparative mapping and marker-assisted selection in Rosaceae fruit crops

    PubMed Central

    Dirlewanger, Elisabeth; Graziano, Enrique; Joobeur, Tarek; Garriga-Calderé, Francesc; Cosson, Patrick; Howad, Werner; Arús, Pere

    2004-01-01

    The development of saturated linkage maps using transferable markers, restriction fragment length polymorphisms, and micro-satellites has provided a foundation for fruit tree genetics and breeding. A Prunus reference map with 562 such markers is available, and a further set of 13 maps constructed with a subset of these markers has allowed genome comparison among seven Prunus diploid (x = 8) species (almond, peach, apricot, cherry, Prunus ferganensis, Prunus davidiana, and Prunus cerasifera); marker colinearity was the rule with all of them. Preliminary results of the comparison between apple and Prunus maps suggest a high level of synteny between these two genera. Conserved genomic regions have also been detected between Prunus and Arabidopsis. By using the data from different linkage maps anchored with the reference Prunus map, it has been possible to establish, in a general map, the position of 28 major genes affecting agronomic characters found in different species. Markers tightly linked to the major genes responsible for the expression of important traits (disease/pest resistances, fruit/nut quality, self-incompatibility, etc.) have been developed in apple and Prunus and are currently in use for marker-assisted selection in breeding programs. Quantitative character dissection using linkage maps and candidate gene approaches has already started. Genomic tools such as the Prunus physical map, large EST collections in both Prunus and Malus, and the establishment of the map position of high numbers of ESTs are required for a better understanding of the Rosaceae genome and to foster additional research and applications on fruit tree genetics. PMID:15159547

  10. Mild Cognitive Impairment is Associated with Selected Functional Markers: Integrating Concurrent, Longitudinal, and Stability Effects

    PubMed Central

    Dolcos, Sanda; MacDonald, Stuart W.S.; Braslavsky, Anna; Camicioli, Richard; Dixon, Roger A.

    2013-01-01

    Objective We examined functional performance on multiple indicators for two cognitive status groups: (a) not impaired controls (NIC) and (b) mild cognitive impairment (MCI). We identified functional markers associated with differences, changes, and stability in cognitive status. Method In the Victoria Longitudinal Study (VLS) we examined cognitive status group effects in (a) cross-sectional functional performance, (b) longitudinal stability, (c) longitudinal functional performance change, and (d) functional marker prediction of later cognitive status. We assembled markers from five continuous clusters of MCI-related functional factors: biological vitality, activity lifestyle, psychosocial affect, subjective health, and global cognition. We used a cross-sectional sample and a two-wave longitudinal sample, stratified by age (mid-old, old-old) and cognitive status (MCI, NIC). Results First, cross-sectional results showed that eight markers differentiated MCI and NIC adults, with the latter performing uniformly better. The groups differed on diastolic blood pressure, body mass index, positive and negative affect, MMSE, and the lifestyle indicators of self-maintenance, travel, and novel cognitive activities. Second, Wave1 to Wave2 stabilities in cognitive status classification were high. Third, several markers differentiated the stable (NIC-to-NIC, MCI-to-MCI) from the unstable (NIC-to-MCI, MCI-to-NIC) cognitive status groups. Fourth, five relevant markers for identifying older adults at risk for cognitive status changes were: diastolic blood pressure, self-maintenance activities, novel cognitive activities, positive affect, and global cognitive status. Conclusion Selected risk and protective factors differentiate persons classified with MCI from those not currently cognitively impaired, both cross-sectionally and longitudinally. PMID:22251311

  11. Genome-wide identification of markers for selecting higher oil content in oil palm.

    PubMed

    Bai, Bin; Wang, Le; Lee, May; Zhang, Yingjun; Rahmadsyah; Alfiko, Yuzer; Ye, Bao Qing; Wan, Zi Yi; Lim, Chin Huat; Suwanto, Antonius; Chua, Nam-Hai; Yue, Gen Hua

    2017-05-30

    Oil palm (Elaeis guineensis, Jacq.) is the most important source of edible oil. The improvement of oil yield is currently slow in conventional breeding programs due to long generation intervals. Marker-assisted selection (MAS) has the potential to accelerate genetic improvement. To identify DNA markers associated with oil content traits for MAS, we performed quantitative trait loci (QTL) mapping using genotyping by sequencing (GBS) in a breeding population derived from a cross between Deli Dura and Ghana Pisifera, containing 153 F1 trees. We constructed a high-density linkage map containing 1357 SNPs and 123 microsatellites. The 16 linkage groups (LGs) spanned 1527 cM, with an average marker space of 1.03 cM. One significant and three suggestive QTL for oil to bunch (O/B) and oil to dry mesocarp (O/DM) were mapped on LG1, LG8, and LG10 in a F1 breeding population, respectively. These QTL explained 7.6-13.3% of phenotypic variance. DNA markers associated with oil content in these QTL were identified. Trees with beneficial genotypes at two QTL for O/B showed an average O/B of 30.97%, significantly (P < 0.01) higher than that of trees without any beneficial QTL genotypes (average O/B of 28.24%). QTL combinations showed that the higher the number of QTL with beneficial genotypes, the higher the resulting average O/B in the breeding population. A linkage map with 1480 DNA markers was constructed and used to identify QTL for oil content traits. Pyramiding the identified QTL with beneficial genotypes associated with oil content traits using DNA markers has the potential to accelerate genetic improvement for oil yield in the breeding population of oil palm.

  12. Comparative mapping and marker-assisted selection in Rosaceae fruit crops.

    PubMed

    Dirlewanger, Elisabeth; Graziano, Enrique; Joobeur, Tarek; Garriga-Calderé, Francesc; Cosson, Patrick; Howad, Werner; Arús, Pere

    2004-06-29

    The development of saturated linkage maps using transferable markers, restriction fragment length polymorphisms, and micro-satellites has provided a foundation for fruit tree genetics and breeding. A Prunus reference map with 562 such markers is available, and a further set of 13 maps constructed with a subset of these markers has allowed genome comparison among seven Prunus diploid (x = 8) species (almond, peach, apricot, cherry, Prunus ferganensis, Prunus davidiana, and Prunus cerasifera); marker colinearity was the rule with all of them. Preliminary results of the comparison between apple and Prunus maps suggest a high level of synteny between these two genera. Conserved genomic regions have also been detected between Prunus and Arabidopsis. By using the data from different linkage maps anchored with the reference Prunus map, it has been possible to establish, in a general map, the position of 28 major genes affecting agronomic characters found in different species. Markers tightly linked to the major genes responsible for the expression of important traits (disease/pest resistances, fruit/nut quality, self-incompatibility, etc.) have been developed in apple and Prunus and are currently in use for marker-assisted selection in breeding programs. Quantitative character dissection using linkage maps and candidate gene approaches has already started. Genomic tools such as the Prunus physical map, large EST collections in both Prunus and Malus, and the establishment of the map position of high numbers of ESTs are required for a better understanding of the Rosaceae genome and to foster additional research and applications on fruit tree genetics.

  13. Mild cognitive impairment is associated with selected functional markers: integrating concurrent, longitudinal, and stability effects.

    PubMed

    Dolcos, Sanda; MacDonald, Stuart W S; Braslavsky, Anna; Camicioli, Richard; Dixon, Roger A

    2012-03-01

    We examined functional performance on multiple indicators for two cognitive status groups: (a) not impaired controls (NIC) and (b) mild cognitive impairment (MCI). We identified functional markers associated with differences, changes, and stability in cognitive status. In the Victoria Longitudinal Study (VLS) we examined cognitive status group effects in (a) cross-sectional functional performance, (b) longitudinal stability, (c) longitudinal functional performance change, and (d) functional marker prediction of later cognitive status. We assembled markers from five continuous clusters of MCI-related functional factors: biological vitality, activity lifestyle, psychosocial affect, subjective health, and global cognition. We used a cross-sectional sample and a two-wave longitudinal sample, stratified by age (mid-old, old-old) and cognitive status (MCI, NIC). First, cross-sectional results showed that eight markers differentiated MCI and NIC adults, with the latter performing uniformly better. The groups differed on diastolic blood pressure, body mass index, positive and negative affect, MMSE, and the lifestyle indicators of self-maintenance, travel, and novel cognitive activities. Second, Wave 1 to Wave 2 stabilities in cognitive status classification were high. Third, several markers differentiated the stable (NIC-to-NIC, MCI-to-MCI) from the unstable (NIC-to-MCI, MCI-to-NIC) cognitive status groups. Fourth, five relevant markers for identifying older adults at risk for cognitive status changes were: diastolic blood pressure, self-maintenance activities, novel cognitive activities, positive affect, and global cognitive status. Selected risk and protective factors differentiate persons classified with MCI from those not currently cognitively impaired, both cross-sectionally and longitudinally.

  14. [Genetic diversity revealed by ISSR molecular marker in common wheat, spelt, compactum and progeny of recurrent selection].

    PubMed

    Du, Jin-Kun; Yao, Ying-Yin; Ni, Zhong-Fu; Peng, Hui-Ru; Sun, Qi-Xin

    2002-05-01

    It is important to estimate the genetic diversity between the parents for improving the heterosis of hybrid wheat. In this study, ISSR(inter-simple sequence repeat) marker was used to measure the genetic diversity within and among common wheat (Triticum aestivum L.), spelt (Triticum spelta L.), compactum (Triticum compactum Host.) and progeny of foreign wheat-based recurrent selection, and the possibility of establishing the new heterotic group was also assessed. Forty seven genotypes were used for ISSR analysis, which included 14 common wheat, 10 spelt wheat, 11 compactum and 12 progeny of recurrent selection. Eleven of 33 ISSR primers that can produce distinguishable bands were selected for PCR amplification. A total of 238 bands were amplified, among which 207 (87%) bands were polymorphic. The polymorphic bands amplified by each primer ranged from 11 to 38, with an averaged of 18.8. The percentage of polymorphic band (80.3%) in common wheat was higher than that in progeny of recurrent selection (78.7%), spelt (75.0%) and compactum (74.9%). The 238 polymorphic products were used to calculate Nei's similarity index (GS) and the genetic distance (GD). It was found that the mean genetic distance between different wheat types (0.3115-0.3442) was obviously higher than that within common wheat (0.2743), spelt (0.2351), compactum (0.2622). In addition, progeny of recurrent selection also showed much higher genetic distance with other three wheat types (0.3217, 0.3256, 0.3198). The cluster analysis was performed based on the genetic distance (GD) matrix by using UPGMA method. Common wheat, spelt, compactum and progeny of recurrent selection were classified into four different groups. In this study, ISSR marker was firstly used to assess genetic diversity among common wheat, spelt, compactum and progeny of recurrent selection, and can differentiate the wheat cultivars (lines) that selected from the same cross combination. It was concluded that spelt, compactum and progeny

  15. New Advances in Marker Assisted Selection for Winter Hardiness in Oats.

    USDA-ARS?s Scientific Manuscript database

    Oat (Avena sativa L.) breeding and genetics research has lagged behind other small grains, such as wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.), in the development of PCR based markers and map construction due to fewer oat researchers and reduced research funding. As a result, marke...

  16. Marker assisted selection of low phytic acid trait in maize (Zea mays L.).

    PubMed

    Sureshkumar, S; Tamilkumar, P; Senthil, N; Nagarajan, P; Thangavelu, A U; Raveendran, M; Vellaikumar, S; Ganesan, K N; Balagopal, R; Vijayalakshmi, G; Shobana, V

    2014-02-01

    Maize is the third important major food crop. Breeding for low phytate maize genotypes is an effective strategy for decreasing the content of kernel phytic acid (a chelator of cations such as Ca(2+) and Fe(3+) ) and thereby increasing the bioavailability of nutritive minerals in human diet and animal feed. Previous studies have established that a mutant plant with a lpa2-2 allele accumulates less phytic acid in seeds. Therefore, the marker assisted backcross breeding (MABB), which involves introgression of lpa2-2 recessive allele (which confer low phytate trait) from a lpa2-2 mutant line into a well-adapted line using backcrosses and selection of lines possessing lpa2-2 allele in each backcross population using molecular markers, is an effective strategy for developing low phytate maize. So far, no studies have developed any lpa2-2 allele specific molecular markers for this purpose. Here, using backcross and selfed progenies, obtained by crossing low phytate mutant line 'EC 659418' (i.e. donor of lpa2-2 allele) into agronomically superior line 'UMI395', we have validated that a SSR marker 'umc2230', located 0.4 cM downstream of lpa2-2, cosegregate, in a Mendelian fashion, with low phytic acid trait. Therefore umc2230 can be dependably used in MABB for the development of low phytate maize.

  17. Objective evaluation measures of genetic marker selection in large-scale SNP genotyping.

    PubMed

    Kaminuma, Eli; Masuya, Hiroshi; Miura, Ikuo; Motegi, Hiromi; Takahasi, Kenzi R; Nakazawa, Miki; Matsui, Minami; Gondo, Yoichi; Noda, Tetsuo; Shiroishi, Toshihiko; Wakana, Shigeharu; Toyoda, Tetsuro

    2008-10-01

    High-throughput single nucleotide polymorphism (SNP) genotyping systems provide two kinds of fluorescent signals detected from different alleles. In current technologies, the process of genotype discrimination requires subjective judgments by expert operators, even when using clustering algorithms. Here, we propose two evaluation measures to manage fluorescent scatter data with nonclear plot aggregation. The first is the marker ranking measure, which provides a ranking system for the SNP markers based on the distance between the scatter plot distribution and a user-defined ideal distribution. The second measure, called individual genotype membership, uses the membership probability of each genotype related to an individual plot in the scatter data. In verification experiments, the marker ranking measure determined the ranking of SNP markers correlated with the subjective order of SNP markers judged by an expert operator. The experiment using the individual genotype membership measure clarified that the total number of unclassified individuals was remarkably reduced compared to that of manually unclassified ones. These two evaluation measures were implemented as the GTAssist software. GTAssist provides objective standards and avoids subjective biases in SNP genotyping workflows.

  18. Evidence of Multiple Disease Resistance (MDR) and implication of meta-analysis in marker assisted selection.

    PubMed

    Ali, Farhan; Pan, Qingchun; Chen, Genshen; Zahid, Kashif Rafiq; Yan, Jianbing

    2013-01-01

    Meta-analysis was performed for three major foliar diseases with the aim to find out the total number of QTL responsible for these diseases and depict some real QTL for molecular breeding and marker assisted selection (MAS) in maize. Furthermore, we confirmed our results with some major known disease resistance genes and most well-known gene family of nucleotide binding site (NBS) encoding genes. Our analysis revealed that disease resistance QTL were randomly distributed in maize genome, but were clustered at different regions of the chromosomes. Totally 389 QTL were observed for these three major diseases in diverse maize germplasm, out of which 63 QTL were controlling more than one disease revealing the presence of multiple disease resistance (MDR). 44 real-QTLs were observed based on 4 QTL as standard in a specific region of genome. We also confirmed the Ht1 and Ht2 genes within the region of real QTL and 14 NBS-encoding genes. On chromosome 8 two NBS genes in one QTL were observed and on chromosome 3, several cluster and maximum MDR QTL were observed indicating that the apparent clustering could be due to genes exhibiting pleiotropic effect. Significant relationship was observed between the number of disease QTL and total genes per chromosome based on the reference genome B73. Therefore, we concluded that disease resistance genes are abundant in maize genome and these results can unleash the phenomenon of MDR. Furthermore, these results could be very handy to focus on hot spot on different chromosome for fine mapping of disease resistance genes and MAS.

  19. Contrasting responses to selection in class I and class IIα major histocompatibility-linked markers in salmon

    PubMed Central

    Consuegra, S; de Eyto, E; McGinnity, P; Stet, R J M; Jordan, W C

    2011-01-01

    Comparison of levels and patterns of genetic variation in natural populations either across loci or against neutral expectation can yield insight into locus-specific differences in the strength and direction of evolutionary forces. We used both approaches to test the hypotheses on patterns of selection on major histocompatibility (MH)-linked markers. We performed temporal analyses of class I and class IIα MH-linked markers and eight microsatellite loci in two Atlantic salmon populations in Ireland on two temporal scales: over six decades and 9 years in the rivers Burrishoole and Delphi, respectively. We also compared contemporary Burrishoole and Delphi samples with nearby populations for the same loci. On comparing patterns of temporal and spatial differentiation among classes of loci, the class IIα MH-linked marker was consistently identified as an outlier compared with patterns at the other microsatellite loci or neutral expectation. We found higher levels of temporal and spatial heterogeneity in heterozygosity (but not in allelic richness) for the class IIα MH-linked marker compared with microsatellites. Tests on both within- and among-population differentiation are consistent with directional selection acting on the class IIα-linked marker in both temporal and spatial comparisons, but only in temporal comparisons for the class I-linked marker. Our results indicate a complex pattern of selection on MH-linked markers in natural populations of Atlantic salmon. These findings highlight the importance of considering selection on MH-linked markers when using these markers for management and conservation purposes. PMID:21266985

  20. Identification and characterization of gene-based SSR markers in date palm (Phoenix dactylifera L.)

    PubMed Central

    2012-01-01

    Background Date palm (Phoenix dactylifera L.) is an important tree in the Middle East and North Africa due to the nutritional value of its fruit. Molecular Breeding would accelerate genetic improvement of fruit tree through marker assisted selection. However, the lack of molecular markers in date palm restricts the application of molecular breeding. Results In this study, we analyzed 28,889 EST sequences from the date palm genome database to identify simple-sequence repeats (SSRs) and to develop gene-based markers, i.e. expressed sequence tag-SSRs (EST-SSRs). We identified 4,609 ESTs as containing SSRs, among which, trinucleotide motifs (69.7%) were the most common, followed by tetranucleotide (10.4%) and dinucleotide motifs (9.6%). The motif AG (85.7%) was most abundant in dinucleotides, while motifs AGG (26.8%), AAG (19.3%), and AGC (16.1%) were most common among trinucleotides. A total of 4,967 primer pairs were designed for EST-SSR markers from the computational data. In a follow up laboratory study, we tested a sample of 20 random selected primer pairs for amplification and polymorphism detection using genomic DNA from date palm cultivars. Nearly one-third of these primer pairs detected DNA polymorphism to differentiate the twelve date palm cultivars used. Functional categorization of EST sequences containing SSRs revealed that 3,108 (67.4%) of such ESTs had homology with known proteins. Conclusion Date palm EST sequences exhibits a good resource for developing gene-based markers. These genic markers identified in our study may provide a valuable genetic and genomic tool for further genetic research and varietal development in date palm, such as diversity study, QTL mapping, and molecular breeding. PMID:23241238

  1. Development and application of SINE-based markers for genotyping of potato varieties.

    PubMed

    Seibt, Kathrin M; Wenke, Torsten; Wollrab, Cora; Junghans, Holger; Muders, Katja; Dehmer, Klaus J; Diekmann, Kerstin; Schmidt, Thomas

    2012-06-01

    Potato variety discrimination based on morphological traits is laborious and influenced by the environment, while currently applied molecular markers are either expensive or time-consuming in development or application. SINEs, short interspersed nuclear elements, are retrotransposons with a high copy number in plant genomes representing a potential source for new markers. We developed a marker system for potato genotyping, designated inter-SINE amplified polymorphism (ISAP). Based on nine potato SINE families recently characterized (Wenke et al. in Plant Cell 23:3117-3128, 2011), we designed species-specific SINE primers. From the resulting 153 primer combinations, highly informative primer sets were selected for potato variety analysis regarding number of bands, quality of the banding pattern, and the degree of polymorphism. Fragments representing ISAPs can be separated by conventional agarose gel electrophoresis; however, automation with a capillary sequencer is feasible. Two selected SINE families, SolS-IIIa and SolS-IV, were shown to be highly but differently amplified in Solanaceae, Solaneae tribe, including wild and cultivated potatoes, tomato, and eggplant. Fluorescent in situ hybridization demonstrated the genome-wide distribution of SolS-IIIa and SolS-IV along potato chromosomes, which is the basis for genotype discrimination and differentiation of somaclonal variants by ISAP markers.

  2. Base deficit as an early marker of coagulopathy in trauma.

    PubMed

    Cheddie, Shalen; Muckart, David J J; Hardcastle, Timothy C

    2013-07-31

    The acute coagulopathy of trauma is associated with hypoperfusion, metabolic acidosis and an increased mortality rate. Biochemical markers of hypoperfusion, namely base deficit (BD) and lactate, are commonly used to assess the degree of hypoperfusion. Early identification of hypoperfusion and acidosis using BD and lactate may help predict the development of coagulopathy in trauma patients and direct therapy. To identify whether a correlation exists between BD, lactate, injury severity, early-onset coagulopathy and mortality. A retrospective chart analysis was undertaken of patients transferred directly from scene to the level I trauma unit at Inkosi Albert Luthuli Central Hospital, Durban, South Africa, from 2007 to 2008. Patients with evidence of hypoperfusion were selected. Hypoperfusion was defined as a base deficit >-2 and coagulopathy as an International Normalized Ratio (INR) of >1.2. BD, lactate, chloride, temperature, Injury Severity Score (ISS), INR and mortality were recorded in this cohort. Student's t-test and Fisher's exact test were used for continuous and categorical variables, respectively. Correlation curves were used to determine the degree of association between the variables BD, lactate and ISS with respect to the INR. A p-value of <0.05 was considered statistically significant. Of the 28 patients, males (n=18) accounted for 64.3% of admissions. The mean age was 31 years (range 1 - 75 years, median 30 years). The mechanism of injury was penetrating trauma in 5 cases (17.9%) and blunt trauma in 23 (82.1%). The median ISS was 24 (range 4 - 59). In 16 patients (57.1%) the INR was within normal limits, but in 12 (42.9%) it was over 1.2. There was a significant correlation between BD, ISS and INR (r=0.393; p=0.019 and r=0.565, respectively; p<0.001). Lactate showed a weak and non-significant association with the INR (r=0.232; p=0.18). There were a total of 12 deaths (42.8%) in this cohort of patients with biochemical evidence of hypoperfusion. There

  3. Genetic diversity and parentage in farmer selections of cacao from Southern Sulawesi, Indonesia revealed by microsatellite markers.

    PubMed

    Dinarti, Diny; Susilo, Agung W; Meinhardt, Lyndel W; Ji, Kun; Motilal, Lambert A; Mischke, Sue; Zhang, Dapeng

    2015-12-01

    Indonesia is the third largest cocoa-producing country in the world. Knowledge of genetic diversity and parentage of farmer selections is important for effective selection and rational deployment of superior cacao clones in farmers' fields. We assessed genetic diversity and parentage of 53 farmer selections of cacao in Sulawesi, Indonesia, using 152 international clones as references. Cluster analysis, based on 15 microsatellite markers, showed that these Sulawesi farmer selections are mainly comprised of hybrids derived from Trinitario and two Upper Amazon Forastero groups. Bayesian assignment and likelihood-based parentage analysis further demonstrated that only a small number of germplasm groups, dominantly Trinitario and Parinari, contributed to these farmer selections, in spite of diverse parental clones having been used in the breeding program and seed gardens in Indonesia since the 1950s. The narrow parentage predicts a less durable host resistance to cacao diseases. Limited access of the farmers to diverse planting materials or the strong preference for large pods and large bean size by local farmers, may have affected the selection outcome. Diverse sources of resistance, harbored in different cacao germplasm groups, need to be effectively incorporated to broaden the on-farm diversity and ensure sustainable cacao production in Sulawesi.

  4. Genetic diversity and parentage in farmer selections of cacao from Southern Sulawesi, Indonesia revealed by microsatellite markers

    PubMed Central

    Dinarti, Diny; Susilo, Agung W.; Meinhardt, Lyndel W.; Ji, Kun; Motilal, Lambert A.; Mischke, Sue; Zhang, Dapeng

    2015-01-01

    Indonesia is the third largest cocoa-producing country in the world. Knowledge of genetic diversity and parentage of farmer selections is important for effective selection and rational deployment of superior cacao clones in farmers’ fields. We assessed genetic diversity and parentage of 53 farmer selections of cacao in Sulawesi, Indonesia, using 152 international clones as references. Cluster analysis, based on 15 microsatellite markers, showed that these Sulawesi farmer selections are mainly comprised of hybrids derived from Trinitario and two Upper Amazon Forastero groups. Bayesian assignment and likelihood-based parentage analysis further demonstrated that only a small number of germplasm groups, dominantly Trinitario and Parinari, contributed to these farmer selections, in spite of diverse parental clones having been used in the breeding program and seed gardens in Indonesia since the 1950s. The narrow parentage predicts a less durable host resistance to cacao diseases. Limited access of the farmers to diverse planting materials or the strong preference for large pods and large bean size by local farmers, may have affected the selection outcome. Diverse sources of resistance, harbored in different cacao germplasm groups, need to be effectively incorporated to broaden the on-farm diversity and ensure sustainable cacao production in Sulawesi. PMID:26719747

  5. [Valuation for usefulness of selected chromosomal markers for Bacillus anthracis identification. II. Valuation for markers SSH and rpoB].

    PubMed

    Zasada, Aleksandra Anna; Jagielski, Marek

    2006-01-01

    The article presents results of valuation for B. anthracis-specificity and usefulness for its identification obtained for different chromosomal markers. In the second part of the study markers SSH241, SSH196, SSH163, SSH133 as well as a fragment of the house-keeping gene rpoB were analyzed. For the investigation MSSCP and multiplex-PCR assays were used. There were also tested different techniques of electrophoresis. The results gave an information about specificity of tested markers and their usefulness for B. anthracis identification.

  6. Expression Marker-Based Strategy to Improve Beef Quality

    PubMed Central

    Cassar-Malek, Isabelle; Picard, Brigitte

    2016-01-01

    For beef cattle research, a main objective is to control concomitantly the development of muscles and the qualities of beef cuts. Beef quality is a complex phenotype that is only detectable after slaughter and is highly variable. The beef industry is in need of tools to estimate beef quality of live cattle or online in abattoirs, with specific attention towards sensory attributes (tenderness, juiciness, flavour, and colour). Identification of relevant genetic and genomic markers is ongoing, especially for tenderness—a top priority quality attribute. In this paper, we describe the steps of an expression marker-based strategy to improve beef sensory quality, from the discovery of biomarkers that identify consistent beef and the biological functions governing beef tenderness to the integration of the knowledge into detection tests for desirable animals. These tools should soon be available for the management of sensory quality in the beef production chain for meeting market's demands and assuring good quality standards. PMID:27066527

  7. Identification, utilisation and mapping of novel transcriptome-based markers from blackcurrant (Ribes nigrum)

    PubMed Central

    2011-01-01

    Background Deep-level second generation sequencing (2GS) technologies are now being applied to non-model species as a viable and favourable alternative to Sanger sequencing. Large-scale SNP discovery was undertaken in blackcurrant (Ribes nigrum L.) using transcriptome-based 2GS 454 sequencing on the parental genotypes of a reference mapping population, to generate large numbers of novel markers for the construction of a high-density linkage map. Results Over 700,000 reads were produced, from which a total of 7,000 SNPs were found. A subset of polymorphic SNPs was selected to develop a 384-SNP OPA assay using the Illumina BeadXpress platform. Additionally, the data enabled identification of 3,000 novel EST-SSRs. The selected SNPs and SSRs were validated across diverse Ribes germplasm, including mapping populations and other selected Ribes species. SNP-based maps were developed from two blackcurrant mapping populations, incorporating 48% and 27% of assayed SNPs respectively. A relatively high proportion of visually monomorphic SNPs were investigated further by quantitative trait mapping of theta score outputs from BeadStudio analysis, and this enabled additional SNPs to be placed on the two maps. Conclusions The use of 2GS technology for the development of markers is superior to previously described methods, in both numbers of markers and biological informativeness of those markers. Whilst the numbers of reads and assembled contigs were comparable to similar sized studies of other non-model species, here a high proportion of novel genes were discovered across a wide range of putative function and localisation. The potential utility of markers developed using the 2GS approach in downstream breeding applications is discussed. PMID:22035129

  8. Development of selective markers linked to a major QTL for parthenocarpy in eggplant (Solanum melongena L.).

    PubMed

    Miyatake, Koji; Saito, Takeo; Negoro, Satomi; Yamaguchi, Hirotaka; Nunome, Tsukasa; Ohyama, Akio; Fukuoka, Hiroyuki

    2012-05-01

    Parthenocarpy, the ability to set fruits without pollination, is a useful trait for setting fruit under unfavorable conditions. To identify the loci controlling parthenocarpy in eggplant (Solanum melongena L.), we constructed linkage maps by using co-dominant simple sequence repeat and single nucleotide polymorphism markers in F(2) populations derived from intraspecific crosses between two non-parthenocarpic lines (LS1934 and Nakate-Shinkuro) and a parthenocarpic line (AE-P03). Total map distances were 1,414.6 cM (ALF2: LS1934 x AE-P03) and 1,153.8 cM (NAF2: Nakate-Shinkuro x AE-P03), respectively. Quantitative trait locus (QTL) analyses revealed two QTLs on chromosomes 3 and 8, which we denoted as Controlling parthenocarpy3.1 (Cop3.1) and Cop8.1, respectively. The percentage of phenotypic variance explained (PVE) of Cop3.1 was 6.3% in ALF2 (LOD = 4.2) and 10.6% in NAF2 (LOD = 3.0). The PVE of Cop8.1 was 45.7% in ALF2 (LOD = 23.8) and 29.7% in NAF2 (LOD = 7.9). Using a population of backcross inbred lines, we confirmed the effect of Cop8.1, but there was no evidence to support the contribution of Cop3.1. We need to verify the effect of Cop3.1 under various temperature conditions. In addition, we clarified the effectiveness of selective SSR markers, emf21H22 and emh11J10, mapped on each side of Cop8.1 in other F(2) populations derived from various parental combinations. This is the first report concerning QTL analysis of parthenocarpy in eggplant using molecular markers. It will be useful in marker-assisted selection and in revealing the genomic mechanism underlying parthenocarpy in eggplant.

  9. Development of universal genetic markers based on single-copy orthologous (COSII) genes in Poaceae.

    PubMed

    Liu, Hailan; Guo, Xiaoqin; Wu, Jiasheng; Chen, Guo-Bo; Ying, Yeqing

    2013-03-01

    KEY MESSAGE : We develop a set of universal genetic markers based on single-copy orthologous (COSII) genes in Poaceae. Being evolutionary conserved, single-copy orthologous (COSII) genes are particularly useful in comparative mapping and phylogenetic investigation among species. In this study, we identified 2,684 COSII genes based on five sequenced Poaceae genomes including rice, maize, sorghum, foxtail millet, and brachypodium, and then developed 1,072 COSII markers whose transferability and polymorphism among five bamboo species were further evaluated with 46 pairs of randomly selected primers. 91.3 % of the 46 primers obtained clear amplification in at least one bamboo species, and 65.2 % of them produced polymorphism in more than one species. We also used 42 of them to construct the phylogeny for the five bamboo species, and it might reflect more precise evolutionary relationship than the one based on the vegetative morphology. The results indicated a promising prospect of applying these markers to the investigation of genetic diversity and the classification of Poaceae. To ease and facilitate access of the information of common interest to readers, a web-based database of the COSII markers is provided ( http://www.sicau.edu.cn/web/yms/PCOSWeb/PCOS.html ).

  10. Salt tolerance in Lycopersicon species. IV. Efficiency of marker-assisted selection for salt tolerance improvement.

    PubMed

    Monforte, A J; Asíns, M J; Carbonell, E A

    1996-10-01

    The usefulness of marker-assisted selection (MAS) to develop salt-tolerant breeding lines from a F2 derived from L. esculentum x L. pimpinellifolium has been studied. Interval mapping methodology of quantitative trait locus (QTL) analysis was used to locate more precisely previously detected salt tolerance QTLs. A new QTL for total fruit weight under salinity (TW) near TG24 was detected. Most of the detected QTLs [3 for TW, 5 for fruit number, (FN) and 4 for fruit weight (FW)] had low R (2) values, except the FW QTL in the TG180-TG48 interval, which explains 36.6% of the total variance. Dominant and overdominant effects were detected at the QTLs for TW, whereas gene effects at the QTLs for FJV and FW ranged from additive to partial dominance. Phenotypic selection of F2 familes and marker-assisted selection of F3 families were carried out. Yield under salinity decreased in the F2 generation. F3 means were similar to those of the F1 as a consequence of phentoypic selection. The most important selection response for every trait was obtained from the F3 to F4 where MAS was applied. While F3 variation was mainly due to the within-family component, in the F4 the FN and FW between-family component was larger than the within-family one, indicating an efficient compartmentalization and fixation of QTLs into the F4 families. Comparison of the yield of these families under control versus saline conditions showed that fruit weight is a key trait to success in tomato salt-tolerance improvement using wild Lycopersicon germplasm. The QTLs we have detected under salinity seem to be also working under control conditions, although the interaction family x treatment was significant for TW, thereby explaining the fact that the selected families responded differently to salinity.

  11. Segregation and genetic linkage analyses of river catfish, Mystus nemurus, based on microsatellite markers.

    PubMed

    Hoh, B P; Siraj, S S; Tan, S G; Yusoff, K

    2013-02-28

    The river catfish Mystus nemurus is an important fresh water species for aquaculture in Malaysia. We report the first genetic linkage map of M. nemurus based on segregation analysis and a linkage map using newly developed microsatellite markers of M. nemurus. A total of 70 of the newly developed polymorphic DNA microsatellite markers were analyzed on pedigrees generated using a pseudo-testcross strategy from 2 mapping families. In the first mapping family, 100 offspring were produced from randomly selected dams of the same populations; dams of the second family were selected from 2 different populations, and this family had 50 offspring. Thirty-one of the 70 markers segregated according to the Mendelian segregation ratio. Linkage analysis revealed that 17 microsatellite markers belonging to 7 linkage groups were obtained at a logarithm of the odds score of 1.2 spanning 584 cM by the Kosambi mapping function, whereas the other 14 remained unlinked. The results from this study will act as primer to a more extensive genetic mapping study aimed towards identifying genetic loci involved in determining economically important traits.

  12. Successful recovery of transgenic cowpea (Vigna unguiculata) using the 6-phosphomannose isomerase gene as the selectable marker.

    PubMed

    Bakshi, Souvika; Saha, Bedabrata; Roy, Nand Kishor; Mishra, Sagarika; Panda, Sanjib Kumar; Sahoo, Lingaraj

    2012-06-01

    A new method for obtaining transgenic cowpea was developed using positive selection based on the Escherichia coli 6-phosphomannose isomerase gene as the selectable marker and mannose as the selective agent. Only transformed cells were capable of utilizing mannose as a carbon source. Cotyledonary node explants from 4-day-old in vitro-germinated seedlings of cultivar Pusa Komal were inoculated with Agrobacterium tumefaciens strain EHA105 carrying the vector pNOV2819. Regenerating transformed shoots were selected on medium supplemented with a combination of 20 g/l mannose and 5 g/l sucrose as carbon source. The transformed shoots were rooted on medium devoid of mannose. Transformation efficiency based on PCR analysis of individual putative transformed shoots was 3.6%. Southern blot analysis on five randomly chosen PCR-positive plants confirmed the integration of the pmi transgene. Qualitative reverse transcription (qRT-PCR) analysis demonstrated the expression of pmi in T₀ transgenic plants. Chlorophenol red (CPR) assays confirmed the activity of PMI in transgenic plants, and the gene was transmitted to progeny in a Mendelian fashion. The transformation method presented here for cowpea using mannose selection is efficient and reproducible, and could be used to introduce a desirable gene(s) into cowpea for biotic and abiotic stress tolerance.

  13. DNA-Based Genetic Markers for Rapid Cycling Brassica Rapa (Fast Plants Type) Designed for the Teaching Laboratory.

    PubMed

    Slankster, Eryn E; Chase, Jillian M; Jones, Lauren A; Wendell, Douglas L

    2012-01-01

    We have developed DNA-based genetic markers for rapid cycling Brassica rapa (RCBr), also known as Fast Plants. Although markers for B. rapa already exist, ours were intentionally designed for use in a teaching laboratory environment. The qualities we selected for were robust amplification in PCR, polymorphism in RCBr strains, and alleles that can be easily resolved in simple agarose slab gels. We have developed two single nucleotide polymorphism (SNP) based markers and 14 variable number tandem repeat (VNTR)-type markers spread over four chromosomes. The DNA sequences of these markers represent variation in a wide range of genomic features. Among the VNTR-type markers, there are examples of variation in a non-genic region, variation within an intron, and variation in the coding sequence of a gene. Among the SNP-based markers there are examples of polymorphism in intronic DNA and synonymous substitution in a coding sequence. Thus these markers can serve laboratory exercises in both transmission genetics and molecular biology.

  14. DNA-Based Genetic Markers for Rapid Cycling Brassica Rapa (Fast Plants Type) Designed for the Teaching Laboratory

    PubMed Central

    Slankster, Eryn E.; Chase, Jillian M.; Jones, Lauren A.; Wendell, Douglas L.

    2012-01-01

    We have developed DNA-based genetic markers for rapid cycling Brassica rapa (RCBr), also known as Fast Plants. Although markers for B. rapa already exist, ours were intentionally designed for use in a teaching laboratory environment. The qualities we selected for were robust amplification in PCR, polymorphism in RCBr strains, and alleles that can be easily resolved in simple agarose slab gels. We have developed two single nucleotide polymorphism (SNP) based markers and 14 variable number tandem repeat (VNTR)-type markers spread over four chromosomes. The DNA sequences of these markers represent variation in a wide range of genomic features. Among the VNTR-type markers, there are examples of variation in a non-genic region, variation within an intron, and variation in the coding sequence of a gene. Among the SNP-based markers there are examples of polymorphism in intronic DNA and synonymous substitution in a coding sequence. Thus these markers can serve laboratory exercises in both transmission genetics and molecular biology. PMID:22675329

  15. Improved age determination of blood and teeth samples using a selected set of DNA methylation markers

    PubMed Central

    Kamalandua, Aubeline

    2015-01-01

    Age estimation from DNA methylation markers has seen an exponential growth of interest, not in the least from forensic scientists. The current published assays, however, can still be improved by lowering the number of markers in the assay and by providing more accurate models to predict chronological age. From the published literature we selected 4 age-associated genes (ASPA, PDE4C, ELOVL2, and EDARADD) and determined CpG methylation levels from 206 blood samples of both deceased and living individuals (age range: 0–91 years). This data was subsequently used to compare prediction accuracy with both linear and non-linear regression models. A quadratic regression model in which the methylation levels of ELOVL2 were squared showed the highest accuracy with a Mean Absolute Deviation (MAD) between chronological age and predicted age of 3.75 years and an adjusted R2 of 0.95. No difference in accuracy was observed for samples obtained either from living and deceased individuals or between the 2 genders. In addition, 29 teeth from different individuals (age range: 19–70 years) were analyzed using the same set of markers resulting in a MAD of 4.86 years and an adjusted R2 of 0.74. Cross validation of the results obtained from blood samples demonstrated the robustness and reproducibility of the assay. In conclusion, the set of 4 CpG DNA methylation markers is capable of producing highly accurate age predictions for blood samples from deceased and living individuals PMID:26280308

  16. Fine Mapping for Identification of Citrus Alternaria Brown Spot Candidate Resistance Genes and Development of New SNP Markers for Marker-Assisted Selection.

    PubMed

    Cuenca, Jose; Aleza, Pablo; Garcia-Lor, Andres; Ollitrault, Patrick; Navarro, Luis

    2016-01-01

    Alternaria brown spot (ABS) is a serious disease affecting susceptible citrus genotypes, which is a strong concern regarding citrus breeding programs. Resistance is conferred by a recessive locus (ABSr) previously located by our group within a 3.3 Mb genome region near the centromere in chromosome III. This work addresses fine-linkage mapping of this region for identifying candidate resistance genes and develops new molecular markers for ABS-resistance effective marker-assisted selection (MAS). Markers closely linked to ABSr locus were used for fine mapping using a 268-segregating diploid progeny derived from a heterozygous susceptible × resistant cross. Fine mapping limited the genomic region containing the ABSr resistance gene to 366 kb, flanked by markers at 0.4 and 0.7 cM. This region contains nine genes related to pathogen resistance. Among them, eight are resistance (R) gene homologs, with two of them harboring a serine/threonine protein kinase domain. These two genes along with a gene encoding a S-adenosyl-L-methionine-dependent-methyltransferase protein, should be considered as strong candidates for ABS-resistance. Moreover, the closest SNP was genotyped in 40 citrus varieties, revealing very high association with the resistant/susceptible phenotype. This new marker is currently used in our citrus breeding program for ABS-resistant parent and cultivar selection, at diploid, triploid and tetraploid level.

  17. Fine Mapping for Identification of Citrus Alternaria Brown Spot Candidate Resistance Genes and Development of New SNP Markers for Marker-Assisted Selection

    PubMed Central

    Cuenca, Jose; Aleza, Pablo; Garcia-Lor, Andres; Ollitrault, Patrick; Navarro, Luis

    2016-01-01

    Alternaria brown spot (ABS) is a serious disease affecting susceptible citrus genotypes, which is a strong concern regarding citrus breeding programs. Resistance is conferred by a recessive locus (ABSr) previously located by our group within a 3.3 Mb genome region near the centromere in chromosome III. This work addresses fine-linkage mapping of this region for identifying candidate resistance genes and develops new molecular markers for ABS-resistance effective marker-assisted selection (MAS). Markers closely linked to ABSr locus were used for fine mapping using a 268-segregating diploid progeny derived from a heterozygous susceptible × resistant cross. Fine mapping limited the genomic region containing the ABSr resistance gene to 366 kb, flanked by markers at 0.4 and 0.7 cM. This region contains nine genes related to pathogen resistance. Among them, eight are resistance (R) gene homologs, with two of them harboring a serine/threonine protein kinase domain. These two genes along with a gene encoding a S-adenosyl-L-methionine-dependent-methyltransferase protein, should be considered as strong candidates for ABS-resistance. Moreover, the closest SNP was genotyped in 40 citrus varieties, revealing very high association with the resistant/susceptible phenotype. This new marker is currently used in our citrus breeding program for ABS-resistant parent and cultivar selection, at diploid, triploid and tetraploid level. PMID:28066498

  18. Comparison of phenotypic versus marker-assisted background selection for the SUB1 QTL during backcrossing in rice

    PubMed Central

    Iftekharuddaula, Khandakar M.; Salam, Muhammad A.; Newaz, Muhammad A.; Ahmed, Helal U.; Collard, Bertrand C. Y.; Septiningsih, Endang M.; Sanchez, Darlene L.; Pamplona, Alvaro M.; Mackill, David J.

    2012-01-01

    Marker assisted backcrossing has been used effectively to transfer the submergence tolerance gene SUB1 into popular rice varieties, but the approach can be costly. The selection strategy comprising foreground marker and phenotypic selection was investigated as an alternative. The non-significant correlation coefficients between ranking of phenotypic selection and ranking of background marker selection in BC2F1, BC3F1 and BC3F2 generations indicated inefficiency of phenotypic selection compared to marker-assisted background selection with respect to recovery of the recipient genome. In addition, the introgression size of the chromosome fragment containing SUB1 was approximately 17 Mb, showing the effects of linkage drag. The significant correlation coefficient between rankings of phenotypic selection with the percentage of recipient alleles in the BC1F1 generation suggested that background selection could be avoided in this generation to minimize the genotyping cost. The phenotypically selected best plant of the BC3F1 generation was selfed and backcross recombinant lines were selected in the resulting BC3F4 generation. The selection strategy could be appropriate for the introgression of SUB1 QTL in countries that lack access to high-throughput genotyping facilities. PMID:23226081

  19. ABCG2 is a selectable marker for enhanced multilineage differentiation potential in periodontal ligament stem cells.

    PubMed

    Szepesi, Áron; Matula, Zsolt; Szigeti, Anna; Várady, György; Szabó, Gyula; Uher, Ferenc; Sarkadi, Balázs; Német, Katalin

    2015-01-15

    Periodontal ligament stem cells (PDLSCs) provide an important source for tissue regeneration and may become especially useful in the formation of osteogenic seeds. PDLSCs can be cultured, expanded, and differentiated in vitro; thus, they may be applied in the long-term treatment of the defects in the dental regions. Here we studied numerous potential markers allowing the selection of human PDLSCs with a maximum differentiation potential. We followed the expression of the ATP-binding cassette subfamily G member 2 (ABCG2) membrane transporter protein and isolated ABCG2-expressing cells by using a monoclonal antibody, recognizing the transporter at the cell surface in intact cells. The expression of the ABCG2 protein, corresponding to the so-called side-population phenotype in various tissue-derived stem cells, was found to be a useful marker for the selection of PDLSCs with enhanced osteogenic, chondrogenic, and adipogenic differentiation. These findings may have important applications in achieving efficient dental tissue regeneration by using stem cells from extracted teeth.

  20. ABCG2 Is a Selectable Marker for Enhanced Multilineage Differentiation Potential in Periodontal Ligament Stem Cells

    PubMed Central

    Szepesi, Áron; Matula, Zsolt; Szigeti, Anna; Várady, György; Szabó, Gyula; Uher, Ferenc; Sarkadi, Balázs

    2015-01-01

    Periodontal ligament stem cells (PDLSCs) provide an important source for tissue regeneration and may become especially useful in the formation of osteogenic seeds. PDLSCs can be cultured, expanded, and differentiated in vitro; thus, they may be applied in the long-term treatment of the defects in the dental regions. Here we studied numerous potential markers allowing the selection of human PDLSCs with a maximum differentiation potential. We followed the expression of the ATP-binding cassette subfamily G member 2 (ABCG2) membrane transporter protein and isolated ABCG2-expressing cells by using a monoclonal antibody, recognizing the transporter at the cell surface in intact cells. The expression of the ABCG2 protein, corresponding to the so-called side-population phenotype in various tissue-derived stem cells, was found to be a useful marker for the selection of PDLSCs with enhanced osteogenic, chondrogenic, and adipogenic differentiation. These findings may have important applications in achieving efficient dental tissue regeneration by using stem cells from extracted teeth. PMID:25101689

  1. The use and abuse of genetic marker-based estimates of relatedness and inbreeding

    PubMed Central

    Taylor, Helen R

    2015-01-01

    Genetic marker-based estimators remain a popular tool for measuring relatedness (rxy) and inbreeding (F) coefficients at both the population and individual level. The performance of these estimators fluctuates with the number and variability of markers available, and the relatedness composition and demographic history of a population. Several methods are available to evaluate the reliability of the estimates of rxy and F, some of which are implemented in the program COANCESTRY. I used the simulation module in COANCESTRY since assess the performance of marker-based estimators of rxy and F in a species with very low genetic diversity, New Zealand’s little spotted kiwi (Apteryx owenii). I also conducted a review of published papers that have used COANCESTRY as its release to assess whether and how the reliability of the estimates of rxy and F produced by genetic markers are being measured and reported in published studies. My simulation results show that even when the correlation between true (simulated) and estimated rxy or F is relatively high (Pearson’s r = 0.66–0.72 and 0.81–0.85, respectively) the imprecision of the estimates renders them highly unreliable on an individual basis. The literature review demonstrates that the majority of studies do not report the reliability of marker-based estimates of rxy and F. There is currently no standard practice for selecting the best estimator for a given data set or reporting an estimator’s performance. This could lead to experimental results being interpreted out of context and render the robustness of conclusions based on measures of rxy and F debatable. PMID:26357542

  2. The use and abuse of genetic marker-based estimates of relatedness and inbreeding.

    PubMed

    Taylor, Helen R

    2015-08-01

    Genetic marker-based estimators remain a popular tool for measuring relatedness (r xy ) and inbreeding (F) coefficients at both the population and individual level. The performance of these estimators fluctuates with the number and variability of markers available, and the relatedness composition and demographic history of a population. Several methods are available to evaluate the reliability of the estimates of r xy and F, some of which are implemented in the program COANCESTRY. I used the simulation module in COANCESTRY since assess the performance of marker-based estimators of r xy and F in a species with very low genetic diversity, New Zealand's little spotted kiwi (Apteryx owenii). I also conducted a review of published papers that have used COANCESTRY as its release to assess whether and how the reliability of the estimates of r xy and F produced by genetic markers are being measured and reported in published studies. My simulation results show that even when the correlation between true (simulated) and estimated r xy or F is relatively high (Pearson's r = 0.66-0.72 and 0.81-0.85, respectively) the imprecision of the estimates renders them highly unreliable on an individual basis. The literature review demonstrates that the majority of studies do not report the reliability of marker-based estimates of r xy and F. There is currently no standard practice for selecting the best estimator for a given data set or reporting an estimator's performance. This could lead to experimental results being interpreted out of context and render the robustness of conclusions based on measures of r xy and F debatable.

  3. A next-generation marker genotyping platform (AmpSeq) in heterozygous crops: a case study for marker-assisted selection in grapevine.

    PubMed

    Yang, Shanshan; Fresnedo-Ramírez, Jonathan; Wang, Minghui; Cote, Linda; Schweitzer, Peter; Barba, Paola; Takacs, Elizabeth M; Clark, Matthew; Luby, James; Manns, David C; Sacks, Gavin; Mansfield, Anna Katharine; Londo, Jason; Fennell, Anne; Gadoury, David; Reisch, Bruce; Cadle-Davidson, Lance; Sun, Qi

    2016-01-01

    Marker-assisted selection (MAS) is often employed in crop breeding programs to accelerate and enhance cultivar development, via selection during the juvenile phase and parental selection prior to crossing. Next-generation sequencing and its derivative technologies have been used for genome-wide molecular marker discovery. To bridge the gap between marker development and MAS implementation, this study developed a novel practical strategy with a semi-automated pipeline that incorporates trait-associated single nucleotide polymorphism marker discovery, low-cost genotyping through amplicon sequencing (AmpSeq) and decision making. The results document the development of a MAS package derived from genotyping-by-sequencing using three traits (flower sex, disease resistance and acylated anthocyanins) in grapevine breeding. The vast majority of sequence reads (⩾99%) were from the targeted regions. Across 380 individuals and up to 31 amplicons sequenced in each lane of MiSeq data, most amplicons (83 to 87%) had <10% missing data, and read depth had a median of 220-244×. Several strengths of the AmpSeq platform that make this approach of broad interest in diverse crop species include accuracy, flexibility, speed, high-throughput, low-cost and easily automated analysis.

  4. A next-generation marker genotyping platform (AmpSeq) in heterozygous crops: a case study for marker-assisted selection in grapevine

    PubMed Central

    Yang, Shanshan; Fresnedo-Ramírez, Jonathan; Wang, Minghui; Cote, Linda; Schweitzer, Peter; Barba, Paola; Takacs, Elizabeth M; Clark, Matthew; Luby, James; Manns, David C; Sacks, Gavin; Mansfield, Anna Katharine; Londo, Jason; Fennell, Anne; Gadoury, David; Reisch, Bruce; Cadle-Davidson, Lance; Sun, Qi

    2016-01-01

    Marker-assisted selection (MAS) is often employed in crop breeding programs to accelerate and enhance cultivar development, via selection during the juvenile phase and parental selection prior to crossing. Next-generation sequencing and its derivative technologies have been used for genome-wide molecular marker discovery. To bridge the gap between marker development and MAS implementation, this study developed a novel practical strategy with a semi-automated pipeline that incorporates trait-associated single nucleotide polymorphism marker discovery, low-cost genotyping through amplicon sequencing (AmpSeq) and decision making. The results document the development of a MAS package derived from genotyping-by-sequencing using three traits (flower sex, disease resistance and acylated anthocyanins) in grapevine breeding. The vast majority of sequence reads (⩾99%) were from the targeted regions. Across 380 individuals and up to 31 amplicons sequenced in each lane of MiSeq data, most amplicons (83 to 87%) had <10% missing data, and read depth had a median of 220–244×. Several strengths of the AmpSeq platform that make this approach of broad interest in diverse crop species include accuracy, flexibility, speed, high-throughput, low-cost and easily automated analysis. PMID:27257505

  5. Genomic relationships based on X chromosome markers and accuracy of genomic predictions with and without X chromosome markers

    PubMed Central

    2014-01-01

    Background Although the X chromosome is the second largest bovine chromosome, markers on the X chromosome are not used for genomic prediction in some countries and populations. In this study, we presented a method for computing genomic relationships using X chromosome markers, investigated the accuracy of imputation from a low density (7K) to the 54K SNP (single nucleotide polymorphism) panel, and compared the accuracy of genomic prediction with and without using X chromosome markers. Methods The impact of considering X chromosome markers on prediction accuracy was assessed using data from Nordic Holstein bulls and different sets of SNPs: (a) the 54K SNPs for reference and test animals, (b) SNPs imputed from the 7K to the 54K SNP panel for test animals, (c) SNPs imputed from the 7K to the 54K panel for half of the reference animals, and (d) the 7K SNP panel for all animals. Beagle and Findhap were used for imputation. GBLUP (genomic best linear unbiased prediction) models with or without X chromosome markers and with or without a residual polygenic effect were used to predict genomic breeding values for 15 traits. Results Averaged over the two imputation datasets, correlation coefficients between imputed and true genotypes for autosomal markers, pseudo-autosomal markers, and X-specific markers were 0.971, 0.831 and 0.935 when using Findhap, and 0.983, 0.856 and 0.937 when using Beagle. Estimated reliabilities of genomic predictions based on the imputed datasets using Findhap or Beagle were very close to those using the real 54K data. Genomic prediction using all markers gave slightly higher reliabilities than predictions without X chromosome markers. Based on our data which included only bulls, using a G matrix that accounted for sex-linked relationships did not improve prediction, compared with a G matrix that did not account for sex-linked relationships. A model that included a polygenic effect did not recover the loss of prediction accuracy from exclusion of X

  6. Genomic relationships based on X chromosome markers and accuracy of genomic predictions with and without X chromosome markers.

    PubMed

    Su, Guosheng; Guldbrandtsen, Bernt; Aamand, Gert P; Strandén, Ismo; Lund, Mogens S

    2014-07-30

    Although the X chromosome is the second largest bovine chromosome, markers on the X chromosome are not used for genomic prediction in some countries and populations. In this study, we presented a method for computing genomic relationships using X chromosome markers, investigated the accuracy of imputation from a low density (7K) to the 54K SNP (single nucleotide polymorphism) panel, and compared the accuracy of genomic prediction with and without using X chromosome markers. The impact of considering X chromosome markers on prediction accuracy was assessed using data from Nordic Holstein bulls and different sets of SNPs: (a) the 54K SNPs for reference and test animals, (b) SNPs imputed from the 7K to the 54K SNP panel for test animals, (c) SNPs imputed from the 7K to the 54K panel for half of the reference animals, and (d) the 7K SNP panel for all animals. Beagle and Findhap were used for imputation. GBLUP (genomic best linear unbiased prediction) models with or without X chromosome markers and with or without a residual polygenic effect were used to predict genomic breeding values for 15 traits. Averaged over the two imputation datasets, correlation coefficients between imputed and true genotypes for autosomal markers, pseudo-autosomal markers, and X-specific markers were 0.971, 0.831 and 0.935 when using Findhap, and 0.983, 0.856 and 0.937 when using Beagle. Estimated reliabilities of genomic predictions based on the imputed datasets using Findhap or Beagle were very close to those using the real 54K data. Genomic prediction using all markers gave slightly higher reliabilities than predictions without X chromosome markers. Based on our data which included only bulls, using a G matrix that accounted for sex-linked relationships did not improve prediction, compared with a G matrix that did not account for sex-linked relationships. A model that included a polygenic effect did not recover the loss of prediction accuracy from exclusion of X chromosome markers. The

  7. A Molecular Selection Index Method Based on Eigenanalysis

    PubMed Central

    Cerón-Rojas, J. Jesús; Castillo-González, Fernando; Sahagún-Castellanos, Jaime; Santacruz-Varela, Amalio; Benítez-Riquelme, Ignacio; Crossa, José

    2008-01-01

    The traditional molecular selection index (MSI) employed in marker-assisted selection maximizes the selection response by combining information on molecular markers linked to quantitative trait loci (QTL) and phenotypic values of the traits of the individuals of interest. This study proposes an MSI based on an eigenanalysis method (molecular eigen selection index method, MESIM), where the first eigenvector is used as a selection index criterion, and its elements determine the proportion of the trait's contribution to the selection index. This article develops the theoretical framework of MESIM. Simulation results show that the genotypic means and the expected selection response from MESIM for each trait are equal to or greater than those from the traditional MSI. When several traits are simultaneously selected, MESIM performs well for traits with relatively low heritability. The main advantages of MESIM over the traditional molecular selection index are that its statistical sampling properties are known and that it does not require economic weights and thus can be used in practical applications when all or some of the traits need to be improved simultaneously. PMID:18716338

  8. A novel homologous dominant selection marker for genetic transformation of Penicillium chrysogenum: overexpression of squalene epoxidase-encoding ergA.

    PubMed

    Sigl, Claudia; Handler, Monika; Sprenger, Georg; Kürnsteiner, Hubert; Zadra, Ivo

    2010-11-01

    Genetic engineering requires genetic selection markers. For generation of biosafe strains in industrial applications, homologous dominant selection markers allowing "self-cloning" are best suited but scarce. Here we describe a novel homologous dominant genetic selection system for the filamentous fungus Penicillium chrysogenum based on overexpression of the P. chrysogenum squalene epoxidase-encoding ergA gene, which confers resistance against terbinafine. Terbinafine (TRB) is a potent antifungal drug used in therapy of fungal infections. Overexpression of ergA was driven by the P. chrysogenum endoxylanase xylP promoter that is highly inducible by xylose. The suitability of the novel selection marker cassette for genetic manipulation was proven by its use for targeted deletion of the transcription factor nosA in P. chrysogenum. NosA-deficiency did not affect growth rates on solid or in liquid media, conidiation in light or darkness, and resistance to hydrogen peroxide. However, NosA-deficiency significantly decreased penicillin productivity. As TRB inhibits the growth of a variety of fungal species, this novel selection marker is expected to be suitable for genetic engineering of diverse fungal species.

  9. Development of a RAD-Seq Based DNA Polymorphism Identification Software, AgroMarker Finder, and Its Application in Rice Marker-Assisted Breeding.

    PubMed

    Fan, Wei; Zong, Jie; Luo, Zhijing; Chen, Mingjiao; Zhao, Xiangxiang; Zhang, Dabing; Qi, Yiping; Yuan, Zheng

    2016-01-01

    Rapid and accurate genome-wide marker detection is essential to the marker-assisted breeding and functional genomics studies. In this work, we developed an integrated software, AgroMarker Finder (AMF: http://erp.novelbio.com/AMF), for providing graphical user interface (GUI) to facilitate the recently developed restriction-site associated DNA (RAD) sequencing data analysis in rice. By application of AMF, a total of 90,743 high-quality markers (82,878 SNPs and 7,865 InDels) were detected between rice varieties JP69 and Jiaoyuan5A. The density of the identified markers is 0.2 per Kb for SNP markers, and 0.02 per Kb for InDel markers. Sequencing validation revealed that the accuracy of genome-wide marker detection by AMF is 93%. In addition, a validated subset of 82 SNPs and 31 InDels were found to be closely linked to 117 important agronomic trait genes, providing a basis for subsequent marker-assisted selection (MAS) and variety identification. Furthermore, we selected 12 markers from 31 validated InDel markers to identify seed authenticity of variety Jiaoyuanyou69, and we also identified 10 markers closely linked to the fragrant gene BADH2 to minimize linkage drag for Wuxiang075 (BADH2 donor)/Jiachang1 recombinants selection. Therefore, this software provides an efficient approach for marker identification from RAD-seq data, and it would be a valuable tool for plant MAS and variety protection.

  10. Development of a RAD-Seq Based DNA Polymorphism Identification Software, AgroMarker Finder, and Its Application in Rice Marker-Assisted Breeding

    PubMed Central

    Luo, Zhijing; Chen, Mingjiao; Zhao, Xiangxiang; Zhang, Dabing; Qi, Yiping; Yuan, Zheng

    2016-01-01

    Rapid and accurate genome-wide marker detection is essential to the marker-assisted breeding and functional genomics studies. In this work, we developed an integrated software, AgroMarker Finder (AMF: http://erp.novelbio.com/AMF), for providing graphical user interface (GUI) to facilitate the recently developed restriction-site associated DNA (RAD) sequencing data analysis in rice. By application of AMF, a total of 90,743 high-quality markers (82,878 SNPs and 7,865 InDels) were detected between rice varieties JP69 and Jiaoyuan5A. The density of the identified markers is 0.2 per Kb for SNP markers, and 0.02 per Kb for InDel markers. Sequencing validation revealed that the accuracy of genome-wide marker detection by AMF is 93%. In addition, a validated subset of 82 SNPs and 31 InDels were found to be closely linked to 117 important agronomic trait genes, providing a basis for subsequent marker-assisted selection (MAS) and variety identification. Furthermore, we selected 12 markers from 31 validated InDel markers to identify seed authenticity of variety Jiaoyuanyou69, and we also identified 10 markers closely linked to the fragrant gene BADH2 to minimize linkage drag for Wuxiang075 (BADH2 donor)/Jiachang1 recombinants selection. Therefore, this software provides an efficient approach for marker identification from RAD-seq data, and it would be a valuable tool for plant MAS and variety protection. PMID:26799713

  11. Integration of DNA into bacterial chromosomes from plasmids without a counter-selection marker

    PubMed Central

    Heap, John T.; Ehsaan, Muhammad; Cooksley, Clare M.; Ng, Yen-Kuan; Cartman, Stephen T.; Winzer, Klaus; Minton, Nigel P.

    2012-01-01

    Most bacteria can only be transformed with circular plasmids, so robust DNA integration methods for these rely upon selection of single-crossover clones followed by counter-selection of double-crossover clones. To overcome the limited availability of heterologous counter-selection markers, here we explore novel DNA integration strategies that do not employ them, and instead exploit (i) activation or inactivation of genes leading to a selectable phenotype, and (ii) asymmetrical regions of homology to control the order of recombination events. We focus here on the industrial biofuel-producing bacterium Clostridium acetobutylicum, which previously lacked robust integration tools, but the approach we have developed is broadly applicable. Large sequences can be delivered in a series of steps, as we demonstrate by inserting the chromosome of phage lambda (minus a region apparently unstable in Escherichia coli in our cloning context) into the chromosome of C. acetobutylicum in three steps. This work should open the way to reliable integration of DNA including large synthetic constructs in diverse microorganisms. PMID:22259038

  12. Molecular markers for identifying a new selected variety of Pacific white shrimp Litopenaeus vannamei

    NASA Astrophysics Data System (ADS)

    Yu, Yang; Zhang, Xiaojun; Liu, Jingwen; Li, Fuhua; Huang, Hao; Li, Yijun; Liu, Xiaolin; Xiang, Jianhai

    2015-01-01

    Selective breeding of the Pacific white shrimp Litopenaeus vannamei during the last decade has produced new varieties exhibiting high growth rates and disease resistance. However, the identification of new varieties of shrimps from their phenotypic characters is difficult. This study introduces a new approach for identifying varieties of shrimps using molecular markers of microsatellites and mitochondrial control region sequences. The method was employed to identify a new selected variety, Kehai No. 1 (KH-1), from three representative stocks (control group): Zhengda; Tongwei; and a stock collected from Fujian Province, which is now cultured in mainland China. By pooled genotyping of KH-1 and the control group, five microsatellites showing differences between KH-1 and the control group were screened out. Individual genotyping data confirmed the results from pooled genotyping. The genotyping data for the five microsatellites were applied to the assignment analysis of the KH-1 group and the control group using the partial Bayesian assignment method in GENECLASS2. By sequencing the mitochondrial control regions of individuals from the KH-1 and control group, four haplotypes were observed in the KH-1 group, whereas 14 haplotypes were obtained in the control group. By combining the microsatellite assignment analysis with mitochondrial control region analysis, the average accuracy of identification of individuals in the KH-1 group and control group reached 89%. The five selected microsatellite loci and mitochondrial control region sequences were highly polymorphic and could be used to distinguish new selected varieties of L. vannamei from other populations cultured in China.

  13. Glyphosate Resistance as a Novel Select-Agent-Compliant, Non-Antibiotic-Selectable Marker in Chromosomal Mutagenesis of the Essential Genes asd and dapB of Burkholderia pseudomallei▿

    PubMed Central

    Norris, Michael H.; Kang, Yun; Lu, Diana; Wilcox, Bruce A.; Hoang, Tung T.

    2009-01-01

    Genetic manipulation of the category B select agents Burkholderia pseudomallei and Burkholderia mallei has been stifled due to the lack of compliant selectable markers. Hence, there is a need for additional select-agent-compliant selectable markers. We engineered a selectable marker based on the gat gene (encoding glyphosate acetyltransferase), which confers resistance to the common herbicide glyphosate (GS). To show the ability of GS to inhibit bacterial growth, we determined the effective concentrations of GS against Escherichia coli and several Burkholderia species. Plasmids based on gat, flanked by unique flip recombination target (FRT) sequences, were constructed for allelic-replacement. Both allelic-replacement approaches, one using the counterselectable marker pheS and the gat-FRT cassette and one using the DNA incubation method with the gat-FRT cassette, were successfully utilized to create deletions in the asd and dapB genes of wild-type B. pseudomallei strains. The asd and dapB genes encode an aspartate-semialdehyde dehydrogenase (BPSS1704, chromosome 2) and dihydrodipicolinate reductase (BPSL2941, chromosome 1), respectively. Mutants unable to grow on media without diaminopimelate (DAP) and other amino acids of this pathway were PCR verified. These mutants displayed cellular morphologies consistent with the inability to cross-link peptidoglycan in the absence of DAP. The B. pseudomallei 1026b Δasd::gat-FRT mutant was complemented with the B. pseudomallei asd gene on a site-specific transposon, mini-Tn7-bar, by selecting for the bar gene (encoding bialaphos/PPT resistance) with PPT. We conclude that the gat gene is one of very few appropriate, effective, and beneficial compliant markers available for Burkholderia select-agent species. Together with the bar gene, the gat cassette will facilitate various genetic manipulations of Burkholderia select-agent species. PMID:19648360

  14. Genetic relationships among Heliconia (Heliconiaceae) species based on RAPD markers.

    PubMed

    Marouelli, L P; Inglis, P W; Ferreira, M A; Buso, G S C

    2010-07-13

    The family Heliconiaceae contains a single genus, Heliconia, with approximately 180 species of Neotropical origin. This genus was formerly allocated to the family Musaceae, but today forms its own family, in the order Zingiberales. The combination of inverted flowers, a single staminode and drupe fruits is an exclusive characteristic of Heliconia. Heliconias are cultivated as ornamental garden plants, and are of increasing importance as cut flowers. However, there are taxonomic confusions and uncertainties about the number of species and the relationships among them. Molecular studies are therefore necessary for better understanding of the species boundaries of these plants. We examined the genetic variability and the phylogenetic relationships of 124 accessions of the genus Heliconia based on RAPD markers. Phenetic and cladistic analyses, using 231 polymorphic RAPD markers, demonstrated that the genus Heliconia is monophyletic. Groupings corresponding to currently recognized species and some subgenera were found, and cultivars and hybrids were found to cluster with their parents. RAPD analysis generally agreed with morphological species classification, except for the position of the subgenus Stenochlamys, which was found to be polyphyletic.

  15. Citrus (Rutaceae) SNP markers based on Competitive Allele-Specific PCR; transferability across the Aurantioideae subfamily1

    PubMed Central

    Garcia-Lor, Andres; Ancillo, Gema; Navarro, Luis; Ollitrault, Patrick

    2013-01-01

    • Premise of the study: Single nucleotide polymorphism (SNP) markers based on Competitive Allele-Specific PCR (KASPar) were developed from sequences of three Citrus species. Their transferability was tested in 63 Citrus genotypes and 19 relative genera of the subfamily Aurantioideae to estimate the potential of SNP markers, selected from a limited intrageneric discovery panel, for ongoing broader diversity analysis at the intra- and intergeneric levels and systematic germplasm bank characterization. • Methods and Results: Forty-two SNP markers were developed using KASPar technology. Forty-one were successfully genotyped in all of the Citrus germplasm, where intra- and interspecific polymorphisms were observed. The transferability and diversity decreased with increasing taxonomic distance. • Conclusions: SNP markers based on the KASPar method developed from sequence data of a limited intrageneric discovery panel provide a valuable molecular resource for genetic diversity analysis of germplasm within a genus and should be useful for germplasm fingerprinting at a much broader diversity level. PMID:25202535

  16. Estimates of epistatic and pleiotropic effects of casein alpha s1 (CSN1S1) and thyroglobulin (TG) genetic markers on beef heifer performance traits enhanced by selection

    USDA-ARS?s Scientific Manuscript database

    Genetic marker effects and type of inheritance are estimated with poor precision when minor marker allele frequencies are low. A stable composite population (MARC II) was subjected to marker assisted selection for two years to equalize CSN1S1 and TG genetic marker frequencies to evaluate the epista...

  17. Marker-assisted selection for recognizing wheat mutant genotypes carrying HMW glutenin alleles related to baking quality.

    PubMed

    Zamani, Mohammad Javad; Bihamta, Mohammad Reza; Naserian Khiabani, Behnam; Tahernezhad, Zahra; Hallajian, Mohammad Taher; Shamsi, Marzieh Varasteh

    2014-01-01

    Allelic diversity of HMW glutenin loci in several studies revealed that allelic combinations affect dough quality. Dx5 + Dy10 subunits are related to good baking quality and Dx2 + Dy12 are related to undesirable baking quality. One of the most regular methods to evaluate the baking quality is SDS-PAGE which is used to improve baking quality labs. Marker-assisted selection is the method which can recognize the alleles related to baking quality and this method is based on polymerase chain reaction. 10 pairs of specific primers related to Dx2, Dx2.1, Dx5, Dy10, and Dy12 subunits were used for recognizing baking quality of some wheat varieties and some mutant genotypes. Only 5 pairs of them could show the specific bands. All subunits were recognized by the primers except Dx2.1. Some of the primers were extracted from previous studies and the others were designed based on D genome subunits of wheat. SDS-PAGE method accomplished having confidence in these marker's results. To realize the effect of mutation, seed storage proteins were measured. It showed that mutation had effect on the amount of seed storage protein on the mutant seeds (which showed polymorphism).

  18. A New Selectable Marker System for Genetic Studies of Bacteria: Final Report

    SciTech Connect

    Parsons, D; Tolmasky, M; Chain, P; Segelke, B W

    2011-03-18

    Genetic manipulations in bacteria currently rely on the introduction of antibiotic resistance genes into a bacterial strain; for those organisms that will be used for commercial or industrial applications, the genetic cassette encoding the antibiotic resistance is sometimes removed after selection. it is clear that if alternative technologies could obviate the need to introduce antibiotic resistance into bacteria, they would most certainly become a standard tool in molecular micriobiology for commercial, industrial as well as research applications. Here, they present the development of a novel genetic engineering technology based on toxin-antitoxin systems to modify bacterial genomes without the use of antibiotic resistance in the mutagenesis process. The primary goal is to develop antibiotic-free selection for genetically altered select agent pathogens. They are adapting the toxinc-antitoxin system to enable gene replacement in select agent pathogens since the NIH restrictions introducing antibiotic resistance into select agent pathogens have hindered research with select agent pathogens.

  19. Advances towards a Marker-Assisted Selection Breeding Program in Prairie Cordgrass, a Biomass Crop

    PubMed Central

    Gedye, K. R.; Gonzalez-Hernandez, J. L.; Owens, V.; Boe, A.

    2012-01-01

    Prairie cordgrass (Spartina pectinata Bosc ex Link) is an indigenous, perennial grass of North America that is being developed into a cellulosic biomass crop suitable for biofuel production. Limited research has been performed into the breeding of prairie cordgrass; this research details an initial investigation into the development of a breeding program for this species. Genomic libraries enriched for four simple sequence repeat (SSR) motifs were developed, 25 clones from each library were sequenced, identifying 70 SSR regions, and primers were developed for these regions, 35 of which were amplified under standard PCR conditions. These SSR markers were used to validate the crossing methodology of prairie cordgrass and it was found that crosses between two plants occurred without the need for emasculation. The successful cross between two clones of prairie cordgrass indicates that this species is not self-incompatible. The results from this research will be used to instigate the production of a molecular map of prairie cordgrass which can be used to incorporate marker-assisted selection (MAS) protocols into a breeding program to improve this species for cellulosic biomass production. PMID:23227036

  20. Changes of B and T lymphocytes and selected apopotosis markers in Hashimoto's thyroiditis.

    PubMed

    Kaczmarek, Elzbieta; Lacka, Katarzyna; Jarmolowska-Jurczyszyn, Donata; Sidor, Anna; Majewski, Przemyslaw

    2011-07-01

    The aim was to assess changes of B and T lymphocytes and selected apoptotic markers in Hashimoto thyroiditis (HT) cases on the basis of quantitative immunohistochemical studies (CD20, CD43, CD8, Bcl-2, caspase-3). The control group comprised colloid goitres without inflammatory infiltrate taken from 10 female patients. Thyroid specimens were obtained retrospectively from 40 patients. The immunohistochemical reactions were subject to quantitative evaluation performed using image-processing methods, including a spatial visualisation of the markers' expression. The percentage of Bcl-2 reactions in HT (mean 3.65%, SD 2.94%) was significantly lower than in the control group (mean 13.99%, SD 5.04%), while the thyroid follicles in HT samples exhibited a higher degree of staining for caspase-3 (mean 1.10%, SD 1.03%) in contrast to normal control tissues (mean 0.48%, SD 1.02%). The results from this study indicate that apoptosis plays a major role in the patogenesis of autoimmune thyroid diseases containing the main pathogenic events in the lesion of thyroid follicular cells in HT. Moreover, the reactivity of CD43 and CD20 was significantly higher in Hashimoto disease, while CD8 was not significantly different from the control group.

  1. Targeted insertion of foreign genes into the tobacco plastid genome without physical linkage to the selectable marker gene

    SciTech Connect

    Carrer, H.; Maliga, P.

    1995-08-01

    To determine whether targeted DNA insertion into the tobacco plastid genome can be obtained without physical linkage to a selectable marker gene, we carried out biolistic transformation of chloroplasts in tobacco leaf segments with a 1:1 mix of two independently targeted antibiotic resistance genes. Plastid transformants were selected by spectinomycin resistance due to expression of an integrated aadA gene. Integration of the unselected kanamycin resistance (kan) gene into the same plastid genome was established by Southern probing in {approx}20% of the spectinomycin-selected clones. Efficient cotransformation will facilitate targeted plastid genome modification without physical linkage to a marker gene. 26 refs., 5 figs., 1 tab.

  2. A bacterial gene codA encoding cytosine deaminase is an effective conditional negative selectable marker in Glycine max

    USDA-ARS?s Scientific Manuscript database

    Background Conditional negative selection is a powerful technique whereby the absence of a gene product allows survival in otherwise lethal conditions. In plants, the Escherichia coli gene codA has been employed as a negative selection marker. CodA is a conditionally lethal dominant gene encoding cy...

  3. Response to Phenotypic and Marker-Assisted Selection for Yield and Quality Component Traits in Cucumber (Cucumis Sativus L.)

    USDA-ARS?s Scientific Manuscript database

    Even though the potential benefits of marker-assisted selection (MAS) for line and population development to improve yield in cucumber have been demonstrated, its application during tandem selection for yield and quality components has not been investigated. Therefore, two cucumber recombinant inbr...

  4. Effect of selected spices on chemical and sensory markers in fortified rye-buckwheat cakes.

    PubMed

    Przygodzka, Małgorzata; Zieliński, Henryk; Ciesarová, Zuzana; Kukurová, Kristina; Lamparski, Grzegorz

    2016-07-01

    The aim of this study was to find out the effect of selected spices on chemical and sensorial markers in cakes formulated on rye and light buckwheat flour fortified with spices. Among collection of spices, rye-buckwheat cakes fortified individually with cloves, nutmeg, allspice, cinnamon, vanilla, and spice mix revealed the highest sensory characteristics and overall quality. Cakes fortified with cloves, allspice, and spice mix showed the highest antioxidant capacity, total phenolics, rutin, and almost threefold higher available lysine contents. The reduced furosine content as well as free and total fluorescent intermediatory compounds were observed as compared to nonfortified cakes. The FAST index was significantly lowered in all cakes enriched with spices, especially with cloves, allspice, and mix. In contrast, browning index increased in compare to cakes without spices. It can be suggested that clove, allspice, vanilla, and spice mix should be used for production of safety and good quality cakes.

  5. Development of PCR-Based Markers Linked to Quantitative Resistance to Late Blight in a Diploid Hybrid Potato Population of Solanum phureja × S. stenotomum

    USDA-ARS?s Scientific Manuscript database

    A diploid hybrid Solanum phureja × S. stenotomum population which has shown relatively high levels of quantitative resistance to late blight was used to develop PCR–based markers for marker assisted selection (MAS) of disease resistance in potato breeding. Two random amplified polymorphic DNA (RAPD...

  6. Increased prediction accuracy in wheat breeding trials using a marker × environment interaction genomic selection model.

    PubMed

    Lopez-Cruz, Marco; Crossa, Jose; Bonnett, David; Dreisigacker, Susanne; Poland, Jesse; Jannink, Jean-Luc; Singh, Ravi P; Autrique, Enrique; de los Campos, Gustavo

    2015-02-06

    Genomic selection (GS) models use genome-wide genetic information to predict genetic values of candidates of selection. Originally, these models were developed without considering genotype × environment interaction(G×E). Several authors have proposed extensions of the single-environment GS model that accommodate G×E using either covariance functions or environmental covariates. In this study, we model G×E using a marker × environment interaction (M×E) GS model; the approach is conceptually simple and can be implemented with existing GS software. We discuss how the model can be implemented by using an explicit regression of phenotypes on markers or using co-variance structures (a genomic best linear unbiased prediction-type model). We used the M×E model to analyze three CIMMYT wheat data sets (W1, W2, and W3), where more than 1000 lines were genotyped using genotyping-by-sequencing and evaluated at CIMMYT's research station in Ciudad Obregon, Mexico, under simulated environmental conditions that covered different irrigation levels, sowing dates and planting systems. We compared the M×E model with a stratified (i.e., within-environment) analysis and with a standard (across-environment) GS model that assumes that effects are constant across environments (i.e., ignoring G×E). The prediction accuracy of the M×E model was substantially greater of that of an across-environment analysis that ignores G×E. Depending on the prediction problem, the M×E model had either similar or greater levels of prediction accuracy than the stratified analyses. The M×E model decomposes marker effects and genomic values into components that are stable across environments (main effects) and others that are environment-specific (interactions). Therefore, in principle, the interaction model could shed light over which variants have effects that are stable across environments and which ones are responsible for G×E. The data set and the scripts required to reproduce the analysis are

  7. Quantitative trait locus mapping in chickens by selective DNA pooling with dinucleotide microsatellite markers by using purified DNA and fresh or frozen red blood cells as applied to marker-assisted selection.

    PubMed

    Lipkin, E; Fulton, J; Cheng, H; Yonash, N; Soller, M

    2002-03-01

    Many large, half-sib sire families are an integral component of chicken genetic improvement programs. These family structures include a sufficient number of individuals for mapping quantitative trait loci (QTL) at high statistical power. However, realizing this statistical power through individual or selective genotyping is yet too costly to be feasible under current genotyping methodologies. Genotyping costs can be greatly reduced through selective DNA pooling, involving densitometric estimates of marker allele frequencies in pooled DNA samples. When using dinucleotide microsatellite markers, however, such estimates are often confounded by overlapping "shadow" bands and can be confounded further by differential amplification of alleles. In the present study a shadow correction procedure provided accurate densitometric estimates of allele frequency for dinucleotide microsatellite markers in pools made from chicken purified DNA samples, fresh blood samples, and frozen-thawed blood samples. In a retrospective study, selective DNA pooling with thawed blood samples successfully identified two QTL previously shown by selective genotyping to affect resistance in chickens to Marek's disease. It is proposed that use of selective DNA pooling can provide relatively low-cost mapping and use in marker-assisted selection of QTL that affect production traits in chickens.

  8. Selection and use of SNP markers for animal identification and paternity analysis in U.S. beef cattle.

    PubMed

    Heaton, Michael P; Harhay, Gregory P; Bennett, Gary L; Stone, Roger T; Grosse, W Michael; Casas, Eduardo; Keele, John W; Smith, Timothy P L; Chitko-McKown, Carol G; Laegreid, William W

    2002-05-01

    DNA marker technology represents a promising means for determining the genetic identity and kinship of an animal. Compared with other types of DNA markers, single nucleotide polymorphisms (SNPs) are attractive because they are abundant, genetically stable, and amenable to high-throughput automated analysis. In cattle, the challenge has been to identify a minimal set of SNPs with sufficient power for use in a variety of popular breeds and crossbred populations. This report describes a set of 32 highly informative SNP markers distributed among 18 autosomes and both sex chromosomes. Informativity of these SNPs in U.S. beef cattle populations was estimated from the distribution of allele and genotype frequencies in two panels: one consisting of 96 purebred sires representing 17 popular breeds, and another with 154 purebred American Angus from six herds in four Midwestern states. Based on frequency data from these panels, the estimated probability that two randomly selected, unrelated individuals will possess identical genotypes for all 32 loci was 2.0 x 10(-13) for multi-breed composite populations and 1.9 x 10(-10) for purebred Angus populations. The probability that a randomly chosen candidate sire will be excluded from paternity was estimated to be 99.9% and 99.4% for the same respective populations. The DNA immediately surrounding the 32 target SNPs was sequenced in the 96 sires of the multi-breed panel and found to contain an additional 183 polymorphic sites. Knowledge of these additional sites, together with the 32 target SNPs, allows the design of robust, accurate genotype assays on a variety of high-throughput SNP genotyping platforms.

  9. Performance and grain yield stability of maize populations developed using marker-assisted recurrent selection and pedigree selection procedures.

    PubMed

    Beyene, Yoseph; Semagn, Kassa; Mugo, Stephen; Prasanna, Boddupalli M; Tarekegne, Amsal; Gakunga, John; Sehabiague, Pierre; Meisel, Barbara; Oikeh, Sylvester O; Olsen, Michael; Crossa, Jose

    A marker-assisted recurrent selection (MARS) program was undertaken in sub-Saharan Africa to improve grain yield under drought-stress in 10 biparental tropical maize populations. The objectives of the present study were to evaluate the performance of C1S2-derived hybrids obtained after three MARS cycles (one cycle of recombination (C1), followed by two generations of selfing (S2), and to study yield stability under both drought-stress (DS) and well-watered (WW) conditions. For each of the 10 populations, we evaluated hybrids developed by crossing 47-74 C1S2 lines advanced through MARS, the best five S5 lines developed through pedigree selection, and the founder parents with a single-cross tester from a complementary heterotic group. The hybrids and five commercial checks were evaluated in Kenya under 1-3 DS and 3-5 WW conditions with two replications. Combined across DS locations, the top 10 C1S2-derived hybrids from each of the 10 biparental populations produced 0.5-46.3 and 11.1-55.1 % higher mean grain yields than hybrids developed using pedigree selection and the commercial checks, respectively. Across WW locations, the best 10 hybrids derived from C1S2 of each population produced 3.4-13.3 and 7.9-36.5 % higher grain yields than hybrids derived using conventional pedigree breeding and the commercial checks, respectively. Mean days to anthesis of the best 10 C1S2 hybrids were comparable to those of hybrids developed using the pedigree method, the founder parents and the commercial checks, with a maximum difference of 3.5 days among the different groups. However, plant height was significantly (P < 0.01) different in most pairwise comparisons. Our results showed the superiority of MARS over pedigree selection for improving diverse tropical maize populations as sources of improved lines for stress-prone environments and thus MARS can be effectively integrated into mainstream maize breeding programs.

  10. Saccharomyces cerevisiae single-copy plasmids for auxotrophy compensation, multiple marker selection, and for designing metabolically cooperating communities

    PubMed Central

    Matsarskaia, Olga; Eckerstorfer, Florian; Ralser, Markus

    2016-01-01

    Auxotrophic markers are useful tools in cloning and genome editing, enable a large spectrum of genetic techniques, as well as facilitate the study of metabolite exchange interactions in microbial communities. If unused background auxotrophies are left uncomplemented however, yeast cells need to be grown in nutrient supplemented or rich growth media compositions, which precludes the analysis of biosynthetic metabolism, and which leads to a profound impact on physiology and gene expression. Here we present a series of 23 centromeric plasmids designed to restore prototrophy in typical Saccharomyces cerevisiae laboratory strains. The 23 single-copy plasmids complement for deficiencies in HIS3, LEU2, URA3, MET17 or LYS2 genes and in their combinations, to match the auxotrophic background of the popular functional-genomic yeast libraries that are based on the S288c strain. The plasmids are further suitable for designing self-establishing metabolically cooperating (SeMeCo) communities, and possess a uniform multiple cloning site to exploit multiple parallel selection markers in protein expression experiments. PMID:27830062

  11. Development of swine-specific DNA markers for biosensor-based halal authentication.

    PubMed

    Ali, M E; Hashim, U; Kashif, M; Mustafa, S; Che Man, Y B; Abd Hamid, S B

    2012-06-29

    The pig (Sus scrofa) mitochondrial genome was targeted to design short (15-30 nucleotides) DNA markers that would be suitable for biosensor-based hybridization detection of target DNA. Short DNA markers are reported to survive harsh conditions in which longer ones are degraded into smaller fragments. The whole swine mitochondrial-genome was in silico digested with AluI restriction enzyme. Among 66 AluI fragments, five were selected as potential markers because of their convenient lengths, high degree of interspecies polymorphism and intraspecies conservatism. These were confirmed by NCBI blast analysis and ClustalW alignment analysis with 11 different meat-providing animal and fish species. Finally, we integrated a tetramethyl rhodamine-labeled 18-nucleotide AluI fragment into a 3-nm diameter citrate-tannate coated gold nanoparticle to develop a swine-specific hybrid nanobioprobe for the determination of pork adulteration in 2.5-h autoclaved pork-beef binary mixtures. This hybrid probe detected as low as 1% pork in deliberately contaminated autoclaved pork-beef binary mixtures and no cross-species detection was recorded, demonstrating the feasibility of this type of probe for biosensor-based detection of pork adulteration of halal and kosher foods.

  12. Development and validation of functional CAPS markers for the FAE genes in Brassica juncea and their use in marker-assisted selection

    PubMed Central

    Saini, Navinder; Singh, Naveen; Kumar, Anil; Vihan, Nitika; Yadav, Sangita; Vasudev, Sujata; Yadava, D.K.

    2016-01-01

    Low erucic acid is a major breeding target to improve the edible oil quality in Brassica juncea. The single nucleotide polymorphism (SNP) in fatty acid elongase 1 (FAE1.1 and FAE1.2) gene was exploited to expedite the breeding program. The paralogs of FAE1 gene were sequenced from low erucic acid genotype Pusa Mustard 30 and SNPs were identified through homologous alignment with sequence downloaded from NCBI GenBank. Two SNPs in FAE1.1 at position 591 and 1265 and one in FAE1.2 at 237 were found polymorphic among low and high erucic acid genotypes. These SNPs either create or change the recognition site of restriction enzymes. Transition of a single nucleotide at position 591 and 1265 in FAE1.1, and at position 237 in FAE1.2, leads to a change in the recognition site of Hpy99I, BglII and MnlI restriction enzymes, respectively. Two CAPS markers for FAE1.1 and one for FAE1.2 were developed to differentiate low and high erucic acid genotypes. The efficiency of these CAPS markers was found 100 per cent when validated in Brassica juncea, and B. nigra genotypes and used in back-cross breeding. These CAPS markers will facilitate in marker-assisted selection for improvement of oil quality in Brassica juncea. PMID:28163599

  13. Augmenting the Genetic Toolbox for Sulfolobus islandicus with a Stringent Positive Selectable Marker for Agmatine Prototrophy

    PubMed Central

    Cooper, Tara E.; Krause, David J.

    2013-01-01

    Sulfolobus species have become the model organisms for studying the unique biology of the crenarchaeal division of the archaeal domain. In particular, Sulfolobus islandicus provides a powerful opportunity to explore natural variation via experimental functional genomics. To support these efforts, we further expanded genetic tools for S. islandicus by developing a stringent positive selection for agmatine prototrophs in strains in which the argD gene, encoding arginine decarboxylase, has been deleted. Strains with deletions in argD were shown to be auxotrophic for agmatine even in nutrient-rich medium, but growth could be restored by either supplementation of exogenous agmatine or reintroduction of a functional copy of the argD gene from S. solfataricus P2 into the ΔargD host. Using this stringent selection, a robust targeted gene knockout system was established via an improved next generation of the MID (marker insertion and unmarked target gene deletion) method. Application of this novel system was validated by targeted knockout of the upsEF genes involved in UV-inducible cell aggregation formation. PMID:23835176

  14. EST-based Microsatellite Marker Data Mining and Characterizing

    USDA-ARS?s Scientific Manuscript database

    Peanut (Arachis hypogaea L.) is an important crop for oil production. In the recent years, molecular marker technologies have been widely applied to genetic diversity analysis, genetic mapping, molecular marker-assisted breeding, gene tagging and QTLs analysis. However, it is expensive, labor-intens...

  15. Intelligent DNA-based molecular diagnostics using linked genetic markers

    SciTech Connect

    Pathak, D.K.; Perlin, M.W.; Hoffman, E.P.

    1994-12-31

    This paper describes a knowledge-based system for molecular diagnostics, and its application to fully automated diagnosis of X-linked genetic disorders. Molecular diagnostic information is used in clinical practice for determining genetic risks, such as carrier determination and prenatal diagnosis. Initially, blood samples are obtained from related individuals, and PCR amplification is performed. Linkage-based molecular diagnosis then entails three data analysis steps. First, for every individual, the alleles (i.e., DNA composition) are determined at specified chromosomal locations. Second, the flow of genetic material among the individuals is established. Third, the probability that a given individual is either a carrier of the disease or affected by the disease is determined. The current practice is to perform each of these three steps manually, which is costly, time consuming, labor-intensive, and error-prone. As such, the knowledge-intensive data analysis and interpretation supersede the actual experimentation effort as the major bottleneck in molecular diagnostics. By examining the human problem solving for the task, we have designed and implemented a prototype knowledge-based system capable of fully automating linkage-based molecular diagnostics in X-linked genetic disorders, including Duchenne Muscular Dystrophy (DMD). Our system uses knowledge-based interpretation of gel electrophoresis images to determine individual DNA marker labels, a constraint satisfaction search for consistent genetic flow among individuals, and a blackboard-style problem solver for risk assessment. We describe the system`s successful diagnosis of DMD carrier and affected individuals from raw clinical data.

  16. Comparison of protocols for walking and running kinematics based on skin surface markers and rigid clusters of markers.

    PubMed

    Miana, A N; Prudêncio, M V; Barros, R M L

    2009-11-01

    The purpose of this study was to compare the two main types of marker sets for human body representation based on rigid clusters of markers and skin surface markers for measuring kinematics during walking and running. Velocity, body segment, and joint angle were considered in the comparison of both protocols. Six male athletes were studied during treadmill gait at 1.4 and 5.5 m/s and recorded with 8 high speed video cameras. The subjects used simultaneously both protocols in the same walking and running cycles, in order to compare the variability in the determination of the joint centers' positions and the joint angles calculated from each protocol. The three-way ANOVA results showed that the variability of the inter-markers distance in the skin surface protocol was higher than that in the rigid clusters of markers, as reported in the literature. However, no statistical differences between the protocols were found in the variability of the determination of the joint centers' positions. Therefore no advantage was verified to rigid cluster protocols even for the upper body segments. Another conclusion is that increases in velocity produced increases in variability of the joint centers' distances and increases in the maximum differences between the joint angles.

  17. Phenotypic versus marker-assisted selection for stalk strength and second-generation European corn borer resistance in maize.

    PubMed

    Flint-Garcia, S A; Darrah, L L; McMullen, M D; Hibbard, B E

    2003-11-01

    Maize ( Zea mays L.) stalk lodging is breakage of the stalk at or below the ear, which may result in loss of the ear at harvest. Stalk lodging is often intensified by the stalk tunneling action of the second-generation of the European corn borer (2-ECB) [Ostrinia nubilalis (Hübner)]. Rind penetrometer resistance (RPR) has been used to measure stalk strength and improve stalk lodging resistance, and quantitative trait loci (QTL) have been identified for both RPR and 2-ECB damage. Phenotypic recurrent selection (PS) increases the frequency of favorable alleles over cycles of selection. Several studies have indicated that marker-assisted selection (MAS) is also a potentially valuable selection tool. The objective of this study was to compare the efficiency of PS versus MAS for RPR and 2-ECB. Marker-assisted selection for high and low RPR was effective in the three populations studied. Phenotypic selection for both high and low RPR was more effective than MAS in two of the populations. However, in a third population, MAS for high RPR using QTL effects from the same population was more effective than PS, and using QTL effects from a separate population was just as effective as PS. Marker-assisted selection for resistance and susceptibility to 2-ECB using QTL effects from the same population was effective in increasing susceptibility, but not in increasing resistance. Marker-assisted selection using QTL effects from a separate population was effective in both directions of selection. Thus, MAS was effective in selecting for both resistance and susceptibility to 2-ECB. These results demonstrated that MAS can be an effective selection tool for both RPR and 2-ECB resistance. These results also validate the locations and effects of QTL for RPR and 2-ECB resistance identified in earlier studies.

  18. Multivariate analysis for the estimation of target localization errors in fiducial marker-based radiotherapy.

    PubMed

    Takamiya, Masanori; Nakamura, Mitsuhiro; Akimoto, Mami; Ueki, Nami; Yamada, Masahiro; Tanabe, Hiroaki; Matsuo, Yukinori; Mizowaki, Takashi; Kokubo, Masaki; Hiraoka, Masahiro; Itoh, Akio

    2016-04-01

    To assess the target localization error (TLE) in terms of the distance between the target and the localization point estimated from the surrogates (|TMD|), the average of respiratory motion for the surrogates and the target (|aRM|), and the number of fiducial markers used for estimating the target (n). This study enrolled 17 lung cancer patients who subsequently underwent four fractions of real-time tumor tracking irradiation. Four or five fiducial markers were implanted around the lung tumor. The three-dimensional (3D) distance between the tumor and markers was at maximum 58.7 mm. One of the markers was used as the target (Pt), and those markers with a 3D |TMDn| ≤ 58.7 mm at end-exhalation were then selected. The estimated target position (Pe) was calculated from a localization point consisting of one to three markers except Pt. Respiratory motion for Pt and Pe was defined as the root mean square of each displacement, and |aRM| was calculated from the mean value. TLE was defined as the root mean square of each difference between Pt and Pe during the monitoring of each fraction. These procedures were performed repeatedly using the remaining markers. To provide the best guidance on the answer with n and |TMD|, fiducial markers with a 3D |aRM ≥ 10 mm were selected. Finally, a total of 205, 282, and 76 TLEs that fulfilled the 3D |TMD| and 3D |aRM| criteria were obtained for n = 1, 2, and 3, respectively. Multiple regression analysis (MRA) was used to evaluate TLE as a function of |TMD| and |aRM| in each n. |TMD| for n = 1 was larger than that for n = 3. Moreover, |aRM| was almost constant for all n, indicating a similar scale for the marker's motion near the lung tumor. MRA showed that |aRM| in the left-right direction was the major cause of TLE; however, the contribution made little difference to the 3D TLE because of the small amount of motion in the left-right direction. The TLE calculated from the MRA ((MRA)TLE) increased as |TMD| and |aRM| increased and

  19. Semen characteristics and selected biochemical markers of canine seminal plasma in various seasons of the year.

    PubMed

    Strzeżek, R; Szemplińska, K; Filipowicz, K; Kordan, W

    2015-01-01

    The aim of this study was to evaluate the influence of season on selected qualitative semen characteristics and biochemical markers of canine seminal plasma. Whole ejaculates were collected from 5 crossbred dogs aged 2-8 years. The study covered a period of one year divided into four seasons: spring (March, April, May), summer (June, July, August), autumn (September, October, November) and winter (December, January, February). Semen samples were subjected to macroscopic and microscopic analyses to determine semen volume, total sperm counts and sperm morphology parameters. The study also involved the determination of sperm motility parameters (CASA system), sperm plasma membrane integrity (SPMI, fluorescent staining SYBR-14/PI), sperm mitochondrial membrane potential (MMP, fluorescent staining JC-1/PI) and the ATP content of sperm cells. Total protein content (TPC) and the activity of alkaline phosphatase (AP) and acid phosphatase (AcP) were determined in biochemical analyses of seminal plasma. No significant differences in ejaculate volume, SMPI or ATP content of sperm cells were observed between seasons. The highest total sperm counts were reported in ejaculates acquired in summer and autumn. The lowest MMP values were determined in summer ejaculates. No significant differences in sperm motility (MOT) were observed throughout the experiment, but ejaculates collected in autumn and winter were characterized by the highest progressive motility (PMOT). AP activity and TPC were not significantly affected by season. However, AcP activity levels were significantly lower in autumn than in the remaining seasons. Seasonal variations in the analyzed macroscopic and microscopic parameters of ejaculates and biochemical markers of seminal plasma did not exert a clear negative effect on the quality of canine semen.

  20. Bacterial inosine 5'-monophosphate dehydrogenase ("IMPDH") DNA as a dominant selectable marker in mammals and other eukaryotes

    DOEpatents

    Huberman, Eliezer; Baccam, Mekhine J.

    2007-02-27

    The present invention relates to a nucleic acid sequence and its corresponding protein sequence useful as a dominant selectable marker in eukaryotes. More specifically the invention relates to a nucleic acid encoding a bacterial IMPDH gene that has been engineered into a eukaryotic expression vectors, thereby permitting bacterial IMPDH expression in mammalian cells. Bacterial IMPDH expression confers resistance to MPA which can be used as dominant selectable marker in eukaryotes including mammals. The invention also relates to expression vectors and cells that express the bacterial IMPDH gene as well as gene therapies and protein synthesis.

  1. Fluorescence/luminescence-based markers for the assessment of Schistosoma mansoni schistosomula drug assays.

    PubMed

    Panic, Gordana; Flores, Dayana; Ingram-Sieber, Katrin; Keiser, Jennifer

    2015-12-08

    Schistosomiasis is responsible for a tremendous public health burden, yet only a single drug, praziquantel, is available. New antischistosomal treatments should therefore be developed. The accuracy, speed and objectivity of in vitro drug screening depend on the assay read-out. Microscopy is still the current gold standard and is in need of updating to an automated format. The aim of the present study was to investigate a panel of fluorescence/luminescence dyes for their applicability as viability markers in drug sensitivity assays for Schistosoma mansoni schistosomula. A search for available viability and cytotoxicity marker assays and dyes was carried out and a short-list of the most interesting candidates was created. The selected kits and dyes were tested on S. mansoni Newly Transformed Schistosomula (NTS), first to assess whether they correlate with parasite viability, with comparatively low background noise, and to optimise assay conditions. Markers fulfilling these criteria were then tested in a dose-response drug assay using standard and experimental drugs and those for which an IC50 value could be accurately and reproducibly calculated were also tested on a subset of a compound library to determine their hit-identification accuracy. Of the 11 markers selected for testing, resazurin, Vybrant® and CellTiter-Glo® correlated best with NTS viability, produced signals ≥ 3-fold stronger than background noise and revealed a significant signal-to-NTS concentration relationship. Of these, CellTiter-Glo® could be used to accurately determine IC50 values for antischistosomals. Use of CellTiter-Glo® in a compound subset screen identified 100% of hits that were identified using standard microscopic evaluation. This study presents a comprehensive overview of the utility of colorimetric markers in drug screening. Our study demonstrates that it is difficult to develop a simple, cheap "just add" colorimetric marker-based drug assay for the larval stage of S

  2. Smart markers for watershed-based cell segmentation.

    PubMed

    Koyuncu, Can Fahrettin; Arslan, Salim; Durmaz, Irem; Cetin-Atalay, Rengul; Gunduz-Demir, Cigdem

    2012-01-01

    Automated cell imaging systems facilitate fast and reliable analysis of biological events at the cellular level. In these systems, the first step is usually cell segmentation that greatly affects the success of the subsequent system steps. On the other hand, similar to other image segmentation problems, cell segmentation is an ill-posed problem that typically necessitates the use of domain-specific knowledge to obtain successful segmentations even by human subjects. The approaches that can incorporate this knowledge into their segmentation algorithms have potential to greatly improve segmentation results. In this work, we propose a new approach for the effective segmentation of live cells from phase contrast microscopy. This approach introduces a new set of "smart markers" for a marker-controlled watershed algorithm, for which the identification of its markers is critical. The proposed approach relies on using domain-specific knowledge, in the form of visual characteristics of the cells, to define the markers. We evaluate our approach on a total of 1,954 cells. The experimental results demonstrate that this approach, which uses the proposed definition of smart markers, is quite effective in identifying better markers compared to its counterparts. This will, in turn, be effective in improving the segmentation performance of a marker-controlled watershed algorithm.

  3. Role of endocytosis in localization and maintenance of the spatial markers for bud-site selection in yeast.

    PubMed

    Tuo, Shanshan; Nakashima, Kenichi; Pringle, John R

    2013-01-01

    The yeast Saccharomyces cerevisiae normally selects bud sites (and hence axes of cell polarization) in one of two distinct patterns, the axial pattern of haploid cells and the bipolar pattern of diploid cells. These patterns depend on distinct sets of cortical-marker proteins that transmit positional information through a common signaling pathway based on a Ras-type GTPase. It has been reported previously that various proteins of the endocytic pathway may be involved in determining the bipolar pattern but not the axial pattern. To explore this question systematically, we constructed and analyzed congenic haploid and diploid deletion mutants for 14 genes encoding proteins that are involved in endocytosis. The mutants displayed a wide range of severities in their overall endocytosis defects, as judged by their growth rates and abilities to take up the lipophilic dye FM 4-64. Consistent with the previous reports, none of the mutants displayed a significant defect in axial budding, but they displayed defects in bipolar budding that were roughly correlated with the severities of their overall endocytosis defects. Both the details of the mutant budding patterns and direct examination of GFP-tagged marker proteins suggested that both initial formation and maintenance of the normally persistent bipolar marks depend on endocytosis, as well as polarized exocytosis, in actively growing cells. Interestingly, maintenance of the bipolar marks in non-growing cells did not appear to require normal levels of endocytosis. In some cases, there was a striking lack of correlation between the overall severities of the general-endocytosis defect and the bud-site selection defect, suggesting that various endocytosis proteins may differ in their importance for the uptake of various plasma-membrane targets.

  4. Selectable Markers for Use in Genetic Manipulation of Extensively Drug-Resistant (XDR) Acinetobacter baumannii HUMC1

    PubMed Central

    Ulhaq, Amber; Yan, Jun; Pantapalangkoor, Paul; Nielsen, Travis B.; Davies, Bryan W.; Actis, Luis A.; Spellberg, Brad

    2017-01-01

    ABSTRACT Acinetobacter baumannii is one of the most antibiotic-resistant pathogens in clinical medicine, and extensively drug-resistant (XDR) strains are commonly isolated from infected patients. Such XDR strains are already resistant to traditional selectable genetic markers, limiting the ability to conduct pathogenesis research by genetic disruption. Optimization of selectable markers is therefore critical for the advancement of fundamental molecular biology techniques to use in these strains. We screened 23 drugs that constitute a broad array of antibiotics spanning multiple drug classes against HUMC1, a highly virulent and XDR A. baumannii clinical blood and lung isolate. HUMC1 is resistant to all clinically useful antibiotics that are reported by the clinical microbiology laboratory, except for colistin. Ethical concerns about intentionally establishing pan-resistance, including to the last-line agent, colistin, in a clinical isolate made identification of other markers desirable. We screened additional antibiotics that are in clinical use and those that are useful only in a lab setting to identify selectable markers that were effective at selecting for transformants in vitro. We show that supraphysiological levels of tetracycline can overcome innate drug resistance displayed by this XDR strain. Last, we demonstrate that transformation of the tetA (tetracycline resistance) and Sh ble (zeocin resistance), but not pac (puromycin resistance), resistance cassettes allow for selection of drug-resistant transformants. These results make the genetic manipulation of XDR A. baumannii strains easily achieved. IMPORTANCE Multidrug-resistant (MDR), extensively drug-resistant (XDR), and pan-drug-resistant (PDR) strains of Acinetobacter baumannii have frequently been characterized. The ability of A. baumannii to develop resistance to antibiotics is a key reason this organism has been difficult to study using genetic and molecular biology approaches. Here we report

  5. Selectable Markers for Use in Genetic Manipulation of Extensively Drug-Resistant (XDR) Acinetobacter baumannii HUMC1.

    PubMed

    Luna, Brian M; Ulhaq, Amber; Yan, Jun; Pantapalangkoor, Paul; Nielsen, Travis B; Davies, Bryan W; Actis, Luis A; Spellberg, Brad

    2017-01-01

    Acinetobacter baumannii is one of the most antibiotic-resistant pathogens in clinical medicine, and extensively drug-resistant (XDR) strains are commonly isolated from infected patients. Such XDR strains are already resistant to traditional selectable genetic markers, limiting the ability to conduct pathogenesis research by genetic disruption. Optimization of selectable markers is therefore critical for the advancement of fundamental molecular biology techniques to use in these strains. We screened 23 drugs that constitute a broad array of antibiotics spanning multiple drug classes against HUMC1, a highly virulent and XDR A. baumannii clinical blood and lung isolate. HUMC1 is resistant to all clinically useful antibiotics that are reported by the clinical microbiology laboratory, except for colistin. Ethical concerns about intentionally establishing pan-resistance, including to the last-line agent, colistin, in a clinical isolate made identification of other markers desirable. We screened additional antibiotics that are in clinical use and those that are useful only in a lab setting to identify selectable markers that were effective at selecting for transformants in vitro. We show that supraphysiological levels of tetracycline can overcome innate drug resistance displayed by this XDR strain. Last, we demonstrate that transformation of the tetA (tetracycline resistance) and Sh ble (zeocin resistance), but not pac (puromycin resistance), resistance cassettes allow for selection of drug-resistant transformants. These results make the genetic manipulation of XDR A. baumannii strains easily achieved. IMPORTANCE Multidrug-resistant (MDR), extensively drug-resistant (XDR), and pan-drug-resistant (PDR) strains of Acinetobacter baumannii have frequently been characterized. The ability of A. baumannii to develop resistance to antibiotics is a key reason this organism has been difficult to study using genetic and molecular biology approaches. Here we report selectable

  6. Multilocus phylogeography and phylogenetics using sequence-based markers.

    PubMed

    Brito, Patrícia H; Edwards, Scott V

    2009-04-01

    We review recent trends in phylogeography and phylogenetics and argue that these two fields stand to be reunited by the common yardstick provided by sequence and SNP data and by new multilocus methods for phylogenetic analysis. Whereas the modern incarnation of both fields was spawned by PCR approaches applied to mitochondrial DNA in the late 1980s, the two fields diverged during the 1990s largely due to the adoption by phylogeographers of microsatellites, in contrast to the adoption of nuclear sequence data by phylogeneticists. Sequence-based markers possess a number of advantages over microsatellites, even on the recent time scales that are the purview of phylogeography. Using examples primarily from vertebrates, we trace the maturation of nuclear gene phylogeography and phylogenetics and suggest that the abundant instances of gene tree heterogeneity beckon a new generation of phylogenetic methods that focus on estimating species trees as distinct from gene trees. Whole genomes provide a powerful common yardstick on which both phylogeography and phylogenetics can assume their proper place as ends of a continuum.

  7. Kazusa Marker DataBase: a database for genomics, genetics, and molecular breeding in plants

    PubMed Central

    Shirasawa, Kenta; Isobe, Sachiko; Tabata, Satoshi; Hirakawa, Hideki

    2014-01-01

    In order to provide useful genomic information for agronomical plants, we have established a database, the Kazusa Marker DataBase (http://marker.kazusa.or.jp). This database includes information on DNA markers, e.g., SSR and SNP markers, genetic linkage maps, and physical maps, that were developed at the Kazusa DNA Research Institute. Keyword searches for the markers, sequence data used for marker development, and experimental conditions are also available through this database. Currently, 10 plant species have been targeted: tomato (Solanum lycopersicum), pepper (Capsicum annuum), strawberry (Fragaria × ananassa), radish (Raphanus sativus), Lotus japonicus, soybean (Glycine max), peanut (Arachis hypogaea), red clover (Trifolium pratense), white clover (Trifolium repens), and eucalyptus (Eucalyptus camaldulensis). In addition, the number of plant species registered in this database will be increased as our research progresses. The Kazusa Marker DataBase will be a useful tool for both basic and applied sciences, such as genomics, genetics, and molecular breeding in crops. PMID:25320561

  8. Kazusa Marker DataBase: a database for genomics, genetics, and molecular breeding in plants.

    PubMed

    Shirasawa, Kenta; Isobe, Sachiko; Tabata, Satoshi; Hirakawa, Hideki

    2014-09-01

    In order to provide useful genomic information for agronomical plants, we have established a database, the Kazusa Marker DataBase (http://marker.kazusa.or.jp). This database includes information on DNA markers, e.g., SSR and SNP markers, genetic linkage maps, and physical maps, that were developed at the Kazusa DNA Research Institute. Keyword searches for the markers, sequence data used for marker development, and experimental conditions are also available through this database. Currently, 10 plant species have been targeted: tomato (Solanum lycopersicum), pepper (Capsicum annuum), strawberry (Fragaria × ananassa), radish (Raphanus sativus), Lotus japonicus, soybean (Glycine max), peanut (Arachis hypogaea), red clover (Trifolium pratense), white clover (Trifolium repens), and eucalyptus (Eucalyptus camaldulensis). In addition, the number of plant species registered in this database will be increased as our research progresses. The Kazusa Marker DataBase will be a useful tool for both basic and applied sciences, such as genomics, genetics, and molecular breeding in crops.

  9. Use of Both Cumulus Cells’ Transcriptomic Markers and Zona Pellucida Birefringence to Select Developmentally Competent Oocytes in Human Assisted Reproductive Technologies

    PubMed Central

    2015-01-01

    Background Selection of the best oocyte for subsequent steps of fertilization and embryo transfer was shown to be the crucial step in human infertility treatment procedure. Oocyte selection using morphological criteria mainly Zona pellucida (ZP) has been the gold standard method in assisted reproductive technologies (ART) clinics, but this selection approach has limitations in terms of accuracy, objectivity and constancy. Recent studies using OMICs-based approaches have allowed the identification of key molecular markers that quantitatively and non-invasively predict the oocyte quality for higher pregnancy rates and efficient infertility treatment. These biomarkers are a valuable reinforcement of the morphological selection criteria widely used in in vitro fertilization (IVF) clinics. In this context, this study was designed to investigate the relationship between transcriptomic predictors of oocyte quality found by our group and the conventional morphological parameters of oocyte quality mainly the ZP birefringence. Results Microarray data revealed that 48 and 27 differentially expressed candidate genes in cumulus cells (CCs) were respectively overexpressed and underexpressed in the ZGP (Zona Good Pregnant) versus ZBNP (Zona Bad Non Pregnant) groups. More than 70% of previously reported transcriptomic biomarkers of oocyte developmental competence were confirmed in this study. The analysis of possible association between ZP birefringence versus molecular markers approach showed an absence of correlation between them using the current set of markers. Conclusions This study suggested a new integrative approach that matches morphological and molecular approaches used to select developmentally competent oocytes able to lead to successful pregnancy and the delivery of healthy baby. For each ZP birefringence score, oocytes displayed a particular CCs' gene expression pattern. However, no correlations were found between the 7 gene biomarkers of oocyte developmental

  10. Comparison of ECG-based physiological markers for hypoxia in a preterm ovine model.

    PubMed

    Zwanenburg, Alex; Hermans, Ben Jm; Andriessen, Peter; Niemarkt, Hendrik J; Jellema, Reint K; Ophelders, Daan Rmg; Vullings, Rik; Wolfs, Tim Gam; Kramer, Boris W; Delhaas, Tammo

    2016-06-01

    Current methods for assessing perinatal hypoxic conditions did not improve infant outcomes. Various waveform-based and interval-based ECG markers have been suggested, but not directly compared. We compare performance of ECG markers in a standardized ovine model for fetal hypoxia. Sixty-nine fetal sheep of 0.7 gestation had ECG recorded 4 h before, during, and 4 h after a 25-min period of umbilical cord occlusion (UCO), leading to severe hypoxia. Various ECG markers were calculated, among which were heart rate (HR), HR-corrected ventricular depolarization/repolarization interval (QTc), and ST-segment analysis (STAN) episodic and baseline rise markers, analogue to clinical STAN device alarms. Performance of interval- and waveform-based ECG markers was assessed by correlating predicted and actual hypoxic/normoxic state. Of the markers studied, HR and QTc demonstrated high sensitivity (≥86%), specificity (≥96%), and positive predictive value (PPV) (≥86%) and detected hypoxia in ≥90% of fetuses at 4 min after UCO. In contrast, STAN episodic and baseline rise markers displayed low sensitivity (≤20%) and could not detect severe fetal hypoxia in 65 and 28% of the animals, respectively. Interval-based HR and QTc markers could assess the presence of severe hypoxia. Waveform-based STAN episodic and baseline rise markers were ineffective as markers for hypoxia.

  11. A flowchart for selecting an ointment base.

    PubMed

    Conway, Jeannine M; Brown, Michael C

    2014-02-12

    OBJECTIVES. To improve students' skills in selecting appropriate ointment bases through the development and implementation of a flowchart. A flowchart was designed to help students select the appropriate base for an ointment. Students used the flowchart throughout the semester in both dry and wet laboratory activities. At the end of the semester, students completed a dry laboratory practical that required them to select an appropriate ointment base and levigating agent. Student performance data from the year prior to implementation was compared to data for 2 years after implementation. Calculation, procedure, and labeling errors also were compared. Prior to implementation of the flowchart, 51 of 101 students selected the correct base. After implementation, 169 of 212 students selected the correct base (p<0.001). Calculation, procedure, and labeling errors were not significantly different. CONCLUSION. Use of a flowchart to select an ointment base improved student performance when used in the context of a dry laboratory assignment.

  12. Multivariate analysis for the estimation of target localization errors in fiducial marker-based radiotherapy

    SciTech Connect

    Takamiya, Masanori; Nakamura, Mitsuhiro Akimoto, Mami; Ueki, Nami; Yamada, Masahiro; Matsuo, Yukinori; Mizowaki, Takashi; Hiraoka, Masahiro; Tanabe, Hiroaki; Kokubo, Masaki; Itoh, Akio

    2016-04-15

    Purpose: To assess the target localization error (TLE) in terms of the distance between the target and the localization point estimated from the surrogates (|TMD|), the average of respiratory motion for the surrogates and the target (|aRM|), and the number of fiducial markers used for estimating the target (n). Methods: This study enrolled 17 lung cancer patients who subsequently underwent four fractions of real-time tumor tracking irradiation. Four or five fiducial markers were implanted around the lung tumor. The three-dimensional (3D) distance between the tumor and markers was at maximum 58.7 mm. One of the markers was used as the target (P{sub t}), and those markers with a 3D |TMD{sub n}| ≤ 58.7 mm at end-exhalation were then selected. The estimated target position (P{sub e}) was calculated from a localization point consisting of one to three markers except P{sub t}. Respiratory motion for P{sub t} and P{sub e} was defined as the root mean square of each displacement, and |aRM| was calculated from the mean value. TLE was defined as the root mean square of each difference between P{sub t} and P{sub e} during the monitoring of each fraction. These procedures were performed repeatedly using the remaining markers. To provide the best guidance on the answer with n and |TMD|, fiducial markers with a 3D |aRM ≥ 10 mm were selected. Finally, a total of 205, 282, and 76 TLEs that fulfilled the 3D |TMD| and 3D |aRM| criteria were obtained for n = 1, 2, and 3, respectively. Multiple regression analysis (MRA) was used to evaluate TLE as a function of |TMD| and |aRM| in each n. Results: |TMD| for n = 1 was larger than that for n = 3. Moreover, |aRM| was almost constant for all n, indicating a similar scale for the marker’s motion near the lung tumor. MRA showed that |aRM| in the left–right direction was the major cause of TLE; however, the contribution made little difference to the 3D TLE because of the small amount of motion in the left–right direction. The TLE

  13. Phenotypic screening and molecular analysis of blast resistance in fragrant rice for marker assisted selection.

    PubMed

    Khan, Mohammad Ashik Iqbal; Sen, Partha Pratim; Bhuiyan, Rejwan; Kabir, Enamul; Chowdhury, Abul Kashem; Fukuta, Yoshimichi; Ali, Ansar; Latif, Mohammad Abdul

    2014-05-01

    Experiments were conducted to identify blast-resistant fragrant genotypes for the development of a durable blast-resistant rice variety during years 2012-2013. The results indicate that out of 140 test materials including 114 fragrant germplasms, 25 differential varieties (DVs) harbouring 23 blast-resistant genes, only 16 fragrant rice germplasms showed comparatively better performance against a virulent isolate of blast disease. The reaction pattern of single-spore isolate of Magnaporthe oryzae to differential varieties showed that Pish, Pi9, Pita-2 and Pita are the effective blast-resistant genes against the tested blast isolates in Bangladesh. The DNA markers profiles of selected 16 rice germplasms indicated that genotype Chinigura contained Pish, Pi9 and Pita genes; on the other hand, both BRRI dhan50 and Bawaibhog contained Pish and Pita genes in their genetic background. Genotypes Jirakatari, BR5, and Gopalbhog possessed Pish gene, while Uknimodhu, Deshikatari, Radhunipagol, Kalijira (3), Chinikanai each contained the Pita gene only. There are some materials that did not contain any target gene(s) in their genetic background, but proved resistant in pathogenicity tests. This information provided valuable genetic information for breeders to develop durable blast-resistant fragrant or aromatic rice varieties in Bangladesh.

  14. Ebf2 is a selective marker of brown and beige adipogenic precursor cells.

    PubMed

    Wang, Wenshan; Kissig, Megan; Rajakumari, Sona; Huang, Li; Lim, Hee-Woong; Won, Kyoung-Jae; Seale, Patrick

    2014-10-07

    Brown adipocytes and muscle and dorsal dermis descend from precursor cells in the dermomyotome, but the factors that regulate commitment to the brown adipose lineage are unknown. Here, we prospectively isolated and determined the molecular profile of embryonic brown preadipose cells. Brown adipogenic precursor activity in embryos was confined to platelet-derived growth factor α(+), myogenic factor 5(Cre)-lineage-marked cells. RNA-sequence analysis identified early B-cell factor 2 (Ebf2) as one of the most selectively expressed genes in this cell fraction. Importantly, Ebf2-expressing cells purified from Ebf2(GFP) embryos or brown fat tissue did not express myoblast or dermal cell markers and uniformly differentiated into brown adipocytes. Interestingly, Ebf2-expressing cells from white fat tissue in adult animals differentiated into brown-like (or beige) adipocytes. Loss of Ebf2 in brown preadipose cells reduced the expression levels of brown preadipose-signature genes, whereas ectopic Ebf2 expression in myoblasts activated brown preadipose-specific genes. Altogether, these results indicate that Ebf2 specifically marks and regulates the molecular profile of brown preadipose cells.

  15. Performance of variable selection methods using stability-based selection.

    PubMed

    Lu, Danny; Weljie, Aalim; de Leon, Alexander R; McConnell, Yarrow; Bathe, Oliver F; Kopciuk, Karen

    2017-04-04

    Variable selection is frequently carried out during the analysis of many types of high-dimensional data, including those in metabolomics. This study compared the predictive performance of four variable selection methods using stability-based selection, a new secondary selection method that is implemented in the R package BioMark. Two of these methods were evaluated using the more well-known false discovery rate (FDR) as well. Simulation studies varied factors relevant to biological data studies, with results based on the median values of 200 partial area under the receiver operating characteristic curve. There was no single top performing method across all factor settings, but the student t test based on stability selection or with FDR adjustment and the variable importance in projection (VIP) scores from partial least squares regression models obtained using a stability-based approach tended to perform well in most settings. Similar results were found with a real spiked-in metabolomics dataset. Group sample size, group effect size, number of significant variables and correlation structure were the most important factors whereas the percentage of significant variables was the least important. Researchers can improve prediction scores for their study data by choosing VIP scores based on stability variable selection over the other approaches when the number of variables is small to modest and by increasing the number of samples even moderately. When the number of variables is high and there is block correlation amongst the significant variables (i.e., true biomarkers), the FDR-adjusted student t test performed best. The R package BioMark is an easy-to-use open-source program for variable selection that had excellent performance characteristics for the purposes of this study.

  16. PL1 fusion gene: a novel visual selectable marker gene that confers tolerance to multiple abiotic stresses in transgenic tomato.

    PubMed

    Jin, Feng; Li, Shu; Dang, Lijie; Chai, Wenting; Li, Pengli; Wang, Ning Ning

    2012-10-01

    Visual selectable markers, including the purple color caused by the accumulation of anthocyanins, have been proposed for use as antibiotic-free alternatives. However, the excessive accumulation of anthocyanins seriously inhibits the growth and development of transgenic plants. In our study, the AtDWF4 promoter from Arabidopsis and the tomato LeANT1 gene, encoding a MYB transcription factor, were used to construct the PL1 fusion gene to test whether it could be used as a visual selectable marker gene for tomato transformation. All the PL1 transgenic shoots exhibited intense purple color on shoot induction medium. In the transgenic tomato plants, PL1 was highly expressed in the cotyledons, but expressed only slightly in the true leaves and other organs. The expression of PL1 had no significantly adverse effects on the growth or development of the transgenic tomato plants, and conferred tolerance to multiple abiotic stresses in them. With the “cut off green shoots” method, multiple independent 35S::GFP transgenic tomato lines were successfully obtained using PL1 as the selectable marker gene. These results suggest that PL1 has potential application of visual selectable marker gene for tomato transformation.

  17. A high efficiency gene disruption strategy using a positive-negative split selection marker and electroporation for Fusarium oxysporum.

    PubMed

    Liang, Liqin; Li, Jianqiang; Cheng, Lin; Ling, Jian; Luo, Zhongqin; Bai, Miao; Xie, Bingyan

    2014-11-01

    The Fusarium oxysporum species complex consists of fungal pathogens that cause serial vascular wilt disease on more than 100 cultivated species throughout the world. Gene function analysis is rapidly becoming more and more important as the whole-genome sequences of various F. oxysporum strains are being completed. Gene-disruption techniques are a common molecular tool for studying gene function, yet are often a limiting step in gene function identification. In this study we have developed a F. oxysporum high-efficiency gene-disruption strategy based on split-marker homologous recombination cassettes with dual selection and electroporation transformation. The method was efficiently used to delete three RNA-dependent RNA polymerase (RdRP) genes. The gene-disruption cassettes of three genes can be constructed simultaneously within a short time using this technique. The optimal condition for electroporation is 10μF capacitance, 300Ω resistance, 4kV/cm field strength, with 1μg of DNA (gene-disruption cassettes). Under these optimal conditions, we were able to obtain 95 transformants per μg DNA. And after positive-negative selection, the transformants were efficiently screened by PCR, screening efficiency averaged 85%: 90% (RdRP1), 85% (RdRP2) and 77% (RdRP3). This gene-disruption strategy should pave the way for high throughout genetic analysis in F. oxysporum. Copyright © 2014 Elsevier GmbH. All rights reserved.

  18. Identification of QTLs for Rust Resistance in the Peanut Wild Species Arachis magna and the Development of KASP Markers for Marker-Assisted Selection

    PubMed Central

    Leal-Bertioli, Soraya C. M.; Cavalcante, Uiara; Gouvea, Ediene G.; Ballén-Taborda, Carolina; Shirasawa, Kenta; Guimarães, Patrícia M.; Jackson, Scott A.; Bertioli, David J.; Moretzsohn, Márcio C.

    2015-01-01

    Rust is a major pathogen of the peanut crop. Development and adoption of rust-resistant cultivars is the most cost efficient and effective way to control the spread of the disease and reduce yield losses. Some cultivated peanut germplasm accessions have a degree of resistance, but the secondary gene pool is a source of much stronger resistance alleles. Wild species, however, have undesirable agronomic traits that are a disincentive to their use in breeding. The identification of genomic regions that harbor disease resistance in wild species is the first step in the implementation of marker-assisted selection that can speed the introgression of wild disease resistances and the elimination of linkage drag. In this work, we identify genome regions that control different components of rust resistance in a recombinant inbred line population developed from a cross between two Arachis species, the susceptible most probable B genome ancestor of cultivated peanut, Arachis ipaënsis, and an accession of its closest relative, Arachis magna, which is resistant to rust. Quantitative trait loci for several components of resistance were placed in the same position on linkage group B08. Single-nucleotide polymorphism Kompetitive allele-specific polymerase chain reaction markers for rust resistance region were designed and validated for marker function in both diploid and tetraploid contexts. PMID:25943521

  19. Identification of QTLs for Rust Resistance in the Peanut Wild Species Arachis magna and the Development of KASP Markers for Marker-Assisted Selection.

    PubMed

    Leal-Bertioli, Soraya C M; Cavalcante, Uiara; Gouvea, Ediene G; Ballén-Taborda, Carolina; Shirasawa, Kenta; Guimarães, Patrícia M; Jackson, Scott A; Bertioli, David J; Moretzsohn, Márcio C

    2015-05-05

    Rust is a major pathogen of the peanut crop. Development and adoption of rust-resistant cultivars is the most cost efficient and effective way to control the spread of the disease and reduce yield losses. Some cultivated peanut germplasm accessions have a degree of resistance, but the secondary gene pool is a source of much stronger resistance alleles. Wild species, however, have undesirable agronomic traits that are a disincentive to their use in breeding. The identification of genomic regions that harbor disease resistance in wild species is the first step in the implementation of marker-assisted selection that can speed the introgression of wild disease resistances and the elimination of linkage drag. In this work, we identify genome regions that control different components of rust resistance in a recombinant inbred line population developed from a cross between two Arachis species, the susceptible most probable B genome ancestor of cultivated peanut, Arachis ipaënsis, and an accession of its closest relative, Arachis magna, which is resistant to rust. Quantitative trait loci for several components of resistance were placed in the same position on linkage group B08. Single-nucleotide polymorphism Kompetitive allele-specific polymerase chain reaction markers for rust resistance region were designed and validated for marker function in both diploid and tetraploid contexts. Copyright © 2015 Leal-Bertioli et al.

  20. A bacterial gene codA encoding cytosine deaminase is an effective conditional negative selectable marker in Glycine max.

    PubMed

    Shao, Min; Michno, Jean-Michel; Hotton, Sara K; Blechl, Ann; Thomson, James

    2015-10-01

    Research describes the practical application of the codA negative selection marker in Soybean. Conditions are given for codA selection at both the shooting and rooting stages of regeneration. Conditional negative selection is a powerful technique whereby the absence of a gene product allows survival in otherwise lethal conditions. In plants, the Escherichia coli gene codA has been employed as a negative selection marker. Our research demonstrates that codA can be used as a negative selection marker in soybean, Glycine max. Like most plants, soybean does not contain cytosine deaminase activity and we show here that wild-type seedlings are not affected by inclusion of 5-FC in growth media. In contrast, transgenic G. max plants expressing codA and grown in the presence of more than 200 μg/mL 5-FC exhibit reductions in hypocotyl and taproot lengths, and severe suppression of lateral root development. We also demonstrate a novel negative selection-rooting assay in which codA-expressing aerial tissues or shoot cuttings are inhibited for root formation in media containing 5-FC. Taken together these techniques allow screening during either the regeneration or rooting phase of tissue culture.

  1. Exploring a new bilateral focal density asymmetry based image marker to predict breast cancer risk

    NASA Astrophysics Data System (ADS)

    Aghaei, Faranak; Mirniaharikandehei, Seyedehnafiseh; Hollingsworth, Alan B.; Wang, Yunzhi; Qiu, Yuchen; Liu, Hong; Zheng, Bin

    2017-03-01

    Although breast density has been widely considered an important breast cancer risk factor, it is not very effective to predict risk of developing breast cancer in a short-term or harboring cancer in mammograms. Based on our recent studies to build short-term breast cancer risk stratification models based on bilateral mammographic density asymmetry, we in this study explored a new quantitative image marker based on bilateral focal density asymmetry to predict the risk of harboring cancers in mammograms. For this purpose, we assembled a testing dataset involving 100 positive and 100 negative cases. In each of positive case, no any solid masses are visible on mammograms. We developed a computer-aided detection (CAD) scheme to automatically detect focal dense regions depicting on two bilateral mammograms of left and right breasts. CAD selects one focal dense region with the maximum size on each image and computes its asymmetrical ratio. We used this focal density asymmetry as a new imaging marker to divide testing cases into two groups of higher and lower focal density asymmetry. The first group included 70 cases in which 62.9% are positive, while the second group included 130 cases in which 43.1% are positive. The odds ratio is 2.24. As a result, this preliminary study supported the feasibility of applying a new focal density asymmetry based imaging marker to predict the risk of having mammography-occult cancers. The goal is to assist radiologists more effectively and accurately detect early subtle cancers using mammography and/or other adjunctive imaging modalities in the future.

  2. Risk prediction of small pulmonary nodules based on novel CT image texture markers

    NASA Astrophysics Data System (ADS)

    Han, Fangfang; Song, Bowen; Ma, He; Qian, Wei; Liang, Zhengrong

    2017-03-01

    Among the detected small nodules sized from 3 to 30mm in CT images, a significant portion is undetermined in terms of malignancy which needs biopsy or other follow-up means, resulting in excessive risk and cost. Therefore, predicting the malignancy of the nodules becomes a clinically desirable task. Based on the previous study of texture features extracted from gray-tone spatial-dependence matrices, this study aims to find more efficient texture features or image texture markers in discriminating the nodule malignancy. Two new image texture markers (median and variance) are proposed to classify the small nodules into different malignant levels, thus the risk prediction could be performed through image analysis. These two new image texture markers can minimize the effect of outliers in the feature series, thus can reduce the noise influence to the feature classification. Total 1,353 nodule samples selected from the Lung Image Database Consortium were used to evaluate the efficiency of the proposed new features. All the classification results are shown in the ROC curves and tabulated by the AUC values. The classification outcomes from (1) the most likely and likely benign nodules vs. the most likely and likely malignant nodules, (2) the most likely vs. likely benign nodules, and (3) the most likely vs. likely malignant nodules, are 0.9125+/-0.0096, 0.9239+/-0.0147, and 0.8888+/-0.0197, respectively, in terms of the largest AUC values. From the experimental outcomes on different malignant levels, the two new image texture markers from nodule volumetric CT image data have shown encouraging performance for the risk prediction.

  3. Levels of selected oxidative stress markers in the vitreous and serum of diabetic retinopathy patients

    PubMed Central

    Brzović-Šarić, Vlatka; Landeka, Irena; Šarić, Borna; Barberić, Monika; Andrijašević, Lidija; Cerovski, Branimir; Oršolić, Nada

    2015-01-01

    the selected oxidative stress markers, SOD and LPO were highly correlative in both the vitreous and serum in PDR compared to patients without metabolic disorders. Their correlations suggested that monitoring their mutual alterations might be informative during PDR development and should be considered in further research. PMID:26120270

  4. Algorithm based on marker-controlled watershed transform for overlapping plant fruit segmentation

    NASA Astrophysics Data System (ADS)

    Zeng, Qingbing; Miao, Yubin; Liu, Chengliang; Wang, Shiping

    2009-02-01

    Overlapping is a major problem for machine vision applications in agriculture. We present a robust marker-controlled watershed transform algorithm to automatically perform the accurate segmentation of overlapping plant fruits. The marker-controlled watershed algorithm mainly involves image preprocessing, marker extraction, and watershed transform. Marker extraction is the most important and difficult step of the whole process. Using K-means clustering, cut point decision making, spline interpolating, and morphological processing, markers can be detected automatically. Due to the good localization performance of detected markers, the accurate contour of separated fruits can be extracted by the watershed transform based on detected markers. The face validity of the segmentation algorithm is tested with a set of grape images, and segmentation results are overlaid onto original images for visual inspection. The algorithm is further evaluated by comparing segmentation results with a ``gold standard'' established by professional agronomists. Quantitative comparison shows that the segmentation algorithm can obtain very good spatial segmentation results.

  5. Optical detection of sepsis markers using liquid crystal based biosensors

    NASA Astrophysics Data System (ADS)

    McCamley, Maureen K.; Artenstein, Andrew W.; Opal, Steven M.; Crawford, Gregory P.

    2007-02-01

    A liquid crystal based biosensor for the detection and diagnosis of sepsis is currently in development. Sepsis, a major clinical syndrome with a significant public health burden in the US due to a large elderly population, is the systemic response of the body to a localized infection and is defined as the combination of pathologic infection and physiological changes. Bacterial infections are responsible for 90% of cases of sepsis in the US. Currently there is no bedside diagnostic available to positively identify sepsis. The basic detection scheme employed in a liquid crystal biosensor contains attributes that would find value in a clinical setting, especially for the early detection of sepsis. Utilizing the unique properties of liquid crystals, such as birefringence, a bedside diagnostic is in development which will optically report the presence of biomolecules. In a septic patient, an endotoxin known as lipopolysaccharide (LPS) is released from the outer membrane of Gram-negative bacteria and can be found in the blood stream. It is hypothesized that this long chained molecule will cause local disruptions to the open surface of a sensor containing aligned liquid crystal. The bulk liquid crystal ampli.es these local changes at the surface due to the presence of the sepsis marker, providing an optical readout through polarizing microscopy images. Liquid crystal sensors consisting of both square and circular grids, 100-200 μm in size, have been fabricated and filled with a common liquid crystal material, 5CB. Homeotropic alignment was confirmed using polarizing microscopy. The grids were then contacted with either saline only (control), or saline with varying concentrations of LPS. Changes in the con.guration of the nematic director of the liquid crystal were observed through the range of concentrations tested (5mg/mL - 1pg/mL) which have been confirmed by a consulting physician as clinically relevant levels.

  6. Biomedical wellness monitoring system based upon molecular markers

    NASA Astrophysics Data System (ADS)

    Ingram, Whitney

    2012-06-01

    We wish to assist caretakers with a sensor monitoring systems for tracking the physiological changes of homealone patients. One goal is seeking biomarkers and modern imaging sensors like stochastic optical reconstruction microscopy (STORM), which has achieved visible imaging at the nano-scale range. Imaging techniques like STORM can be combined with a fluorescent functional marker in a system to capture the early transformation signs from wellness to illness. By exploiting both microscopic knowledge of genetic pre-disposition and the macroscopic influence of epigenetic factors we hope to target these changes remotely. We adopt dual spectral infrared imaging for blind source separation (BSS) to detect angiogenesis changes and use laser speckle imaging for hypertension blood flow monitoring. Our design hypothesis for the monitoring system is guided by the user-friendly, veteran-preferred "4-Non" principles (noninvasive, non-contact, non-tethered, non-stop-to-measure) and by the NIH's "4Ps" initiatives (predictive, personalized, preemptive, and participatory). We augment the potential storage system with the recent know-how of video Compressive Sampling (CSp) from surveillance cameras. In CSp only major changes are saved, which reduces the manpower cost of caretakers and medical analysts. This CSp algorithm is based on smart associative memory (AM) matrix storage: change features and detailed scenes are written by the outer-product and read by the inner product without the usual Harsh index for image searching. From this approach, we attempt to design an effective household monitoring approach to save healthcare costs and maintain the quality of life of seniors.

  7. Development and Validation of Marker-Aided Selection Methods for Wood Property Traits in Loblolly Pine and Hybrid Poplar

    SciTech Connect

    Tuskan, G.A.

    2001-06-20

    Wood properties influence pulp and paper quality. Certainly, overall pulp yields are directly related to the cellulose content, changes in hemicellulose content are associated with changes in pulp cohesiveness, and pulping efficiency is related to lignin content. Despite the importance of wood properties on product quality, little progress has been made in improving such traits because current methods of assessing wood and fiber characteristics are time-consuming, expensive, and often imprecise. Genetic improvement of wood and fiber properties has been further hampered by the large size of trees, delayed reproductive maturity and long harvest cycles. Recent developments in molecular genetics will help overcome the physical, economic and biological constraints in assessing and improving wood properties. Genetic maps consisting of numerous molecular markers are now available for loblolly pine and hybrid poplar. Such markers/maps may be used as part of a marker-aided selection and breeding effort or to expedite the isolation and characterization of genes and/or promoters that directly control wood properties. The objectives of this project are: (1) to apply new and rapid analytical techniques for assessing component wood properties to segregating F2 progeny populations of loblolly pine and hybrid poplar, (2) to map quantitative trait loci and identify molecular markers associated with wood properties in each of the above species and (3) to validate marker-aided selection methods for wood properties in loblolly pine and hybrid poplar.

  8. Selected eosinophil proteins as markers of inflammation in atopic dermatitis patients.

    PubMed

    Jenerowicz, Dorotaz; Czarnecka-Operacz, Magdalena; Silny, Wojciech

    2006-01-01

    Atopic dermatitis (AD) is an inflammatory skin disease characterized by chronic and recurrent course, beginning primarily in early childhood. The etiopathogenesis of AD has not yet been fully understood, although various types of inflammatory cells including eosinophils may be involved in its pathomechanism. The basic aim of the study was to evaluate the usefulness of selected eosinophil proteins in serum and urine of AD patients, as markers of disease severity. The study also aimed to analyze correlations between the level of examined proteins and parameters such as skin prick test (SPT) results, serum concentration of total IgE, and coexistence of symptoms of other atopic diseases. The study included 30 AD patients and two control groups: 30 patients suffering from chronic urticaria and 30 healthy individuals. The mean level of eosinophil proteins measured in serum and urine of AD patients was higher than that in controls, although a significant difference was only recorded for serum and urine level of eosinophil protein X (EPX). Patients with very severe/severe AD presented higher levels of eosinophil proteins than patients presenting with mild/moderate AD, although no significant difference was found between these two groups. AD patients with positive SPT results and detectable specific IgE in serum, and with coexisting symptoms of other atopic diseases presented with higher mean levels of serum and urine eosinophil proteins than AD cases with negative SPT results and without any symptoms of other atopic diseases. In children suffering from AD, serum eosinophil cationic protein level, EPX level and urine EPX level were higher than those in healthy children, however, without statistical significance. Study results suggested a significant role of eosinophils in the etiopathogenesis of AD. Serum and urine levels of selected eosinophil proteins may serve as an important part of diagnostic approach to AD patients, especially in differentiation of allergic and non

  9. Fundamental Vocabulary Selection Based on Word Familiarity

    NASA Astrophysics Data System (ADS)

    Sato, Hiroshi; Kasahara, Kaname; Kanasugi, Tomoko; Amano, Shigeaki

    This paper proposes a new method for selecting fundamental vocabulary. We are presently constructing the Fundamental Vocabulary Knowledge-base of Japanese that contains integrated information on syntax, semantics and pragmatics, for the purposes of advanced natural language processing. This database mainly consists of a lexicon and a treebank: Lexeed (a Japanese Semantic Lexicon) and the Hinoki Treebank. Fundamental vocabulary selection is the first step in the construction of Lexeed. The vocabulary should include sufficient words to describe general concepts for self-expandability, and should not be prohibitively large to construct and maintain. There are two conventional methods for selecting fundamental vocabulary. The first is intuition-based selection by experts. This is the traditional method for making dictionaries. A weak point of this method is that the selection strongly depends on personal intuition. The second is corpus-based selection. This method is superior in objectivity to intuition-based selection, however, it is difficult to compile a sufficiently balanced corpora. We propose a psychologically-motivated selection method that adopts word familiarity as the selection criterion. Word familiarity is a rating that represents the familiarity of a word as a real number ranging from 1 (least familiar) to 7 (most familiar). We determined the word familiarity ratings statistically based on psychological experiments over 32 subjects. We selected about 30,000 words as the fundamental vocabulary, based on a minimum word familiarity threshold of 5. We also evaluated the vocabulary by comparing its word coverage with conventional intuition-based and corpus-based selection over dictionary definition sentences and novels, and demonstrated the superior coverage of our lexicon. Based on this, we conclude that the proposed method is superior to conventional methods for fundamental vocabulary selection.

  10. Genetic diversity and relatedness of sweet cherry (prunus avium L.) cultivars based on single nucleotide polymorphic markers.

    PubMed

    Fernandez I Marti, Angel; Athanson, Blessing; Koepke, Tyson; Font I Forcada, Carolina; Dhingra, Amit; Oraguzie, Nnadozie

    2012-01-01

    Most previous studies on genetic fingerprinting and cultivar relatedness in sweet cherry were based on isoenzyme, RAPD, and simple sequence repeat (SSR) markers. This study was carried out to assess the utility of single nucleotide polymorphism (SNP) markers generated from 3' untranslated regions (UTR) for genetic fingerprinting in sweet cherry. A total of 114 sweet cherry germplasm representing advanced selections, commercial cultivars, and old cultivars imported from different parts of the world were screened with seven SSR markers developed from other Prunus species and with 40 SNPs obtained from 3' UTR sequences of Rainier and Bing sweet cherry cultivars. Both types of marker study had 99 accessions in common. The SSR data was used to validate the SNP results. Results showed that the average number of alleles per locus, mean observed heterozygosity, expected heterozygosity, and polymorphic information content values were higher in SSRs than in SNPs although both set of markers were similar in their grouping of the sweet cherry accessions as shown in the dendrogram. SNPs were able to distinguish sport mutants from their wild type germplasm. For example, "Stella" was separated from "Compact Stella." This demonstrates the greater power of SNPs for discriminating mutants from their original parents than SSRs. In addition, SNP markers confirmed parentage and also determined relationships of the accessions in a manner consistent with their pedigree relationships. We would recommend the use of 3' UTR SNPs for genetic fingerprinting, parentage verification, gene mapping, and study of genetic diversity in sweet cherry.

  11. A bayesian mixed regression based prediction of quantitative traits from molecular marker and gene expression data.

    PubMed

    Bhattacharjee, Madhuchhanda; Sillanpää, Mikko J

    2011-01-01

    Both molecular marker and gene expression data were considered alone as well as jointly to serve as additive predictors for two pathogen-activity-phenotypes in real recombinant inbred lines of soybean. For unobserved phenotype prediction, we used a bayesian hierarchical regression modeling, where the number of possible predictors in the model was controlled by different selection strategies tested. Our initial findings were submitted for DREAM5 (the 5th Dialogue on Reverse Engineering Assessment and Methods challenge) and were judged to be the best in sub-challenge B3 wherein both functional genomic and genetic data were used to predict the phenotypes. In this work we further improve upon this previous work by considering various predictor selection strategies and cross-validation was used to measure accuracy of in-data and out-data predictions. The results from various model choices indicate that for this data use of both data types (namely functional genomic and genetic) simultaneously improves out-data prediction accuracy. Adequate goodness-of-fit can be easily achieved with more complex models for both phenotypes, since the number of potential predictors is large and the sample size is not small. We also further studied gene-set enrichment (for continuous phenotype) in the biological process in question and chromosomal enrichment of the gene set. The methodological contribution of this paper is in exploration of variable selection techniques to alleviate the problem of over-fitting. Different strategies based on the nature of covariates were explored and all methods were implemented under the bayesian hierarchical modeling framework with indicator-based covariate selection. All the models based in careful variable selection procedure were found to produce significant results based on permutation test.

  12. Selective, reliable blood and milk bio-markers for diagnosing clinical and subclinical bovine mastitis.

    PubMed

    Sadek, Kadry; Saleh, Ebeed; Ayoub, Mousa

    2017-02-01

    Mastitis is positioned as the most vital ailment in dairy cows in light of conventional cost examinations. The aim of the present study was to evaluate the diagnostic value of different acute phase proteins (APPs), pro-inflammatory cytokines, and oxidative stress biomarkers in healthy cows and in those with clinical or subclinical mastitis and to localize APP gene expression in the milk of mastitic cows. Therefore, 20 subclinical mastitic cows with positive California Mastitis Test (CMT) results and no clinical signs of mastitis, 15 clinically mastitic cows, and 15 healthy cows with negative CMT results and somatic cell count (SCC) of <600,000 cells/mL were selected. Milk and blood samples were collected. The present findings indicate that the biochemical parameters examined were significantly (p < 0.05) increased in cows with both types of mastitis, except for total protein, albumin, and GSH levels and the TAC, which were significantly (p < 0.05) decreased, compared with values in the controls. Surprisingly, SAA and Hp gene expression were up-regulated in milk from cows with both forms of mastitis, while Fb expression was absent. The present study demonstrates that APPs, pro-inflammatory cytokines, and indicators of oxidative stress may serve as biomarkers of clinical and subclinical mastitis. Interestingly, the expression of SAA and Hp indicates the local de novo synthesis of these APPs within the mammary glands. Furthermore, the presence of SAA and Hp transcripts in milk cells derived from pathogen-free mammary glands proved their constitutive expression. However, future studies with more extensive baseline sampling are still needed to establish and validate the reference values for APPs, cytokines, and oxidative stress markers in cows.

  13. Systematic Differences in the Response of Genetic Variation to Pedigree and Genome Based Selection Methods

    USDA-ARS?s Scientific Manuscript database

    Genomic selection (GS) is a DNA-based method of selecting for quantitative traits in animal and plant breeding, and offers a potentially superior alternative to traditional breeding methods that rely on pedigree and phenotype information. Using a 60 K SNP chip with markers spaced throughout the enti...

  14. Fluorescent sperm in a transparent worm: validation of a GFP marker to study sexual selection

    PubMed Central

    2014-01-01

    Background Sexual selection has initially been thought to occur exclusively at the precopulatory stage in terms of contests among males and female mate choice, but research over the last four decades revealed that it often continues after copulation through sperm competition and cryptic female choice. However, studying these postcopulatory processes remains challenging because they occur internally and therefore are often difficult to observe. In the transparent free-living flatworm Macrostomum lignano, a recently established transgenic line that expresses green fluorescent protein (GFP) in all cell types, including sperm, offers a unique opportunity to non-invasively visualise and quantify the sperm of a GFP-expressing donor inside the reproductive tract of wild-type recipients in vivo. We here test several aspects of the reproductive performance of the transgenic individuals and the accuracy of the techniques involved in assessing the GFP-expressing worms and their sperm. We then show the usefulness of these methods in a study on sperm displacement. Results GFP-expressing worms do not differ from wild-type worms in terms of morphology, mating rate and reproductive success. In addition, we show that the GFP signal is reliably and unequivocally expressed by all GFP-expressing individuals observed under epifluorescence illumination. However, the intensity of the GFP signal emitted by sperm of GFP expressing donors can vary (which we show to be at least in part due to sperm ageing) and the GFP marker is inherited according to Mendel’s laws in most, but not all, of the individuals. Nevertheless, we argue these two issues can be addressed with an appropriate experimental design. Finally, we demonstrate the value of the GFP-techniques by comparing the number of GFP-expressing sperm in a wild-type recipient before and after mating with a competing sperm donor, providing clear experimental evidence for sperm displacement in M. lignano. This result suggests that sperm

  15. Development of New Candidate Gene and EST-Based Molecular Markers for Gossypium Species

    PubMed Central

    Buyyarapu, Ramesh; Kantety, Ramesh V.; Yu, John Z.; Saha, Sukumar; Sharma, Govind C.

    2011-01-01

    New source of molecular markers accelerate the efforts in improving cotton fiber traits and aid in developing high-density integrated genetic maps. We developed new markers based on candidate genes and G. arboreum EST sequences that were used for polymorphism detection followed by genetic and physical mapping. Nineteen gene-based markers were surveyed for polymorphism detection in 26 Gossypium species. Cluster analysis generated a phylogenetic tree with four major sub-clusters for 23 species while three species branched out individually. CAP method enhanced the rate of polymorphism of candidate gene-based markers between G. hirsutum and G. barbadense. Two hundred A-genome based SSR markers were designed after datamining of G. arboreum EST sequences (Mississippi Gossypium arboreum   EST-SSR: MGAES). Over 70% of MGAES markers successfully produced amplicons while 65 of them demonstrated polymorphism between the parents of G. hirsutum and G. barbadense RIL population and formed 14 linkage groups. Chromosomal localization of both candidate gene-based and MGAES markers was assisted by euploid and hypoaneuploid CS-B analysis. Gene-based and MGAES markers were highly informative as they were designed from candidate genes and fiber transcriptome with a potential to be integrated into the existing cotton genetic and physical maps. PMID:22315588

  16. Development of New Candidate Gene and EST-Based Molecular Markers for Gossypium Species.

    PubMed

    Buyyarapu, Ramesh; Kantety, Ramesh V; Yu, John Z; Saha, Sukumar; Sharma, Govind C

    2011-01-01

    New source of molecular markers accelerate the efforts in improving cotton fiber traits and aid in developing high-density integrated genetic maps. We developed new markers based on candidate genes and G. arboreum EST sequences that were used for polymorphism detection followed by genetic and physical mapping. Nineteen gene-based markers were surveyed for polymorphism detection in 26 Gossypium species. Cluster analysis generated a phylogenetic tree with four major sub-clusters for 23 species while three species branched out individually. CAP method enhanced the rate of polymorphism of candidate gene-based markers between G. hirsutum and G. barbadense. Two hundred A-genome based SSR markers were designed after datamining of G. arboreum EST sequences (Mississippi Gossypium arboreum  EST-SSR: MGAES). Over 70% of MGAES markers successfully produced amplicons while 65 of them demonstrated polymorphism between the parents of G. hirsutum and G. barbadense RIL population and formed 14 linkage groups. Chromosomal localization of both candidate gene-based and MGAES markers was assisted by euploid and hypoaneuploid CS-B analysis. Gene-based and MGAES markers were highly informative as they were designed from candidate genes and fiber transcriptome with a potential to be integrated into the existing cotton genetic and physical maps.

  17. Characterization of a likelihood based method and effects of markers informativeness in evaluation of admixture and population group assignment

    PubMed Central

    Yang, Bao-Zhu; Zhao, Hongyu; Kranzler, Henry R; Gelernter, Joel

    2005-01-01

    Background Detection and evaluation of population stratification are crucial issues in the conduct of genetic association studies. Statistical approaches useful for understanding these issues have been proposed; these methods rely on information gained from genotyping sets of markers that reflect population ancestry. Before using these methods, a set of markers informative for differentiating population genetic substructure (PGS) is necessary. We have previously evaluated the performance of a Bayesian clustering method implemented in the software STRUCTURE in detecting PGS with a particular informative marker set. In this study, we implemented a likelihood based method (LBM) in evaluating the informativeness of the same selected marker panel, with respect to assessing potential for stratification in samples of European Americans (EAs) and African Americans (AAs), that are known to be admixed. LBM calculates the probability of a set of genotypes based on observations in a reference population with known specific allele frequencies for each marker, assuming Hardy Weinberg equilibrium (HWE) for each marker and linkage equilibrium among markers. Results In EAs, the assignment accuracy by LBM exceeded 99% using the most efficient marker FY, and reached perfect assignment accuracy using the 10 most efficient markers excluding FY. In AAs, the assignment accuracy reached 96.4% using FY, and >95% when using at least the 9 most efficient markers. The comparison of the observed and reference allele frequencies (which were derived from previous publications and public databases) shows that allele frequencies observed in EAs matched the reference group more accurately than allele frequencies observed in AAs. As a result, the LBM performed better in EAs than AAs, as might be expected given the dependence of LBMs on prior knowledge of allele frequencies. Performance was not dependent on sample size. Conclusion The performance of the LBM depends on the efficiency and number of

  18. Application of the shsp Gene, Encoding a Small Heat Shock Protein, as a Food-Grade Selection Marker for Lactic Acid Bacteria

    PubMed Central

    El Demerdash, Hassan A. M.; Heller, Knut J.; Geis, Arnold

    2003-01-01

    Plasmid pSt04 of Streptococcus thermophilus contains a gene encoding a protein with homology to small heat shock proteins (A. Geis, H. A. M. El Demerdash, and K. J. Heller, Plasmid 50:53-69, 2003). Strains cured from the shsp plasmids showed significantly reduced heat and acid resistance and a lower maximal growth temperature. Transformation of the cloned shsp gene into S. thermophilus St11 lacking a plasmid encoding shsp resulted in increased resistance to incubation at 60°C or pH 3.5 and in the ability to grow at 52°C. A food-grade cloning system for S. thermophilus, based on the plasmid-encoded shsp gene as a selection marker, was developed. This approach allowed selection after transfer of native and recombinant shsp plasmids into different S. thermophilus and Lactococcus lactis strains. Using a recombinant plasmid carrying an erythromycin resistance (Emr) gene in addition to shsp, we demonstrated that both markers are equally efficient in selecting for plasmid-bearing cells. The average transformation rates in S. thermophilus (when we were selecting for heat resistance) were determined to be 2.4 × 104 and 1.0 × 104 CFU/0.5 μg of DNA, with standard deviations of 0.54 × 104 and 0.32 × 104, for shsp and Emr selection, respectively. When we selected for pH resistance, the average transformation rates were determined to be 2.25 × 104 and 3.8 × 103 CFU/0.5 μg of DNA, with standard deviations of 0.63 × 104 and 3.48 × 103, for shsp and Emr selection, respectively. The applicability of shsp as a selection marker was further demonstrated by constructing S. thermophilus plasmid pHRM1 carrying the shsp gene as a selection marker and the restriction-modification genes of another S. thermophilus plasmid as a functional trait. PMID:12902223

  19. Accuracy of an infrared marker-based patient positioning system (ExacTrac®) for stereotactic body radiotherapy in localizing the planned isocenter using fiducial markers

    NASA Astrophysics Data System (ADS)

    Montes-Rodríguez, María de los Ángeles; Hernández-Bojórquez, Mariana; Martínez-Gómez, Alma Angélica; Contreras-Pérez, Agustín; Negrete-Hernández, Ingrid Mireya; Hernández-Oviedo, Jorge Omar; Mitsoura, Eleni; Santiago-Concha, Bernardino Gabriel

    2014-11-01

    Stereotactic Body Radiation Therapy (SBRT) requires a controlled immobilization and position monitoring of patient and target. The purpose of this work is to analyze the performance of the imaging system ExacTrac® (ETX) using infrared and fiducial markers. Materials and methods: In order to assure the accuracy of isocenter localization, a Quality Assurance procedure was applied using an infrared marker-based positioning system. Scans were acquired of an inhouse-agar gel and solid water phantom with infrared spheres. In the inner part of the phantom, three reference markers were delineated as reference and one pellet was place internally; which was assigned as the isocenter. The iPlan® RT Dose treatment planning system. Images were exported to the ETX console. Images were acquired with the ETX to check the correctness of the isocenter placement. Adjustments were made in 6D the reference markers were used to fuse the images. Couch shifts were registered. The procedure was repeated for verification purposes. Results: The data recorded of the verifications in translational and rotational movements showed averaged 3D spatial uncertainties of 0.31 ± 0.42 mm respectively 0.82° ± 0.46° in the phantom and the first correction of these uncertainties were of 1.51 ± 1.14 mm respectively and 1.37° ± 0.61°. Conclusions: This study shows a high accuracy and repeatability in positioning the selected isocenter. The ETX-system for verifying the treatment isocenter position has the ability to monitor the tracing position of interest, making it possible to be used for SBRT positioning within uncertainty ≤1mm.

  20. Accuracy of an infrared marker-based patient positioning system (ExacTrac®) for stereotactic body radiotherapy in localizing the planned isocenter using fiducial markers

    SciTech Connect

    Montes-Rodríguez, María de los Ángeles Mitsoura, Eleni; Hernández-Bojórquez, Mariana; Martínez-Gómez, Alma Angélica; Contreras-Pérez, Agustín; Negrete-Hernández, Ingrid Mireya; Hernández-Oviedo, Jorge Omar; Santiago-Concha, Bernardino Gabriel

    2014-11-07

    Stereotactic Body Radiation Therapy (SBRT) requires a controlled immobilization and position monitoring of patient and target. The purpose of this work is to analyze the performance of the imaging system ExacTrac® (ETX) using infrared and fiducial markers. Materials and methods: In order to assure the accuracy of isocenter localization, a Quality Assurance procedure was applied using an infrared marker-based positioning system. Scans were acquired of an inhouse-agar gel and solid water phantom with infrared spheres. In the inner part of the phantom, three reference markers were delineated as reference and one pellet was place internally; which was assigned as the isocenter. The iPlan® RT Dose treatment planning system. Images were exported to the ETX console. Images were acquired with the ETX to check the correctness of the isocenter placement. Adjustments were made in 6D the reference markers were used to fuse the images. Couch shifts were registered. The procedure was repeated for verification purposes. Results: The data recorded of the verifications in translational and rotational movements showed averaged 3D spatial uncertainties of 0.31 ± 0.42 mm respectively 0.82° ± 0.46° in the phantom and the first correction of these uncertainties were of 1.51 ± 1.14 mm respectively and 1.37° ± 0.61°. Conclusions: This study shows a high accuracy and repeatability in positioning the selected isocenter. The ETX-system for verifying the treatment isocenter position has the ability to monitor the tracing position of interest, making it possible to be used for SBRT positioning within uncertainty ≤1mm.

  1. Compatibility of pedigree-based and marker-based relationship matrices for single-step genetic evaluation.

    PubMed

    Christensen, Ole F

    2012-12-03

    Single-step methods provide a coherent and conceptually simple approach to incorporate genomic information into genetic evaluations. An issue with single-step methods is compatibility between the marker-based relationship matrix for genotyped animals and the pedigree-based relationship matrix. Therefore, it is necessary to adjust the marker-based relationship matrix to the pedigree-based relationship matrix. Moreover, with data from routine evaluations, this adjustment should in principle be based on both observed marker genotypes and observed phenotypes, but until now this has been overlooked. In this paper, I propose a new method to address this issue by 1) adjusting the pedigree-based relationship matrix to be compatible with the marker-based relationship matrix instead of the reverse and 2) extending the single-step genetic evaluation using a joint likelihood of observed phenotypes and observed marker genotypes. The performance of this method is then evaluated using two simulated datasets. The method derived here is a single-step method in which the marker-based relationship matrix is constructed assuming all allele frequencies equal to 0.5 and the pedigree-based relationship matrix is constructed using the unusual assumption that animals in the base population are related and inbred with a relationship coefficient γ and an inbreeding coefficient γ / 2. Taken together, this γ parameter and a parameter that scales the marker-based relationship matrix can handle the issue of compatibility between marker-based and pedigree-based relationship matrices. The full log-likelihood function used for parameter inference contains two terms. The first term is the REML-log-likelihood for the phenotypes conditional on the observed marker genotypes, whereas the second term is the log-likelihood for the observed marker genotypes. Analyses of the two simulated datasets with this new method showed that 1) the parameters involved in adjusting marker-based and pedigree-based

  2. Compatibility of pedigree-based and marker-based relationship matrices for single-step genetic evaluation

    PubMed Central

    2012-01-01

    Background Single-step methods provide a coherent and conceptually simple approach to incorporate genomic information into genetic evaluations. An issue with single-step methods is compatibility between the marker-based relationship matrix for genotyped animals and the pedigree-based relationship matrix. Therefore, it is necessary to adjust the marker-based relationship matrix to the pedigree-based relationship matrix. Moreover, with data from routine evaluations, this adjustment should in principle be based on both observed marker genotypes and observed phenotypes, but until now this has been overlooked. In this paper, I propose a new method to address this issue by 1) adjusting the pedigree-based relationship matrix to be compatible with the marker-based relationship matrix instead of the reverse and 2) extending the single-step genetic evaluation using a joint likelihood of observed phenotypes and observed marker genotypes. The performance of this method is then evaluated using two simulated datasets. Results The method derived here is a single-step method in which the marker-based relationship matrix is constructed assuming all allele frequencies equal to 0.5 and the pedigree-based relationship matrix is constructed using the unusual assumption that animals in the base population are related and inbred with a relationship coefficient γ and an inbreeding coefficient γ / 2. Taken together, this γ parameter and a parameter that scales the marker-based relationship matrix can handle the issue of compatibility between marker-based and pedigree-based relationship matrices. The full log-likelihood function used for parameter inference contains two terms. The first term is the REML-log-likelihood for the phenotypes conditional on the observed marker genotypes, whereas the second term is the log-likelihood for the observed marker genotypes. Analyses of the two simulated datasets with this new method showed that 1) the parameters involved in adjusting marker-based

  3. Breeding of a target genotype variety based on identified chalkiness marker-QTL associations in rice (Oryza sativa L.).

    PubMed

    Liu, X; Du, Y R; Li, X H; Li, X L; Yang, W Q; Wang, Y

    2015-10-21

    The aim of this study was to breed a target genotype variety based on the identified chalkiness marker-QTL (quantitative trait locus) associations in rice. First, a permanent mapping population of rice that consisted of 525 recombinant inbred lines (RILs), which were derived from Zhenshan 97/Minghui 63, was used to identify QTLs with additive effects for rice quantitative traits and percentage of grain chalkiness (PGC). Subsequently, based on the identified QTLs in rice, the molecular marker 68923-PGC was selected to screen the low chalkiness rice line. Then, using the integration of molecular marker breeding and traditional breeding, we analyzed the genotype and phenotype of inbred lines from 525 RILs; we identified one rice variety with particularly high yields, good taste, and broad adaptability. The new variety was temporarily named RIL10, which was a high quality, high yield, and broadly adaptable variety, and it is predominantly a feature that has contributed to its geographical adaptability, which would be planted from 35°E to 18°E in Chinain China, where 2/3 of rice production occurs. RIL10 was a marker-assisted selection breeding achievement for producing a high quality, high yield, and broadly adaptable rice variety.

  4. The origin of the Japanese race based on genetic markers of immunoglobulin G

    PubMed Central

    Matsumoto, Hideo

    2009-01-01

    This review addresses the distribution of genetic markers of immunoglobulin G (Gm) among 130 Mongoloid populations in the world. These markers allowed the populations to be clearly divided into 2 groups, the northern and southern groups. The northern group is characterized by high frequencies of 2 marker genes, ag and ab3st, and an extremely low frequency of the marker gene afb1b3; and the southern group, in contrast, is indicated by a remarkably high frequency of afb1b3 and low frequencies of ag and ab3st. Based on the geographical distribution of the markers and gene flow of Gm ag and ab3st (northern Mongoloid marker genes) from northeast Asia to the Japanese archipelago, the Japanese population belongs basically to the northern Mongoloid group and is thus suggested to have originated in northeast Asia, most likely in the Baikal area of Siberia. PMID:19212099

  5. Transmission/Disequilibrium Test Based on Haplotype Sharing for Tightly Linked Markers

    PubMed Central

    Zhang, Shuanglin; Sha, Qiuying; Chen, Huann-Sheng; Dong, Jianping; Jiang, Renfang

    2003-01-01

    Studies using haplotypes of multiple tightly linked markers are more informative than those using a single marker. However, studies based on multimarker haplotypes have some difficulties. First, if we consider each haplotype as an allele and use the conventional single-marker transmission/disequilibrium test (TDT), then the rapid increase in the degrees of freedom with an increasing number of markers means that the statistical power of the conventional tests will be low. Second, the parental haplotypes cannot always be unambiguously reconstructed. In the present article, we propose a haplotype-sharing TDT (HS-TDT) for linkage or association between a disease-susceptibility locus and a chromosome region in which several tightly linked markers have been typed. This method is applicable to both quantitative traits and qualitative traits. It is applicable to any size of nuclear family, with or without ambiguous phase information, and it is applicable to any number of alleles at each of the markers. The degrees of freedom (in a broad sense) of the test increase linearly as the number of markers considered increases but do not increase as the number of alleles at the markers increases. Our simulation results show that the HS-TDT has the correct type I error rate in structured populations and that, in most cases, the power of HS-TDT is higher than the power of the existing single-marker TDTs and haplotype-based TDTs. PMID:12929082

  6. Toward genetic transformation of mitochondria in mammalian cells using a recoded drug-resistant selection marker.

    PubMed

    Yoon, Young Geol; Koob, Michael Duane

    2011-04-20

    Due to technical difficulties, the genetic transformation of mitochondria in mammalian cells is still a challenge. In this report, we described our attempts to transform mammalian mitochondria with an engineered mitochondrial genome based on selection using a drug resistance gene. Because the standard drug-resistant neomycin phosphotransferase confers resistance to high concentrations of G418 when targeted to the mitochondria, we generated a recoded neomycin resistance gene that uses the mammalian mitochondrial genetic code to direct the synthesis of this protein in the mitochondria, but not in the nucleus (mitochondrial version). We also generated a universal version of the recoded neomycin resistance gene that allows synthesis of the drug-resistant proteins both in the mitochondria and nucleus. When we transfected these recoded neomycin resistance genes that were incorporated into the mouse mitochondrial genome clones into mouse tissue culture cells by electroporation, no DNA constructs were delivered into the mitochondria. We found that the universal version of the recoded neomycin resistance gene was expressed in the nucleus and thus conferred drug resistance to G418 selection, while the synthetic mitochondrial version of the gene produced no background drug-resistant cells from nuclear transformation. These recoded synthetic drug-resistant genes could be a useful tool for selecting mitochondrial genetic transformants as a precise technology for mitochondrial transformation is developed.

  7. A new type of model-based Roentgen stereophotogrammetric analysis for solving the occluded marker problem.

    PubMed

    Kaptein, B L; Valstar, E R; Stoel, B C; Rozing, P M; Reiber, J H C

    2005-11-01

    Roentgen stereophotogrammetric analysis (RSA) measures micromotion of an orthopaedic implant with respect to its surrounding bone. A problem in RSA is that the markers are sometimes overprojected by the implant itself. This study describes the so-called Marker Configuration Model-based RSA (MCM-based RSA) that is able to measure the pose of a rigid body in situations where less than three markers could be detected in both images of an RSA radiograph. MCM-based RSA is based on fitting a Marker Configuration model (MC-model) to the projection lines from the marker projection positions in the image to their corresponding Roentgen foci. An MC-model describes the positions of markers relative to each other and is obtained using conventional RSA. We used data from 15 double examinations of a clinical study of total knee prostheses and removed projections of the three tibial component markers, simulating occlusion of markers. The migration of the tibial component with respect to the bone, which should be zero, for the double examination is a measure of the accuracy of algorithm. With the new algorithm, it is possible to estimate the pose of a rigid body of which one or two markers are occluded in one of the images of the RSA radiograph with high accuracy as long as a proper MC-model of the markers in the rigid body is available. The new algorithm makes RSA more robust for occlusion of markers. This improves the results of clinical RSA studies because the number of lost RSA follow-up moments is reduced.

  8. Iterative h-minima-based marker-controlled watershed for cell nucleus segmentation.

    PubMed

    Koyuncu, Can Fahrettin; Akhan, Ece; Ersahin, Tulin; Cetin-Atalay, Rengul; Gunduz-Demir, Cigdem

    2016-04-01

    Automated microscopy imaging systems facilitate high-throughput screening in molecular cellular biology research. The first step of these systems is cell nucleus segmentation, which has a great impact on the success of the overall system. The marker-controlled watershed is a technique commonly used by the previous studies for nucleus segmentation. These studies define their markers finding regional minima on the intensity/gradient and/or distance transform maps. They typically use the h-minima transform beforehand to suppress noise on these maps. The selection of the h value is critical; unnecessarily small values do not sufficiently suppress the noise, resulting in false and oversegmented markers, and unnecessarily large ones suppress too many pixels, causing missing and undersegmented markers. Because cell nuclei show different characteristics within an image, the same h value may not work to define correct markers for all the nuclei. To address this issue, in this work, we propose a new watershed algorithm that iteratively identifies its markers, considering a set of different h values. In each iteration, the proposed algorithm defines a set of candidates using a particular h value and selects the markers from those candidates provided that they fulfill the size requirement. Working with widefield fluorescence microscopy images, our experiments reveal that the use of multiple h values in our iterative algorithm leads to better segmentation results, compared to its counterparts. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.

  9. A population 'consensus', partial linkage map of Picea abies Karst. based on RAPD markers

    Treesearch

    G. Bucci; Thomas L. Kubisiak; W.L. Nance; P. Menozzi

    1997-01-01

    The authors built a "consensus" partial linkage map based on RAPD markers using 48 sibships of eight megagametophytes each from a natural population of Norway spruce. A RAPD linkage map for a single individual from the same population had previously been constructed. Using 30 random decamers that had yielded 83 RAPD markers in the single-tree map, eight...

  10. Developement of gene and EST-based markers and their chromosomal localization in cotton

    USDA-ARS?s Scientific Manuscript database

    Molecular markers are being used extensively in genetic studies and breeding programs of cotton. To screen the limited genetic diversity in cotton germplasm, we focused our efforts to develop candidate gene and expressed sequence tag (EST) based markers and in finding polymorphisms between genotype...

  11. Development of new candidate gene and EST-based molecular markers for Gossypium species

    USDA-ARS?s Scientific Manuscript database

    New source of molecular markers accelerates the efforts in improving cotton fiber traits and aid in developing high-density integrated genetic maps. We developed new markers based on candidate genes and G. arboreum expressed sequence tag (EST) sequences, and validated them through amplification, ge...

  12. Development of marker-based tracking methods for augmented reality applied to NPP maintenance work support and its experimental evaluation

    SciTech Connect

    Ishii, H.; Fujino, H.; Bian, Z.; Sekiyama, T.; Shimoda, H.; Yoshikawa, H.

    2006-07-01

    In this study, two types of marker-based tracking methods for Augmented Reality have been developed. One is a method which employs line-shaped markers and the other is a method which employs circular-shaped markers. These two methods recognize the markers by means of image processing and calculate the relative position and orientation between the markers and the camera in real time. The line-shaped markers are suitable to be pasted in the buildings such as NPPs where many pipes and tanks exist. The circular-shaped markers are suitable for the case that there are many obstacles and it is difficult to use line-shaped markers because the obstacles hide the part of the line-shaped markers. Both methods can extend the maximum distance between the markers and the camera compared to the legacy marker-based tracking methods. (authors)

  13. A preliminary study for identification of candidate AFLP markers under artificial selection for shell color in pearl oyster Pinctada fucata.

    PubMed

    Zou, Keshu; Zhang, Dianchang; Guo, Huayang; Zhu, Caiyan; Li, Min; Jiang, Shigui

    2014-05-25

    Pearl oyster Pinctada fucata is widely cultured to produce seawater pearl in South China, and the quality of pearl is significantly affected by its shell color. Thus the Pearl Oyster Selective Breeding Program (POSBP) was carried out for the shell color and growth traits. The black (B), gold (G), red (R) and white (W) shell strains with fast growth trait were achieved after five successive generation selection. In this study, AFLP technique was used to scan genome of four strains with different shell colors to identify the candidate markers under artificial selection. Eight AFLP primer combinations were screened and yielded 688 loci, 676 (98.26%) of which were polymorphic. In black, gold, red and white strains, the percentage of polymorphic loci was 90.41%, 87.79%, 93.60% and 93.31%, respectively, Nei's gene diversity was 0.3225, 0.2829, 0.3221 and 0.3292, Shannon's information index was 0.4801, 0.4271, 0.4825 and 0.4923, and the value of FST was 0.1805. These results suggested that the four different shell color strains had high genetic diversity and great genetic differentiation among strains, which had been subjected to the continuous selective pressures during the artificial selective breeding. Furthermore, six outlier loci were considered as the candidate markers under artificial selection for shell color. This study provides a molecular evidence for the inheritance of shell color of P. fucata.

  14. The selectable antibiotic marker, tetA(C), increases Pseudomonas aeruginosa susceptibility to the herbicide/superoxide generator, paraquat.

    PubMed

    Leesukon, Panithi; Wirathorn, Wilaiwan; Chuchue, Tatsanee; Charoenlap, Nisanart; Mongkolsuk, Skorn

    2013-09-01

    The presence of the widely used selectable antibiotic marker, tetA(C), unexpectedly increased the sensitivity of Pseudomonas aeruginosa PAO1 to the superoxide-generating herbicide, paraquat. A DNA fragment spanning the first 99 amino acids of TetA(C) was sufficient to confer paraquat sensitivity. The TetA(C)-induced paraquat sensitive phenotype was observed in other Gram-negative bacteria such as Agrobacterium tumefaciens, Salmonella enterica ser. Typhimurium and Xanthomonas campestris suggesting that this is a general property of tetA(C). This finding serves as a cautionary note for those using tetA(C) as a selectable marker for genetic manipulations in studies using paraquat either as a superoxide stress generator or a redox cycling drug.

  15. Coexpression of osteogenic and adipogenic differentiation markers in selected subpopulations of primary human mesenchymal progenitor cells.

    PubMed

    Ponce, M L; Koelling, S; Kluever, A; Heinemann, D E H; Miosge, N; Wulf, G; Frosch, K-H; Schütze, N; Hufner, M; Siggelkow, H

    2008-07-01

    Knowledge of the basic mechanisms controlling osteogenesis and adipogenesis might provide new insights into the prevention of osteoporosis and age-related osteopenia. With the help of magnetic cell sorting and fluorescence activated cell sorting (FACS), osteoblastic subpopulations of mesenchymal progenitor cells were characterized. Alkaline phosphatase (AP) negative cells expressed low levels of osteoblastic and adipocytic markers. AP positive cells expressed adipocytic markers more strongly than the AP negative cell populations, thus suggesting that committed osteoblasts exhibit a greater adipogenic potential. AP negative cells differentiated to the mature osteoblastic phenotype, as demonstrated by increased AP-activity and osteocalcin secretion under standard osteogenic culture conditions. Surprisingly, this was accompanied by increased expression of adipocytic gene markers such as peroxisome proliferator-activated receptor-gamma2, lipoprotein lipase and fatty acid binding protein. The induction of adipogenic markers was suppressed by transforming growth factor-beta1 (TGF-beta1) and promoted by bone morphogenetic protein 2 (BMP-2). Osteogenic culture conditions including BMP-2 induced both the formation of mineralized nodules and cytoplasmic lipid vacuoles. Upon immunogold electron microscopic analysis, osteoblastic and adipogenic marker proteins were detectable in the same cell. Our results suggest that osteogenic and adipogenic differentiation in human mesenchymal progenitor cells might not be exclusively reciprocal, but rather, a parallel event until late during osteoblast development.

  16. Development of a cassava core collection based on single nucleotide polymorphism markers.

    PubMed

    Oliveira, E J; Ferreira, C F; Santos, V S; Oliveira, G A F

    2014-08-25

    Single nucleotide polymorphism (SNP) markers were used in the largest cassava (Manihot esculenta Crantz) germplasm collection from Brazil to develop core collections based on the maximization strategy. Subsets with 61, 64, 84, 128, 256, and 384 cassava accessions were selected and named PoHEU, MST64, PoRAN, MST128, MST256, and MST384, respectively. All the 798 alleles identified by 402 SNP markers in the entire collection were captured in all core collections. Only small alterations in the diversity parameters were observed for the different core collections compared with the complete collection. Because of the optimal adjustment of the validation parameters representative of the complete collection, the absence of genotypes with high genetic similarity and the maximization of the genetic distances between accessions of the PoHEU core collection, which contained 4.7% of the accessions of the complete collection, maximized the genetic conservation of this important cassava collection. Furthermore, the development of this core collection will allow concentrated efforts toward future characterization and agronomic evaluation of accessions to maximize the diversity and genetic gains in cassava breeding programs.

  17. Impact of reduced marker set estimation of genomic relationship matrices on genomic selection for feed efficiency in Angus cattle

    PubMed Central

    2010-01-01

    Background Molecular estimates of breeding value are expected to increase selection response due to improvements in the accuracy of selection and a reduction in generation interval, particularly for traits that are difficult or expensive to record or are measured late in life. Several statistical methods for incorporating molecular data into breeding value estimation have been proposed, however, most studies have utilized simulated data in which the generated linkage disequilibrium may not represent the targeted livestock population. A genomic relationship matrix was developed for 698 Angus steers and 1,707 Angus sires using 41,028 single nucleotide polymorphisms and breeding values were estimated using feed efficiency phenotypes (average daily feed intake, residual feed intake, and average daily gain) recorded on the steers. The number of SNPs needed to accurately estimate a genomic relationship matrix was evaluated in this population. Results Results were compared to estimates produced from pedigree-based mixed model analysis of 862 Angus steers with 34,864 identified paternal relatives but no female ancestors. Estimates of additive genetic variance and breeding value accuracies were similar for AFI and RFI using the numerator and genomic relationship matrices despite fewer animals in the genomic analysis. Bootstrap analyses indicated that 2,500-10,000 markers are required for robust estimation of genomic relationship matrices in cattle. Conclusions This research shows that breeding values and their accuracies may be estimated for commercially important sires for traits recorded in experimental populations without the need for pedigree data to establish identity by descent between members of the commercial and experimental populations when at least 2,500 SNPs are available for the generation of a genomic relationship matrix. PMID:20403185

  18. Addition of chromic oxide to creep feed as a fecal marker for selection of creep feed-eating suckling pigs.

    PubMed

    Kuller, Wikke I; van Beers-Schreurs, Hetty M G; Soede, Nicoline M; Taverne, Marcel A M; Kemp, Bas; Verheijden, Jos H M

    2007-07-01

    To determine whether the addition of chromic oxide (Cr(2)O(3)) to creep feed could be used as a visual marker in feces for selection of creep feed-eating suckling pigs. 20 suckling pigs. Via syringe, 5 pigs (2 to 3 days old on day 0; 1 pig/treatment) from each of 4 litters received oral administrations of 10, 20, 30, or 40 g of creep feed containing 10 g of Cr(2)O(3)*kg(1) on each of 2 consecutive days (days 20 and 21) or 30 g of creep feed containing 10 g of Cr(2)O(3)*kg(1) on day 20 and 30 g of Cr(2)O(3)-free creep feed on day 21. On days 21 through 24, 6 fecal samples were collected from each pig at regular intervals between 8:00 AM and 6:00 PM. Green-colored feces were considered indicative of creep feed consumption (eaters). Data analyses were based on single and multiple fecal samples. On day 22, evaluation of 1 fecal sample/pig and multiple fecal samples per pig resulted in identification of as many as 40% and only 15% of the feed-treated pigs wrongly as noneaters, respectively. Repeated sampling over multiple days would identify 99% of eaters accurately. Pigs erroneously identified as noneaters were those administered either low amounts of Cr(2)O(3)-supplemented creep feed for 2 days or Cr(2)O(3)-supplemented creep feed on only 1 day. Data suggest that addition of Cr(2)O(3) to creep feed enables selection of individual creep feed-eating suckling pigs via examination of feces, provided that repeated fecal samples are evaluated.

  19. Development of SSR markers for a Tibetan medicinal plant, Lancea tibetica (Phrymaceae), based on RAD sequencing1

    PubMed Central

    Tian, Zunzhe; Zhang, Faqi; Liu, Hairui; Gao, Qingbo; Chen, Shilong

    2016-01-01

    Premise of the study: Lancea tibetica (Phrymaceae), a Tibetan medicinal plant, is endemic to the Qinghai–Tibet Plateau. The over-exploitation of wild L. tibetica has led to the destruction of many populations. To enhance protection and management, biological research, especially population genetic studies, should be carried out on L. tibetica. Simple sequence repeat (SSR) markers of L. tibetica were developed to analyze population diversity. Methods and Results: Four thousand four hundred and forty-one SSR loci were identified for L. tibetica based on restriction-site associated DNA (RAD) sequencing on the Illumina HiSeq platform. One hundred SSR loci were arbitrarily selected for primer design, and 38 of them were successfully amplified. These markers were tested on 56 individuals from three populations of L. tibetica, and 10 markers displayed polymorphisms. The total number of alleles per locus ranged from three to eight, and observed and expected heterozygosities ranged from 0.200 to 1.000 and 0.683 to 0.879, respectively. We tested for cross-amplification of these 10 markers in the related species L. hirsuta and found that nine could be successfully amplified. Conclusions: The SSR markers characterized here are the first to be developed and tested in L. tibetica. They will be useful for future population genetic studies on L. tibetica and closely related species. PMID:27843726

  20. Reference ranges and impact of selected confounders on classic serum and urinary renal markers in neonatal period.

    PubMed

    Miklaszewska, Monika; Korohoda, Przemysław; Kwinta, Przemko; Zachwieja, Katarzyna; Tomasik, Tomasz; Kiliś-Pstrusińska, Katarzyna; Drożdż, Dorota

    2017-03-01

    Renal injury in term and pre-term neonates may be an antecedent to chronic kidney disease in the child's further life. The aim of the paper was to compile a reference range for selected serum and urinary renal markers in the neonatal period for full-term (FT) and pre-term (PT) newborns. The prospective study included 23 FT infants (birth weight - BW≥2500g and gestational age - GA≥37Hbd) and 32 PT children (BW<2500g and GA<37Hbd) in good general condition, without acute kidney injury (AKI) or sepsis. Between the 4th and 28th DOL, urinary concentrations of the studied renal markers (uCr, uNa, uOsm) were determined on a daily basis, while serum creatinine (SCr) was assessed minimum every 48-72h. The mean GA and BW of the FT and PT infants were respectively as follows: 38.5±1.7Hbd; 3433±495.2g and 32.7±2.6Hbd; 1836.7±419.8g. For serum glomerular (SCr, eGFR) and tubular markers (FENa, RFI), the median values with normal ranges were compiled. For urinary renal markers (uCr, uNa, uOsm) and those values standardized for kg of body weight, percentile tables for 4-28DOL were elaborated. The study has resulted in determining the normal ranges of serum glomerular and tubular renal markers, as well as percentile tables of selected urinary renal parameters during the neonatal period. The percentile tables may prove to be helpful for further standardization of other urinary parameters per urinary creatinine in neonatal population. Copyright © 2017 Medical University of Bialystok. Published by Elsevier B.V. All rights reserved.

  1. Homoeologous GSL-ELONG gene replacement for manipulation of aliphatic glucosinolates in Brassica rapa L. by marker assisted selection

    PubMed Central

    Hirani, Arvind H.; Zelmer, Carla D.; McVetty, Peter B. E.; Daayf, Fouad; Li, Genyi

    2013-01-01

    Aliphatic glucosinolates are the predominant sulfur-rich plant secondary metabolites in economically important Brassica crops. Glucosinolates and their hydrolysis products are involved in plant–microbe, plant–insect, plant–animal, and plant–human interactions. It is, therefore, important to manipulate glucosinolate profiles and contents in Brassica species. In this study, aliphatic glucosinolates were genetically manipulated through homoeologous recombination in backcross lines followed by marker assisted selection in B. rapa. A resynthesized B. napus line, from a cross between B. rapa and B. oleracea, was backcrossed with Chinese cabbage doubled haploid line, RI16. Marker assisted selection for non-functional gene was performed in each backcross generations. Advanced backcross progenies (BC3F2) were developed to identify homoeologous gene replacement and/or introgression. Reduction in 5C aliphatic glucosinolates (gluconapoleiferin, glucoalyssin, and glucobrassicanapin) was observed in BC3F2 progenies of the recurrent parent that carried the GSL-ELONG- gene. The GSL-ELONG- positive backcross progenies were also screened by the A-genome and BraGSL-ELONG gene specific marker, which linked with 5C aliphatic glucosinolates. The A-genome specific marker was absent in the plants of advanced backcross progenies which showed reduction in 5C aliphatic glucosinolates. The results suggest that the functional allele had been replaced by the non-functional GSL-ELONG- allele from B. oleracea. Some advanced backcross progenies (BC3F2) positive for the GSL-ELONG- allele and the A-genome specific SCAR marker BraMAM1-1 did not show reduction in 5C aliphatic glucosinolates, suggesting that GSL-ELONG- allele is recessive. Replacement of the functional locus in the A-genome by non-functional counterpart in the C-genome reduced the content of 5C aliphatic glucosinolates in B. rapa seeds with 20 μmol/g. PMID:23532458

  2. Genetic linkage map of Chinese native variety faba bean (Vicia faba L.) based on simple sequence repeat markers

    USDA-ARS?s Scientific Manuscript database

    Simple sequence repeat (SSR) marker is a powerful tool for construction of genetic linkage map which can be applied for locating quantitative trait loci (QTL) and marker-assisted selection (MAS). In this study, a genetic map of faba bean was constructed with SSR markers using a population of 129 F2 ...

  3. Construction of intersubspecific molecular genetic map of lentil based on ISSR, RAPD and SSR markers.

    PubMed

    Gupta, Mamta; Verma, Bhawna; Kumar, Naresh; Chahota, Rakesh K; Rathour, Rajeev; Sharma, Shyam K; Bhatia, Sabhyata; Sharma, Tilak R

    2012-01-01

    Lentil (Lens culinaris ssp. culinaris), is a self-pollinating diploid (2n = 2x = 14), cool-season legume crop and is consumed worldwide as a rich source of protein (~24.0%), largely in vegetarian diets. Here we report development of a genetic linkage map of Lens using 114 F(2) plants derived from the intersubspecific cross between L 830 and ILWL 77. RAPD (random amplified polymorphic DNA) primers revealed more polymorphism than ISSR (intersimple sequence repeat) and SSR (simple sequence repeat) markers. The highest proportion (30.72%) of segregation distortion was observed in RAPD markers. Of the 235 markers (34 SSR, 9 ISSR and 192 RAPD) used in the mapping study, 199 (28 SSRs, 9 ISSRs and 162 RAPDs) were mapped into 11 linkage groups (LGs), varying between 17.3 and 433.8 cM and covering 3843.4 cM, with an average marker spacing of 19.3 cM. Linkage analysis revealed nine major groups with 15 or more markers each and two small LGs with two markers each, and 36 unlinked markers. The study reported assigning of 11 new SSRs on the linkage map. Of the 66 markers with aberrant segregation, 14 were unlinked and the remaining 52 were mapped. ISSR and RAPD markers were found to be useful in map construction and saturation. The current map represents maximum coverage of lentil genome and could be used for identification of QTL regions linked to agronomic traits, and for marker-assisted selection in lentil.

  4. Validation and Implementation of Marker-Assisted Selection (MAS) for PVY Resistance (Ryadg gene) in a Tetraploid Potato Breeding Program

    USDA-ARS?s Scientific Manuscript database

    The gene Ryadg from S. tuberosum ssp. andigena provides extreme resistance to PVY. This gene has been mapped to chromosome XI and linked PCR-based DNA markers have been identified. Advanced tetraploid russeted potato clones developed by the U.S. Pacific Northwest Potato Breeding Program with Ryadg P...

  5. Finding linked markers to En for efficient selection of pea enation Mosaic Virus resistance in pea

    USDA-ARS?s Scientific Manuscript database

    Pea enation mosaic virus (PEMV) causes an important disease of cool season food legumes, resulting in significant yield loss worldwide. The present investigation was carried out to study the inheritance and identify the molecular markers linked with the PEMV resistance gene (En) in field pea (Pisum ...

  6. Selection for Run1-Ren1 dihybrid grapevines using microsatellite markers

    USDA-ARS?s Scientific Manuscript database

    The inheritance of Ren1 and Run1 powdery mildew resistance genes were tracked in a grapevine hybrid family using linked microsatellite markers. Segregation of the powdery mildew resistance phenotype was evaluated under in vitro and greenhouse conditions independently of the genotype data. Combined a...

  7. Apramycin resistance as a selective marker for gene transfer in mycobacteria.

    PubMed Central

    Paget, E; Davies, J

    1996-01-01

    We have explored the potential of using the apramycin resistance gene as a marker in mycobacterial gene transfer studies. Shuttle plasmids available for both electroporation and conjugation studies have been constructed, and we have successfully validated the use of the apramycin resistance gene as a component of cloning vectors for Mycobacterium smegmatis, M. bovis BCG, and M. tuberculosis. PMID:8892841

  8. Marker-assisted selection to pyramid nematode resistance and the high oleic trait in peanut

    USDA-ARS?s Scientific Manuscript database

    The dynamic challenges of peanut farming demand a quick response from breeders to develop new cultivars that can cope with new problems. Application of molecular markers in peanut breeding programs can greatly compress the cultivar development process from ten to fifteen years to less than three yea...

  9. Marker-based quantification of interfractional tumor position variation and the use of markers for setup verification in radiation therapy for esophageal cancer.

    PubMed

    Jin, Peng; van der Horst, Astrid; de Jong, Rianne; van Hooft, Jeanin E; Kamphuis, Martijn; van Wieringen, Niek; Machiels, Melanie; Bel, Arjan; Hulshof, Maarten C C M; Alderliesten, Tanja

    2015-12-01

    The aim of this study was to quantify interfractional esophageal tumor position variation using markers and investigate the use of markers for setup verification. Sixty-five markers placed in the tumor volumes of 24 esophageal cancer patients were identified in computed tomography (CT) and follow-up cone-beam CT. For each patient we calculated pairwise distances between markers over time to evaluate geometric tumor volume variation. We then quantified marker displacements relative to bony anatomy and estimated the variation of systematic (Σ) and random errors (σ). During bony anatomy-based setup verification, we visually inspected whether the markers were inside the planning target volume (PTV) and attempted marker-based registration. Minor time trends with substantial fluctuations in pairwise distances implied tissue deformation. Overall, Σ(σ) in the left-right/cranial-caudal/anterior-posterior direction was 2.9(2.4)/4.1(2.4)/2.2(1.8) mm; for the proximal stomach, it was 5.4(4.3)/4.9(3.2)/1.9(2.4) mm. After bony anatomy-based setup correction, all markers were inside the PTV. However, due to large tissue deformation, marker-based registration was not feasible. Generally, the interfractional position variation of esophageal tumors is more pronounced in the cranial-caudal direction and in the proximal stomach. Currently, marker-based setup verification is not feasible for clinical routine use, but markers can facilitate the setup verification by inspecting whether the PTV covers the tumor volume adequately. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  10. Gene targeting in the red alga Cyanidioschyzon merolae: single- and multi-copy insertion using authentic and chimeric selection markers.

    PubMed

    Fujiwara, Takayuki; Ohnuma, Mio; Yoshida, Masaki; Kuroiwa, Tsuneyoshi; Hirano, Tatsuya

    2013-01-01

    The unicellular red alga Cyanidioschyzon merolae is an emerging model organism for studying organelle division and inheritance: the cell is composed of an extremely simple set of organelles (one nucleus, one mitochondrion and one chloroplast), and their genomes are completely sequenced. Although a fruitful set of cytological and biochemical methods have now been developed, gene targeting techniques remain to be fully established in this organism. Thus far, only a single selection marker, URA Cm-Gs , has been available that complements the uracil-auxotrophic mutant M4. URA Cm-Gs , a chimeric URA5.3 gene of C. merolae and the related alga Galdieria sulphuraria, was originally designed to avoid gene conversion of the mutated URA5.3 allele in the parental strain M4. Although an early example of targeted gene disruption by homologous recombination was reported using this marker, the genome structure of the resultant transformants had never been fully characterized. In the current study, we showed that the use of the chimeric URA Cm-Gs selection marker caused multicopy insertion at high frequencies, accompanied by undesired recombination events at the targeted loci. The copy number of the inserted fragments was variable among the transformants, resulting in high yet uneven levels of transgene expression. In striking contrast, when the authentic URA5.3 gene (URA Cm-Cm ) was used as a selection marker, efficient single-copy insertion was observed at the targeted locus. Thus, we have successfully established a highly reliable and reproducible method for gene targeting in C. merolae. Our method will be applicable to a number of genetic manipulations in this organism, including targeted gene disruption, replacement and tagging.

  11. Gene Targeting in the Red Alga Cyanidioschyzon merolae: Single- and Multi-Copy Insertion Using Authentic and Chimeric Selection Markers

    PubMed Central

    Fujiwara, Takayuki; Ohnuma, Mio; Yoshida, Masaki; Kuroiwa, Tsuneyoshi; Hirano, Tatsuya

    2013-01-01

    The unicellular red alga Cyanidioschyzon merolae is an emerging model organism for studying organelle division and inheritance: the cell is composed of an extremely simple set of organelles (one nucleus, one mitochondrion and one chloroplast), and their genomes are completely sequenced. Although a fruitful set of cytological and biochemical methods have now been developed, gene targeting techniques remain to be fully established in this organism. Thus far, only a single selection marker, URACm-Gs, has been available that complements the uracil-auxotrophic mutant M4. URACm-Gs, a chimeric URA5.3 gene of C. merolae and the related alga Galdieria sulphuraria, was originally designed to avoid gene conversion of the mutated URA5.3 allele in the parental strain M4. Although an early example of targeted gene disruption by homologous recombination was reported using this marker, the genome structure of the resultant transformants had never been fully characterized. In the current study, we showed that the use of the chimeric URACm-Gs selection marker caused multicopy insertion at high frequencies, accompanied by undesired recombination events at the targeted loci. The copy number of the inserted fragments was variable among the transformants, resulting in high yet uneven levels of transgene expression. In striking contrast, when the authentic URA5.3 gene (URACm-Cm) was used as a selection marker, efficient single-copy insertion was observed at the targeted locus. Thus, we have successfully established a highly reliable and reproducible method for gene targeting in C. merolae. Our method will be applicable to a number of genetic manipulations in this organism, including targeted gene disruption, replacement and tagging. PMID:24039997

  12. Cytogenetic Analysis of Populus trichocarpa - Ribosomal DNA, Telomere Repeat Sequence, and Marker-selected BACs

    SciTech Connect

    Tuskan, Gerald A; Gunter, Lee E; DiFazio, Stephen P

    2009-01-01

    The 18S-28S rDNA and 5S rDNA loci in Populus trichocarpa were localized using fluorescent in situ hybridization (FISH). Two 18S-28S rDNA sites and one 5S rDNA site were identified and located at the ends of 3 different chromosomes. FISH signals from the Arabidopsis -type telomere repeat sequence were observed at the distal ends of each chromosome. Six BAC clones selected from 2 linkage groups based on genome sequence assembly (LG-I and LG-VI) were localized on 2 chromosomes, as expected. BACs from LG-I hybridized to the longest chromosome in the complement. All BAC positions were found to be concordant with sequence assembly positions. BAC-FISH will be useful for delineating each of the Populus trichocarpa chromosomes and improving the sequence assembly of this model angiosperm tree species.

  13. Cytogenetic analysis of Populus trichocarpa--ribosomal DNA, telomere repeat sequence, and marker-selected BACs.

    PubMed

    Islam-Faridi, M N; Nelson, C D; DiFazio, S P; Gunter, L E; Tuskan, G A

    2009-01-01

    The 18S-28S rDNA and 5S rDNA loci in Populus trichocarpa were localized using fluorescent in situ hybridization (FISH). Two 18S-28S rDNA sites and one 5S rDNA site were identified and located at the ends of 3 different chromosomes. FISH signals from the Arabidopsis-type telomere repeat sequence were observed at the distal ends of each chromosome. Six BAC clones selected from 2 linkage groups based on genome sequence assembly (LG-I and LG-VI) were localized on 2 chromosomes, as expected. BACs from LG-I hybridized to the longest chromosome in the complement. All BAC positions were found to be concordant with sequence assembly positions. BAC-FISH will be useful for delineating each of the Populus trichocarpa chromosomes and improving the sequence assembly of this model angiosperm tree species.

  14. Tissue-specifically regulated site-specific excision of selectable marker genes in bivalent insecticidal, genetically-modified rice.

    PubMed

    Hu, Zhan; Ding, Xuezhi; Hu, Shengbiao; Sun, Yunjun; Xia, Liqiu

    2013-12-01

    Marker-free, genetically-modified rice was created by the tissue-specifically regulated Cre/loxP system, in which the Cre recombinase gene and hygromycin phosphotransferase gene (hpt) were flanked by two directly oriented loxP sites. Cre expression was activated by the tissue-specific promoter OsMADS45 in flower or napin in seed, resulting in simultaneous excision of the recombinase and marker genes. Segregation of T1 progeny was performed to select recombined plants. The excision was confirmed by PCR, Southern blot and sequence analyses indicating that efficiency varied from 10 to 53 % for OsMADS45 and from 12 to 36 % for napin. The expression of cry1Ac and vip3A was detected by RT-PCR analysis in marker-free transgenic rice. These results suggested that our tissue-specifically regulated Cre/loxP system could auto-excise marker genes from transgenic rice and alleviate public concerns about the security of GM crops.

  15. Establishment of apoptotic regulatory network for genetic markers of colorectal cancer and optimal selection of traditional Chinese medicine target.

    PubMed

    Tian, Tongde; Chen, Chuanliang; Yang, Feng; Tang, Jingwen; Pei, Junwen; Shi, Bian; Zhang, Ning; Zhang, Jianhua

    2017-03-01

    The paper aimed to screen out genetic markers applicable to early diagnosis for colorectal cancer and establish apoptotic regulatory network model for colorectal cancer, and to analyze the current situation of traditional Chinese medicine (TCM) target, thereby providing theoretical evidence for early diagnosis and targeted therapy of colorectal cancer. Taking databases including CNKI, VIP, Wanfang data, Pub Med, and MEDLINE as main sources of literature retrieval, literatures associated with genetic markers that are applied to early diagnosis of colorectal cancer were searched and performed comprehensive and quantitative analysis by Meta analysis, hence screening genetic markers used in early diagnosis of colorectal cancer. KEGG analysis was employed to establish apoptotic regulatory network model based on screened genetic markers, and optimization was conducted on TCM targets. Through Meta analysis, seven genetic markers were screened out, including WWOX, K-ras, COX-2, P53, APC, DCC and PTEN, among which DCC has the highest diagnostic efficiency. Apoptotic regulatory network was built by KEGG analysis. Currently, it was reported that TCM has regulatory function on gene locus in apoptotic regulatory network. The apoptotic regulatory model of colorectal cancer established in this study provides theoretical evidence for early diagnosis and TCM targeted therapy of colorectal cancer in clinic.

  16. The linkage maps of Dendrobium species based on RAPD and SRAP markers.

    PubMed

    Xue, Dawei; Feng, Shangguo; Zhao, Hongyan; Jiang, Hua; Shen, Bo; Shi, Nongnong; Lu, Jiangjie; Liu, Junjun; Wang, Huizhong

    2010-03-01

    Dendrobium plants are used commonly as tonic herbs and health food in many Asian countries, especially in China. Here we report the genetic map construction of two Dendrobium species with a double pseudo-testcross strategy using random amplified polymorphic DNA (RAPD) and sequence-related amplified polymorphism (SRAP) markers. A F(1) mapping population of 90 individuals was developed from a cross between D. officinale and D. hercoglossum. A total of 307 markers, including 209 RAPD and 98 SRAP, were identified and used for genetic linkage group (LG) analysis. The D. officinale linkage map consisted of 11 major linkage groups and 3 doublets, which covered 629.4 cM by a total of 62 markers with an average locus distance of 11.2 cM between two adjacent markers. The D. hercoglossum linkage map contained 112 markers mapped on 15 major and 4 minor linkage groups, spanning a total length of 1,304.6 cM with an average distance of 11.6 cM between two adjacent markers. The maps constructed in this study covered 92.7% and 82.7% of the D. hercoglossum and D. officinale genomes respectively, providing an important basis for the mapping of horticultural and medicinal traits and for the application of marker-assisted selection in Dendrobium breeding program.

  17. Development of Selectable Marker-Free Transgenic Rice Plants with Enhanced Seed Tocopherol Content through FLP/FRT-Mediated Spontaneous Auto-Excision

    PubMed Central

    Woo, Hee-Jong; Qin, Yang; Park, Soo-Yun; Park, Soon Ki; Cho, Yong-Gu; Shin, Kong-Sik; Lim, Myung-Ho; Cho, Hyun-Suk

    2015-01-01

    Development of marker-free transgenic plants is a technical alternative for avoiding concerns about the safety of selectable marker genes used in genetically modified (GM) crops. Here, we describe the construction of a spontaneous self-excision binary vector using an oxidative stress-inducible modified FLP/FRT system and its successful application to produce marker-free transgenic rice plants with enhanced seed tocopherol content. To generate selectable marker-free transgenic rice plants, we constructed a binary vector using the hpt selectable marker gene and the rice codon-optimized FLP (mFLP) gene under the control of an oxidative stress-inducible promoter between two FRT sites, along with multiple cloning sites for convenient cloning of genes of interest. Using this pCMF binary vector with the NtTC gene, marker-free T1 transgenic rice plants expressing NtTC were produced by Agrobacterium-mediated stable transformation using hygromycin as a selective agent, followed by segregation of selectable marker genes. Furthermore, α-, γ-, and total tocopherol levels were significantly increased in seeds of the marker-free transgenic TC line compared with those of wild-type plants. Thus, this spontaneous auto-excision system, incorporating an oxidative stress-inducible mFLP/FRT system to eliminate the selectable marker gene, can be easily adopted and used to efficiently generate marker-free transgenic rice plants. Moreover, nutritional enhancement of rice seeds through elevation of tocopherol content coupled with this marker-free strategy may improve human health and public acceptance of GM rice. PMID:26172549

  18. Genetic Variation and Breeding Signature in Mass Selection Lines of the Pacific Oyster (Crassostrea gigas) Assessed by SNP Markers

    PubMed Central

    Zhong, Xiaoxiao; Feng, Dandan; Yu, Hong; Kong, Lingfeng; Li, Qi

    2016-01-01

    In breeding industries, a challenging problem is how to keep genetic diversity over generations. To investigate genetic variation and identify breeding signatures in mass selected lines of Pacific oyster (Crassostrea gigas), three sixth-generation selected lines and four wild populations were assessed using 103 single nucleotide polymorphism (SNP) markers. The genetic diversity data indicated that the selected lines exhibited a significant reduction in the observed heterozygosity and observed number of alleles per locus compared with the wild populations (P≤0.05), indicating the selected lines tended to lose genetic diversity contrasted with the wild populations. The unweighted pair-group method with arithmetic mean (UPGMA) analysis showed that the wild populations and selected lines were not separated into two groups. Using four outlier tests, a total of 17 loci were found under selection at two levels. The global outlier detection suggested that 4 common outlier loci were subject to selection using both the hierarchical island model and Bayesian likelihood approaches. At regional level, 3 SNPs were detected as outlier using at least two outlier tests and one outlier SNP (CgSNP309) was overlapped in the two wild-selected population comparisons. The candidate outlier SNPs provide valuable resources for future association studies in C. gigas. PMID:26954577

  19. A mutated cytosine deaminase gene, codA (D314A), as an efficient negative selection marker for gene targeting in rice.

    PubMed

    Osakabe, Keishi; Nishizawa-Yokoi, Ayako; Ohtsuki, Namie; Osakabe, Yuriko; Toki, Seiichi

    2014-03-01

    Gene targeting (GT) is a powerful tool manipulating a gene of interest in a given genome specifically and precisely. To achieve efficient GT in higher plants, both positive and negative selection markers are required. In particular, a strong negative selection system is needed for enrichment of cells to eliminate those cells in which random integration of the introduced DNA has occurred in GT experiments. Currently, non-conditional negative selection marker genes are used for GT experiments in rice plants, and no conditional negative selection system is available. In this study, we describe the development of an efficient conditional negative selection system in rice plants using Escherichia coli cytosine deaminase (codA). We found that a mutant codA gene, codA(D314A), acts more efficiently than the wild-type codA for negative selection in rice plants. The codA(D314A) marker was further used as a negative selection marker for GT experiments in rice. Our conditional negative selection system effectively eliminated the cells in which random integration event(s) occurred; the enrichment factor was approximately 100-fold. This enrichment factor was similar to that found when Corynebacterium diphtheriae toxin fragment A was used. Our results suggest the codA(D314A) marker gene as a promising negative selection marker for GT of rice.

  20. Genomic selection and association mapping in rice (Oryza sativa): effect of trait genetic architecture, training population composition, marker number and statistical model on accuracy of rice genomic selection in elite, tropical rice breeding lines.

    PubMed

    Spindel, Jennifer; Begum, Hasina; Akdemir, Deniz; Virk, Parminder; Collard, Bertrand; Redoña, Edilberto; Atlin, Gary; Jannink, Jean-Luc; McCouch, Susan R

    2015-02-01

    Genomic Selection (GS) is a new breeding method in which genome-wide markers are used to predict the breeding value of individuals in a breeding population. GS has been shown to improve breeding efficiency in dairy cattle and several crop plant species, and here we evaluate for the first time its efficacy for breeding inbred lines of rice. We performed a genome-wide association study (GWAS) in conjunction with five-fold GS cross-validation on a population of 363 elite breeding lines from the International Rice Research Institute's (IRRI) irrigated rice breeding program and herein report the GS results. The population was genotyped with 73,147 markers using genotyping-by-sequencing. The training population, statistical method used to build the GS model, number of markers, and trait were varied to determine their effect on prediction accuracy. For all three traits, genomic prediction models outperformed prediction based on pedigree records alone. Prediction accuracies ranged from 0.31 and 0.34 for grain yield and plant height to 0.63 for flowering time. Analyses using subsets of the full marker set suggest that using one marker every 0.2 cM is sufficient for genomic selection in this collection of rice breeding materials. RR-BLUP was the best performing statistical method for grain yield where no large effect QTL were detected by GWAS, while for flowering time, where a single very large effect QTL was detected, the non-GS multiple linear regression method outperformed GS models. For plant height, in which four mid-sized QTL were identified by GWAS, random forest produced the most consistently accurate GS models. Our results suggest that GS, informed by GWAS interpretations of genetic architecture and population structure, could become an effective tool for increasing the efficiency of rice breeding as the costs of genotyping continue to decline.

  1. Genomic Selection and Association Mapping in Rice (Oryza sativa): Effect of Trait Genetic Architecture, Training Population Composition, Marker Number and Statistical Model on Accuracy of Rice Genomic Selection in Elite, Tropical Rice Breeding Lines

    PubMed Central

    Spindel, Jennifer; Begum, Hasina; Akdemir, Deniz; Virk, Parminder; Collard, Bertrand; Redoña, Edilberto; Atlin, Gary; Jannink, Jean-Luc; McCouch, Susan R.

    2015-01-01

    Genomic Selection (GS) is a new breeding method in which genome-wide markers are used to predict the breeding value of individuals in a breeding population. GS has been shown to improve breeding efficiency in dairy cattle and several crop plant species, and here we evaluate for the first time its efficacy for breeding inbred lines of rice. We performed a genome-wide association study (GWAS) in conjunction with five-fold GS cross-validation on a population of 363 elite breeding lines from the International Rice Research Institute's (IRRI) irrigated rice breeding program and herein report the GS results. The population was genotyped with 73,147 markers using genotyping-by-sequencing. The training population, statistical method used to build the GS model, number of markers, and trait were varied to determine their effect on prediction accuracy. For all three traits, genomic prediction models outperformed prediction based on pedigree records alone. Prediction accuracies ranged from 0.31 and 0.34 for grain yield and plant height to 0.63 for flowering time. Analyses using subsets of the full marker set suggest that using one marker every 0.2 cM is sufficient for genomic selection in this collection of rice breeding materials. RR-BLUP was the best performing statistical method for grain yield where no large effect QTL were detected by GWAS, while for flowering time, where a single very large effect QTL was detected, the non-GS multiple linear regression method outperformed GS models. For plant height, in which four mid-sized QTL were identified by GWAS, random forest produced the most consistently accurate GS models. Our results suggest that GS, informed by GWAS interpretations of genetic architecture and population structure, could become an effective tool for increasing the efficiency of rice breeding as the costs of genotyping continue to decline. PMID:25689273

  2. Development of PCR-Based Markers to Determine the Sex of Kelps.

    PubMed

    Lipinska, Agnieszka P; Ahmed, Sophia; Peters, Akira F; Faugeron, Sylvain; Cock, J Mark; Coelho, Susana M

    2015-01-01

    Sex discriminating genetic markers are commonly used to facilitate breeding programs in economically and ecologically important animal and plant species. However, despite their considerable economic and ecological value, the development of sex markers for kelp species has been very limited. In this study, we used the recently described sequence of the sex determining region (SDR) of the brown algal model Ectocarpus to develop novel DNA-based sex-markers for three commercially relevant kelps: Laminaria digitata, Undaria pinnatifida and Macrocystis pyrifera. Markers were designed within nine protein coding genes of Ectocarpus male and female (U/V) sex chromosomes and tested on gametophytes of the three kelp species. Seven primer pairs corresponding to three loci in the Ectocarpus SDR amplified sex-specific bands in the three kelp species, yielding at least one male and one female marker for each species. Our work has generated the first male sex-specific markers for L. digitata and U. pinnatifida, as well as the first sex markers developed for the genus Macrocystis. The markers and methodology presented here will not only facilitate seaweed breeding programs but also represent useful tools for population and demography studies and provide a means to investigate the evolution of sex determination across this largely understudied eukaryotic group.

  3. Development of PCR‐Based Markers to Determine the Sex of Kelps

    PubMed Central

    Lipinska, Agnieszka P.; Ahmed, Sophia; Peters, Akira F.; Faugeron, Sylvain; Cock, J. Mark; Coelho, Susana M.

    2015-01-01

    Sex discriminating genetic markers are commonly used to facilitate breeding programs in economically and ecologically important animal and plant species. However, despite their considerable economic and ecological value, the development of sex markers for kelp species has been very limited. In this study, we used the recently described sequence of the sex determining region (SDR) of the brown algal model Ectocarpus to develop novel DNA-based sex-markers for three commercially relevant kelps: Laminaria digitata, Undaria pinnatifida and Macrocystis pyrifera. Markers were designed within nine protein coding genes of Ectocarpus male and female (U/V) sex chromosomes and tested on gametophytes of the three kelp species. Seven primer pairs corresponding to three loci in the Ectocarpus SDR amplified sex-specific bands in the three kelp species, yielding at least one male and one female marker for each species. Our work has generated the first male sex-specific markers for L. digitata and U. pinnatifida, as well as the first sex markers developed for the genus Macrocystis. The markers and methodology presented here will not only facilitate seaweed breeding programs but also represent useful tools for population and demography studies and provide a means to investigate the evolution of sex determination across this largely understudied eukaryotic group. PMID:26496392

  4. Quasar Selection Based On Photometric Variability

    NASA Astrophysics Data System (ADS)

    MacLeod, Chelsea; Brooks, K.; Ivezic, Z.; Kochanek, C. S.; Gibson, R.; Meisner, A.; Kozlowski, S.; Sesar, B.; Becker, A. C.; deVries, W. H.

    2011-01-01

    We develop a method for separating quasars from other variable point sources using SDSS Stripe 82 light curve data for 10,000 variable objects. To statistically describe quasar variability, we use a damped random walk model parametrized by a damping time scale, tau, and an asymptotic amplitude (structure function), SF_inf. With the aid of an SDSS spectroscopically confirmed quasar sample, we demonstrate that variability selection in typical extragalactic fields with low stellar density can deliver complete samples with reasonable purity (or efficiency, E). Compared to a selection method based solely on the slope of the structure function, the inclusion of the tau information boosts E from 60% to 75% while maintaining a highly complete sample (98%) even in the absence of color information. With the aid of color selection, the purity can be further boosted to 96%, with C=93%. Hence, selection methods based on variability will play an important role in the selection of quasars with data provided by upcoming large sky surveys, such as the Large Synoptic Survey Telescope (LSST). For a typical (simulated) LSST cadence over 10 years and a photometric accuracy of 0.03 mag (achieved at i 22), C is expected to be 88% for a simple sample selection criterion of tau>100 days. In summary, given an adequate survey cadence, photometric variability provides an even better method than color selection for separating quasars from stars. We acknowledge support by NSF grant AST-0807500 to the University of Washington.

  5. Identification of uterine leiomyoma-specific marker genes based on DNA methylation and their clinical application

    PubMed Central

    Sato, Shun; Maekawa, Ryo; Yamagata, Yoshiaki; Tamura, Isao; Lee, Lifa; Okada, Maki; Jozaki, Kosuke; Asada, Hiromi; Tamura, Hiroshi; Sugino, Norihiro

    2016-01-01

    Differential diagnosis of uterine leiomyomas and leiomyosarcomas is needed to determine whether the uterus can be retained. Therefore, biomarkers for uterine leiomyomas, and reliable and objective diagnostic methods have been desired besides the pathological diagnosis. In the present study, we identified 12 genes specific to uterine leiomyomas based on DNA methylation. Using these marker genes specific to uterine leiomyomas, we established a hierarchical clustering system based on the DNA methylation level of the marker genes, which could completely differentiate between uterine leiomyomas and normal myometrium. Furthermore, our hierarchical clustering system completely discriminated uterine cancers and differentiated between uterine leiomyosarcomas and leiomyomas with more than 70% accuracy. In conclusion, this study identified DNA methylation-based marker genes specific to uterine leiomyomas, and our hierarchical clustering system using these marker genes was useful for differential diagnosis of uterine leiomyomas and leiomyosarcomas. PMID:27498619

  6. Marker-assisted selection in breeding silkworm strains with high tolerance to fluoride, scaleless wings, and high silk production.

    PubMed

    Li, M W; Yu, H J; Yi, X L; Li, J; Dai, F Y; Hou, C X

    2015-09-22

    In the silkworm (Bombyx mori), tolerance to fluoride and scaleless wings are controlled by the dominant gene Dtf (dominant tolerance to fluoride) and recessive gene nlw (no Lepidoptera wings), respectively, and these genes have been mapped by using simple sequence repeat and sequence tag site markers. Marker-assisted evaluation and selection of silkworms with fluoride tolerance and scaleless wings were used for predicting fluoride resistance and scaleless wings in backcrossed animals. A silkworm strain was bred using this method, and its economic characteristics were found to be similar to those of commercial silkworms. These methods will therefore be useful for silkworm breeding programs and in screening for two or more characteristics of interest for segregating populations.

  7. Development of a transposon-based marker system for mutation breeding in sorghum (Sorghum bicolor L.).

    PubMed

    Im, S B; Kwon, S-J; Ryu, J; Jeong, S W; Kim, J B; Ahn, J-W; Kim, S H; Jo, Y D; Choi, H-I; Kang, S-Y

    2016-09-16

    Under certain circumstances, transposable elements (TE) can create or reverse mutations and alter the genome size of a cell. Sorghum (Sorghum bicolor L.) is promising for plant transposon tagging due to its small genome size and its low content of repetitive DNA. We developed a marker system based on targeted region amplification polymorphisms (TE-TRAP) that uses the terminal inverted repeats (TIRs) of transposons. A total of 3816 class 2 transposons belonging to the PIF/Harbinger family were identified from the whole sorghum genome that produced five primers, including eight types of TIRs. To define the applicability and utilization of TE-TRAP, we used 21 individuals that had been bred after ɤ-ray irradiation. In total, 31 TE-TRAP, 16 TD, and 21 AFLP primer combinations generated 1133, 223, and 555 amplicons, respectively. The percent polymorphic marker was 62.8, 51.1, and 59.3% for the TE-TRAP, TD, and AFLP markers, respectively. Phylogenetic and principal component analyses revealed that TE-TRAP divided the 21 individuals into three groups. Analysis of molecular variance suggested that TE-TRAP had a higher level of genetic diversity than the other two marker systems. After verifying the efficiency of TE-TRAP, 189 sorghum individuals were used to investigate the associations between the markers and the ɤ-ray doses. Two significant associations were found among the polymorphic markers. This TE-based method provides a useful marker resource for mutation breeding research.

  8. Identifying Objective EEG Based Markers of Linear Vection in Depth

    PubMed Central

    Palmisano, Stephen; Barry, Robert J.; De Blasio, Frances M.; Fogarty, Jack S.

    2016-01-01

    This proof-of-concept study investigated whether a time-frequency EEG approach could be used to examine vection (i.e., illusions of self-motion). In the main experiment, we compared the event-related spectral perturbation (ERSP) data of 10 observers during and directly after repeated exposures to two different types of optic flow display (each was 35° wide by 29° high and provided 20 s of motion stimulation). Displays consisted of either a vection display (which simulated constant velocity forward self-motion in depth) or a control display (a spatially scrambled version of the vection display). ERSP data were decomposed using time-frequency Principal Components Analysis (t–f PCA). We found an increase in 10 Hz alpha activity, peaking some 14 s after display motion commenced, which was positively associated with stronger vection ratings. This followed decreases in beta activity, and was also followed by a decrease in delta activity; these decreases in EEG amplitudes were negatively related to the intensity of the vection experience. After display motion ceased, a series of increases in the alpha band also correlated with vection intensity, and appear to reflect vection- and/or motion-aftereffects, as well as later cognitive preparation for reporting the strength of the vection experience. Overall, these findings provide support for the notion that EEG can be used to provide objective markers of changes in both vection status (i.e., “vection/no vection”) and vection strength. PMID:27559328

  9. Automatic coregistration of volumetric images based on implanted fiducial markers.

    PubMed

    Koch, Martin; Maltz, Jonathan S; Belongie, Serge J; Gangadharan, Bijumon; Bose, Supratik; Shukla, Himanshu; Bani-Hashemi, Ali R

    2008-10-01

    The accurate delivery of external beam radiation therapy is often facilitated through the implantation of radio-opaque fiducial markers (gold seeds). Before the delivery of each treatment fraction, seed positions can be determined via low dose volumetric imaging. By registering these seed locations with the corresponding locations in the previously acquired treatment planning computed tomographic (CT) scan, it is possible to adjust the patient position so that seed displacement is accommodated. The authors present an unsupervised automatic algorithm that identifies seeds in both planning and pretreatment images and subsequently determines a rigid geometric transformation between the two sets. The algorithm is applied to the imaging series of ten prostate cancer patients. Each test series is comprised of a single multislice planning CT and multiple megavoltage conebeam (MVCB) images. Each MVCB dataset is obtained immediately prior to a subsequent treatment session. Seed locations were determined to within 1 mm with an accuracy of 97 +/- 6.1% for datasets obtained by application of a mean imaging dose of 3.5 cGy per study. False positives occurred in three separate instances, but only when datasets were obtained at imaging doses too low to enable fiducial resolution by a human operator, or when the prostate gland had undergone large displacement or significant deformation. The registration procedure requires under nine seconds of computation time on a typical contemporary computer workstation.

  10. Discriminative Projection Selection Based Face Image Hashing

    NASA Astrophysics Data System (ADS)

    Karabat, Cagatay; Erdogan, Hakan

    Face image hashing is an emerging method used in biometric verification systems. In this paper, we propose a novel face image hashing method based on a new technique called discriminative projection selection. We apply the Fisher criterion for selecting the rows of a random projection matrix in a user-dependent fashion. Moreover, another contribution of this paper is to employ a bimodal Gaussian mixture model at the quantization step. Our simulation results on three different databases demonstrate that the proposed method has superior performance in comparison to previously proposed random projection based methods.

  11. A Bio-Economic Case Study of Canadian Honey Bee (Hymenoptera: Apidae) Colonies: Marker-Assisted Selection (MAS) in Queen Breeding Affects Beekeeper Profits.

    PubMed

    Bixby, Miriam; Baylis, Kathy; Hoover, Shelley E; Currie, Rob W; Melathopoulos, Andony P; Pernal, Stephen F; Foster, Leonard J; Guarna, M Marta

    2017-03-16

    Over the past decade in North America and Europe, winter losses of honey bee (Hymenoptera: Apidae) colonies have increased dramatically. Scientific consensus attributes these losses to multifactorial causes including altered parasite and pathogen profiles, lack of proper nutrition due to agricultural monocultures, exposure to pesticides, management, and weather. One method to reduce colony loss and increase productivity is through selective breeding of queens to produce disease-, pathogen-, and mite-resistant stock. Historically, the only method for identifying desirable traits in honey bees to improve breeding was through observation of bee behavior. A team of Canadian scientists have recently identified markers in bee antennae that correspond to behavioral traits in bees and can be tested for in a laboratory. These scientists have demonstrated that this marker-assisted selection (MAS) can be used to produce hygienic, pathogen-resistant honey bee colonies. Based on this research, we present a beekeeping case study where a beekeeper's profit function is used to evaluate the economic impact of adopting colonies selected for hygienic behavior using MAS into an apiary. Our results show a net profit gain from an MAS colony of between 2% and 5% when Varroa mites are effectively treated. In the case of ineffective treatment, MAS generates a net profit benefit of between 9% and 96% depending on the Varroa load. When a Varroa mite population has developed some treatment resistance, we show that MAS colonies generate a net profit gain of between 8% and 112% depending on the Varroa load and degree of treatment resistance.

  12. Reliability of pedigree-based and genomic evaluations in selected populations.

    PubMed

    Gorjanc, Gregor; Bijma, Piter; Hickey, John M

    2015-08-14

    Reliability is an important parameter in breeding. It measures the precision of estimated breeding values (EBV) and, thus, potential response to selection on those EBV. The precision of EBV is commonly measured by relating the prediction error variance (PEV) of EBV to the base population additive genetic variance (base PEV reliability), while the potential for response to selection is commonly measured by the squared correlation between the EBV and breeding values (BV) on selection candidates (reliability of selection). While these two measures are equivalent for unselected populations, they are not equivalent for selected populations. The aim of this study was to quantify the effect of selection on these two measures of reliability and to show how this affects comparison of breeding programs using pedigree-based or genomic evaluations. Two scenarios with random and best linear unbiased prediction (BLUP) selection were simulated, where the EBV of selection candidates were estimated using only pedigree, pedigree and phenotype, genome-wide marker genotypes and phenotype, or only genome-wide marker genotypes. The base PEV reliabilities of these EBV were compared to the corresponding reliabilities of selection. Realized genetic selection intensity was evaluated to quantify the potential of selection on the different types of EBV and, thus, to validate differences in reliabilities. Finally, the contribution of different underlying processes to changes in additive genetic variance and reliabilities was quantified. The simulations showed that, for selected populations, the base PEV reliability substantially overestimates the reliability of selection of EBV that are mainly based on old information from the parental generation, as is the case with pedigree-based prediction. Selection on such EBV gave very low realized genetic selection intensities, confirming the overestimation and importance of genotyping both male and female selection candidates. The two measures of

  13. SNP-based markers for discriminating olive (Olea europaea L.) cultivars.

    PubMed

    Reale, S; Doveri, S; Díaz, A; Angiolillo, A; Lucentini, L; Pilla, F; Martín, A; Donini, P; Lee, D

    2006-09-01

    A set of 11 polymorphic markers (1 cleaved amplified polymorphic sequence (CAPS), 2 sequence-characterized amplified regions (SCARs), and 8 single-nucleotide polymorphism (SNP)-derived markers) was obtained for olive cultivar identification by comparing DNA sequences from different accessions. Marker development was more efficient, using sequences from the database rather than cloning arbitrary DNA fragments. Analyses of the sequences of 3 genes from 11 diverse cultivars revealed an SNP frequency of 1 per 190 base pairs in exons and 1 per 149 base pairs in introns. Most mutations were silent or had little perceptible effect on the polypeptide encoded. The higher incidence of transversions (55%) suggests that methylation is not the major driving force for DNA base changes. Evidence of linkage disequilibrium in 2 pairs of markers has been detected. The set of predominantly SNP-based markers was used to genotype 65 olive samples obtained from Europe and Australia, and was able clearly to discriminate 77% of the cultivars. Samples, putatively of the same cultivar but derived from different sources, were revealed as identical, demonstrating the utility of these markers as tools for resolving nomenclature issues. Genotyping data were used for constructing a dendrogram by UPGMA cluster analysis using the simple matching similarity coefficient. Relationships between cultivars are discussed in relation to the route of olive's spread.

  14. Annotated genetic linkage maps of Pinus pinaster Ait. from a Central Spain population using microsatellite and gene based markers

    PubMed Central

    2012-01-01

    Background Pinus pinaster Ait. is a major resin producing species in Spain. Genetic linkage mapping can facilitate marker-assisted selection (MAS) through the identification of Quantitative Trait Loci and selection of allelic variants of interest in breeding populations. In this study, we report annotated genetic linkage maps for two individuals (C14 and C15) belonging to a breeding program aiming to increase resin production. We use different types of DNA markers, including last-generation molecular markers. Results We obtained 13 and 14 linkage groups for C14 and C15 maps, respectively. A total of 211 and 215 markers were positioned on each map and estimated genome length was between 1,870 and 2,166 cM respectively, which represents near 65% of genome coverage. Comparative mapping with previously developed genetic linkage maps for P. pinaster based on about 60 common markers enabled aligning linkage groups to this reference map. The comparison of our annotated linkage maps and linkage maps reporting QTL information revealed 11 annotated SNPs in candidate genes that co-localized with previously reported QTLs for wood properties and water use efficiency. Conclusions This study provides genetic linkage maps from a Spanish population that shows high levels of genetic divergence with French populations from which segregating progenies have been previously mapped. These genetic maps will be of interest to construct a reliable consensus linkage map for the species. The importance of developing functional genetic linkage maps is highlighted, especially when working with breeding populations for its future application in MAS for traits of interest. PMID:23036012

  15. A system for tumor heterogeneity evaluation and diagnosis based on tumor markers measured routinely in the laboratory.

    PubMed

    Hui, Liu; Rixv, Liu; Xiuying, Zhou

    2015-12-01

    To develop an efficient and reliable approach to estimate tumor heterogeneity and improve tumor diagnosis using multiple tumor markers measured routinely in the clinical laboratory. A total of 161 patients with different cancers were recruited as the cancer group, and 91 patients with non-oncological conditions were required as the non-oncological disease group. The control group comprised 90 randomly selected healthy subjects. AFP, CEA, CYFRA, CA125, CA153, CA199, CA724, and NSE levels were measured in all these subjects with a chemiluminescent microparticle immunoassay. The tumor marker with the maximum S/CO value (sample test value:cutoff value for discriminating individuals with and without tumors) was considered as a specific tumor marker (STM) for an individual. Tumor heterogeneity index (THI)=N/P (N: number of STMs; P: percentage of individuals with STMs in a certain tumor population) was used to quantify tumor heterogeneity: high THI indicated high tumor heterogeneity. The tumor marker index (TMI), TMI = STM×(number of positive tumor markers+1), was used for diagnosis. The THIs of lung, gastric, and liver cancers were 8.33, 9.63, and 5.2, respectively, while the ROC-areas under the curve for TMI were 0.862, 0.809, and 0.966. In this study, we developed a novel index for tumor heterogeneity based on the expression of various routinely evaluated serum tumor markers. Development of an evaluation system for tumor heterogeneity on the basis of this index could provide an effective diagnostic tool for some cancers. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  16. [Genetic linkage map of Betula pendula Roth and Betula platyphylla Suk based on random amplified polymorphisms DNA markers].

    PubMed

    Jiang, Ting-Bo; Li, Shao-Chen; Gao, Fu-Ling; Ding, Bao-Jian; Qu, Yue-Jun; Tang, Xin-Hua; Liu, Gui-Feng; Jiang, Jing; Yang, Chuan-Ping

    2007-07-01

    Based on the genetic inheritance and segregation of random amplified polymorphism DNA (RAPDs) markers, the first mid-density linkage map for silver birch was constructed by using a pseudo-testcross mapping strategy. A segregating population including 80 progenies from the cross between Betula pendula Roth and B. platyphylla Suk was obtained. A set of 1,200 random oligonucleotide primers were screened, and 208 primers were selected to generate RAPD markers within a sample of 80 F1 progenies. A total of 364 segregating sites were identified. Among them, 307 belonged to 1 : 1 segregating site, and 36 belonged to 3 : 1 segregating site, others were found distorted from the normal 1 : 1 ratio. Altogether 307 sites segregating 1 : 1 (testcross configuration) were used to construct parent-specific linkage maps, 145 for B. pendula and 162 for B. platyphylla. The resulting linkage maps consisted of 145 marker sites in 14 groups (four or more sites per group), 6 triples and 6 pairs for B. pendula, which covered the map distance about 955.6 cM (Kosambi units). The average map distance between adjacent markers was 14.9 cM, and 162 linked marker site for B. platyphylla were mapped onto 15 groups (four or more sites per group), 4 triples and 6 pairs, which covered the map distance about 1,545.8 cM, and the average map distance between adjacent markers was 15.2 cM. Further study is warranted to integrate the two maps to one density map and to locate important genes on the maps.

  17. Genetic Diversity and Relatedness of Sweet Cherry (Prunus Avium L.) Cultivars Based on Single Nucleotide Polymorphic Markers

    PubMed Central

    Fernandez i Marti, Angel; Athanson, Blessing; Koepke, Tyson; Font i Forcada, Carolina; Dhingra, Amit; Oraguzie, Nnadozie

    2012-01-01

    Most previous studies on genetic fingerprinting and cultivar relatedness in sweet cherry were based on isoenzyme, RAPD, and simple sequence repeat (SSR) markers. This study was carried out to assess the utility of single nucleotide polymorphism (SNP) markers generated from 3′ untranslated regions (UTR) for genetic fingerprinting in sweet cherry. A total of 114 sweet cherry germplasm representing advanced selections, commercial cultivars, and old cultivars imported from different parts of the world were screened with seven SSR markers developed from other Prunus species and with 40 SNPs obtained from 3′ UTR sequences of Rainier and Bing sweet cherry cultivars. Both types of marker study had 99 accessions in common. The SSR data was used to validate the SNP results. Results showed that the average number of alleles per locus, mean observed heterozygosity, expected heterozygosity, and polymorphic information content values were higher in SSRs than in SNPs although both set of markers were similar in their grouping of the sweet cherry accessions as shown in the dendrogram. SNPs were able to distinguish sport mutants from their wild type germplasm. For example, “Stella” was separated from “Compact Stella.” This demonstrates the greater power of SNPs for discriminating mutants from their original parents than SSRs. In addition, SNP markers confirmed parentage and also determined relationships of the accessions in a manner consistent with their pedigree relationships. We would recommend the use of 3′ UTR SNPs for genetic fingerprinting, parentage verification, gene mapping, and study of genetic diversity in sweet cherry. PMID:22737155

  18. Amygdala activation as a marker for selective attention toward neutral faces in a chronic traumatic brain injury population.

    PubMed

    Young, Leanne R; Yu, Weikei; Holloway, Michael; Rodgers, Barry N; Chapman, Sandra B; Krawczyk, Daniel C

    2017-09-01

    There has been great interest in characterizing the response of the amygdala to emotional faces, especially in the context of social cognition. Although amygdala activation is most often associated with fearful or angry stimuli, there is considerable evidence that the response of the amygdala to neutral faces is both robust and reliable. This characteristic of amygdala function is of particular interest in the context of assessing populations with executive function deficits, such as traumatic brain injuries, which can be evaluated using fMRI attention modulation tasks that evaluate prefrontal control over representations, notably faces. The current study tested the hypothesis that the amygdala may serve as a marker of selective attention to neutral faces. Using fMRI, we gathered data within a chronic traumatic brain injury population. Blood Oxygenation Level Dependent (BOLD) signal change within the left and right amygdalae and fusiform face areas was measured while participants viewed neutral faces and scenes, under conditions requiring participants to (1) categorize pictures of faces and scenes, (2) selectively attend to either faces or scenes, or (3) attend to both faces and scenes. Findings revealed that the amygdala is an effective marker for selective attention to neutral faces and, furthermore, it was more face-specific than the fusiform face area. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Characterization of a higher plant herbicide-resistant phytoene desaturase and its use as a selectable marker.

    PubMed

    Arias, Renée S; Dayan, Franck E; Michel, Albrecht; Howell, J'Lynn; Scheffler, Brian E

    2006-03-01

    Three natural somatic mutations at codon 304 of the phytoene desaturase gene (pds) of Hydrilla verticillata (L. f. Royle) have been reported to provide resistance to the herbicide fluridone. We substituted the arginine 304 present in the wild-type H. verticillata phytoene desaturase (PDS) with all 19 other natural amino acids and tested PDS against fluridone. In in vitro assays, the threonine (Thr), cysteine (Cys), alanine (Ala) and glutamine (Gln) mutations imparted the highest resistance to fluridone. Thr, the three natural mutations [Cys, serine (Ser), histidine (His)] and the wild-type PDS protein were tested in vitro against seven inhibitors of PDS representing several classes of herbicides. These mutations conferred cross-resistance to norflurazon and overall negative cross-resistance to beflubutamid, picolinafen and diflufenican. The T3 generation of transgenic Arabidopsis thaliana plants harbouring the four selected mutations and wild-type pds had similar patterns of cross-resistance to the herbicides as observed in the in vitro assays. The Thr304 Hydrilla pds mutant proved to be an excellent marker for the selection of transgenic plants. Seedlings harbouring Thr304 pds had a maximum resistance to sensitivity (R/S) ratio of 57 and 14 times higher than that of the wild-type for treatments with norflurazon and fluridone, respectively. These plants exhibited normal growth and development, even after long-term exposure to herbicide. As Thr304 pds is of plant origin, it could become more acceptable than other selectable markers for use in genetically modified food.

  20. Multilocus Detection of Wolf x Dog Hybridization in Italy, and Guidelines for Marker Selection

    PubMed Central

    Randi, Ettore; Hulva, Pavel; Fabbri, Elena; Galaverni, Marco; Galov, Ana; Kusak, Josip; Bigi, Daniele; Bolfíková, Barbora Černá; Smetanová, Milena; Caniglia, Romolo

    2014-01-01

    Hybridization and introgression can impact the evolution of natural populations. Several wild canid species hybridize in nature, sometimes originating new taxa. However, hybridization with free-ranging dogs is threatening the genetic integrity of grey wolf populations (Canis lupus), or even the survival of endangered species (e.g., the Ethiopian wolf C. simensis). Efficient molecular tools to assess hybridization rates are essential in wolf conservation strategies. We evaluated the power of biparental and uniparental markers (39 autosomal and 4 Y-linked microsatellites, a melanistic deletion at the β-defensin CBD103 gene, the hypervariable domain of the mtDNA control-region) to identify the multilocus admixture patterns in wolf x dog hybrids. We used empirical data from 2 hybrid groups with different histories: 30 presumptive natural hybrids from Italy and 73 Czechoslovakian wolfdogs of known hybrid origin, as well as simulated data. We assessed the efficiency of various marker combinations and reference samples in admixture analyses using 69 dogs of different breeds and 99 wolves from Italy, Balkans and Carpathian Mountains. Results confirmed the occurrence of hybrids in Italy, some of them showing anomalous phenotypic traits and exogenous mtDNA or Y-chromosome introgression. Hybridization was mostly attributable to village dogs and not strictly patrilineal. The melanistic β-defensin deletion was found only in Italian dogs and in putative hybrids. The 24 most divergent microsatellites (largest wolf-dog FST values) were equally or more informative than the entire panel of 39 loci. A smaller panel of 12 microsatellites increased risks to identify false admixed individuals. The frequency of F1 and F2 was lower than backcrosses or introgressed individuals, suggesting hybridization already occurred some generations in the past, during early phases of wolf expansion from their historical core areas. Empirical and simulated data indicated the identification of the past

  1. Multilocus detection of wolf x dog hybridization in italy, and guidelines for marker selection.

    PubMed

    Randi, Ettore; Hulva, Pavel; Fabbri, Elena; Galaverni, Marco; Galov, Ana; Kusak, Josip; Bigi, Daniele; Bolfíková, Barbora Černá; Smetanová, Milena; Caniglia, Romolo

    2014-01-01

    Hybridization and introgression can impact the evolution of natural populations. Several wild canid species hybridize in nature, sometimes originating new taxa. However, hybridization with free-ranging dogs is threatening the genetic integrity of grey wolf populations (Canis lupus), or even the survival of endangered species (e.g., the Ethiopian wolf C. simensis). Efficient molecular tools to assess hybridization rates are essential in wolf conservation strategies. We evaluated the power of biparental and uniparental markers (39 autosomal and 4 Y-linked microsatellites, a melanistic deletion at the β-defensin CBD103 gene, the hypervariable domain of the mtDNA control-region) to identify the multilocus admixture patterns in wolf x dog hybrids. We used empirical data from 2 hybrid groups with different histories: 30 presumptive natural hybrids from Italy and 73 Czechoslovakian wolfdogs of known hybrid origin, as well as simulated data. We assessed the efficiency of various marker combinations and reference samples in admixture analyses using 69 dogs of different breeds and 99 wolves from Italy, Balkans and Carpathian Mountains. Results confirmed the occurrence of hybrids in Italy, some of them showing anomalous phenotypic traits and exogenous mtDNA or Y-chromosome introgression. Hybridization was mostly attributable to village dogs and not strictly patrilineal. The melanistic β-defensin deletion was found only in Italian dogs and in putative hybrids. The 24 most divergent microsatellites (largest wolf-dog FST values) were equally or more informative than the entire panel of 39 loci. A smaller panel of 12 microsatellites increased risks to identify false admixed individuals. The frequency of F1 and F2 was lower than backcrosses or introgressed individuals, suggesting hybridization already occurred some generations in the past, during early phases of wolf expansion from their historical core areas. Empirical and simulated data indicated the identification of the past

  2. Genetic divergence in elite castor bean lineages based on TRAP markers.

    PubMed

    Simões, K S; Silva, S A; Machado, E L; Silva, M S

    2017-09-27

    Castor bean (Ricinus communis L.) is a tropical plant of great commercial interest and a potential source of biodiesel. The development of genetically improved cultivars with high amounts of oil in the seeds and low ricin toxicity is crucial to increase the productivity of this crop. The use of TRAP (target region amplification polymorphism) markers to develop elite lineages and study genetic divergence is fundamental to advance the genetic improvement of this species. The goal of this study was to evaluate the genetic divergence among 40 elite lineages of R. communis, which belong to the NBIO-UFRB Genetic Improvement Program, using TRAP markers involved in the biosynthesis of oil and ricin. Total DNA was extracted and quantified from the leaf tissue of the castor bean plants, and 70 TRAP combinations (fixed and arbitrary primers) were used to genotype the 40 lineages. Of the 580 fragments amplified, 335 were polymorphic (58%). The genetic dissimilarity among the lineages was calculated by the Jaccard dissimilarity index using the UPGMA grouping method. A dendrogram was generated, and four groups formed, showing divergence among the elite lineages that favors selection. The TRAP molecular markers were efficient at characterizing the genetic variability among the lineages and, because TRAP markers are functional markers involved in the biosynthesis of oil and ricin, they are important when studying the association between a marker and a candidate gene.

  3. Genetic diversity analysis of common beans based on molecular markers

    PubMed Central

    Gill-Langarica, Homar R.; Muruaga-Martínez, José S.; Vargas-Vázquez, M.L. Patricia; Rosales-Serna, Rigoberto; Mayek-Pérez, Netzahualcoyotl

    2011-01-01

    A core collection of the common bean (Phaseolus vulgaris L.), representing genetic diversity in the entire Mexican holding, is kept at the INIFAP (Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias, Mexico) Germplasm Bank. After evaluation, the genetic structure of this collection (200 accessions) was compared with that of landraces from the states of Oaxaca, Chiapas and Veracruz (10 genotypes from each), as well as a further 10 cultivars, by means of four amplified fragment length polymorphisms (AFLP) +3/+3 primer combinations and seven simple sequence repeats (SSR) loci, in order to define genetic diversity, variability and mutual relationships. Data underwent cluster (UPGMA) and molecular variance (AMOVA) analyses. AFLP analysis produced 530 bands (88.5% polymorphic) while SSR primers amplified 174 alleles, all polymorphic (8.2 alleles per locus). AFLP indicated that the highest genetic diversity was to be found in ten commercial-seed classes from two major groups of accessions from Central Mexico and Chiapas, which seems to be an important center of diversity in the south. A third group included genotypes from Nueva Granada, Mesoamerica, Jalisco and Durango races. Here, SSR analysis indicated a reduced number of shared haplotypes among accessions, whereas the highest genetic components of AMOVA variation were found within accessions. Genetic diversity observed in the common-bean core collection represents an important sample of the total Phaseolus genetic variability at the main Germplasm Bank of INIFAP. Molecular marker strategies could contribute to a better understanding of the genetic structure of the core collection as well as to its improvement and validation. PMID:22215964

  4. Genetic diversity analysis of common beans based on molecular markers.

    PubMed

    Gill-Langarica, Homar R; Muruaga-Martínez, José S; Vargas-Vázquez, M L Patricia; Rosales-Serna, Rigoberto; Mayek-Pérez, Netzahualcoyotl

    2011-10-01

    A core collection of the common bean (Phaseolus vulgaris L.), representing genetic diversity in the entire Mexican holding, is kept at the INIFAP (Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias, Mexico) Germplasm Bank. After evaluation, the genetic structure of this collection (200 accessions) was compared with that of landraces from the states of Oaxaca, Chiapas and Veracruz (10 genotypes from each), as well as a further 10 cultivars, by means of four amplified fragment length polymorphisms (AFLP) +3/+3 primer combinations and seven simple sequence repeats (SSR) loci, in order to define genetic diversity, variability and mutual relationships. Data underwent cluster (UPGMA) and molecular variance (AMOVA) analyses. AFLP analysis produced 530 bands (88.5% polymorphic) while SSR primers amplified 174 alleles, all polymorphic (8.2 alleles per locus). AFLP indicated that the highest genetic diversity was to be found in ten commercial-seed classes from two major groups of accessions from Central Mexico and Chiapas, which seems to be an important center of diversity in the south. A third group included genotypes from Nueva Granada, Mesoamerica, Jalisco and Durango races. Here, SSR analysis indicated a reduced number of shared haplotypes among accessions, whereas the highest genetic components of AMOVA variation were found within accessions. Genetic diversity observed in the common-bean core collection represents an important sample of the total Phaseolus genetic variability at the main Germplasm Bank of INIFAP. Molecular marker strategies could contribute to a better understanding of the genetic structure of the core collection as well as to its improvement and validation.

  5. Increased prediction accuracy in wheat breeding trials using a marker x environment interaction genomic selection model

    USDA-ARS?s Scientific Manuscript database

    Genomic selection (GS) models use genome-wide genetic information to predict genetic values of candidates for selection. Originally these models were developed without considering genotype ' environment interaction (GE). Several authors have proposed extensions of the cannonical GS model that accomm...

  6. Historical overview and current status of genomic technology and marker assisted selection in beef cattle

    USDA-ARS?s Scientific Manuscript database

    Genomic technology has long been viewed as a transformative technology in that it has the potential to greatly alter strategies for livestock selection and management. Effective selection requires accurate predictions of genetic merit. Traditional genetic evaluations are obtained from data and ped...

  7. Tagging and mapping of a rice gall midge resistance gene, Gm8, and development of SCARs for use in marker-aided selection and gene pyramiding.

    PubMed

    Jain, A; Ariyadasa, R; Kumar, A; Srivastava, M N; Mohan, M; Nair, S

    2004-11-01

    Using amplified fragment length polymorphisms (AFLPs) and random amplified polymorphic DNAs (RAPDs), we have tagged and mapped Gm8, a gene conferring resistance to the rice gall midge (Orseolia oryzae), a major insect pest of rice, onto rice chromosome 8. Using AFLPs, two fragments, AR257 and AS168, were identified that were linked to the resistant and susceptible phenotypes, respectively. Another resistant phenotype-specific marker, AP19(587), was also identified using RAPDs. SCAR primers based on the sequence of the fragments AR257 and AS168 failed to reveal polymorphism between the resistant and the susceptible parents. However, PCR using primers based on the regions flanking AR257 revealed polymorphism that was phenotype-specific. In contrast, PCR carried out using primers flanking the susceptible phenotype-associated fragment AS168 produced a monomorphic fragment. Restriction digestion of these monomorphic fragments revealed polymorphism between the susceptible and resistant parents. Nucleotide BLAST searches revealed that the three fragments show strong homology to rice PAC and BAC clones that formed a contig representing the short arm of chromosome 8. PCR amplification using the above-mentioned primers on a larger population, derived from a cross between two indica rice varieties, Jhitpiti (resistant parent) and TN1 (susceptible parent), showed that there is a tight linkage between the markers and the Gm8 locus. These markers, therefore, have potential for use in marker-aided selection and pyramiding of Gm8 along with other previously tagged gall midge resistance genes [ Gm2, Gm4(t), and Gm7].

  8. Development of High Tryptophan Maize Near Isogenic Lines Adapted to Temperate Regions through Marker Assisted Selection - Impediments and Benefits

    PubMed Central

    Kostadinovic, Marija; Ignjatovic-Micic, Dragana; Vancetovic, Jelena; Ristic, Danijela; Bozinovic, Sofija; Stankovic, Goran; Mladenovic Drinic, Snezana

    2016-01-01

    Breeding program aimed at converting standard maize inbred lines to their quality protein maize (QPM) counterparts for growing in temperate climate is being conducted at Maize Research Institute (MRI). The objective of the research presented herein was to develop QPM versions of two commercial ZP inbreds through marker assisted selection (MAS) with opaque2 specific molecular markers, while maintaining their good agronomic performances and combining abilities. Donor line was a tropical QPM line CML 144. After two backcross and three selfing generations, six near isogenic lines (NILs) with 93% recovery of the recurrent parent genome were created from one cross. Average increments of 30% in tryptophan content and 36% in quality index were obtained, as well as kernels with less than 25% opaque endosperm. Grain yield was increased by 11–31% and combining abilities of the improved lines were on a par with the original line. Correlations between biochemical and agronomic parameters revealed that selection for plant height, ear length and kernel row number together with tryptophan content could be recommended for development of QPM with this material. However, several impediments emerged during selection. Major drawbacks in NIL development were small number of opaque2 recessive homozygotes (4.5% and 7.6% in BC2F2 of two crosses) and poor seed set throughout selection, which led to the loss of one cross. Moreover, in the other cross many plants in different generations had to be omitted from further selection due to the insufficient number of kernels. This phenomenon could be explained by incompatibility between pollen and style, possibly due to the exotic donor germplasm. Overall, it could be expected that the use of NILs, which are adapted to temperate climate and have high percentage of domestic germplasm, would outbalance the noted impediments and increase MAS efficiency in different breeding programs. PMID:27936171

  9. The use of the phosphomannose isomerase gene as alternative selectable marker for Agrobacterium-mediated transformation of flax (Linum usitatissimum).

    PubMed

    Lamblin, Frédéric; Aimé, Aurélie; Hano, Christophe; Roussy, Isabelle; Domon, Jean-Marc; Van Droogenbroeck, Bart; Lainé, Eric

    2007-06-01

    In order to meet the future requirement of using non-antibiotic resistance genes for the production of transgenic plants, we have adapted the selectable marker system PMI/mannose to be used in Agrobacterium-mediated transformation of flax (Linum usitatissimum L.) cv. Barbara. The Escherichia coli pmi gene encodes a phosphomannose isomerase (E.C. 5.1.3.8) that converts mannose-6-phosphate, an inhibitor of glycolysis, into fructose-6-phosphate (glycolysis intermediate). Its expression in transformed cells allows them to grow on mannose-selective medium. The Agrobacterium tumefaciens strain GV3101 (pGV2260) harbouring the binary vector pNOV2819 that carries the pmi gene under the control of the Cestrum yellow leaf curling virus constitutive promoter was used for transformation experiments. Transgenic flax plants able to root on mannose-containing medium were obtained from hypocotyl-derived calli that had been selected on a combination of 20 g L(-1) sucrose and 10 g L(-1) mannose. Their transgenic state was confirmed by PCR and Southern blotting. Transgene expression was detected by RT-PCR in leaves, stems and roots of in vitro grown primary transformants. The mean transformation efficiency of 3.6%, that reached 6.4% in one experiment was comparable to that obtained when using the nptII selectable marker on the same cultivar. The ability of T1 seeds to germinate on mannose-containing medium confirmed the Mendelian inheritance of the pmi gene in the progeny of primary transformants. These results indicate that the PMI/mannose selection system can be successfully used for the recovery of flax transgenic plants under safe conditions for human health and the environment.

  10. Marker-assisted selection to improve drought adaptation in maize: the backcross approach, perspectives, limitations, and alternatives.

    PubMed

    Ribaut, Jean-Marcel; Ragot, Michel

    2007-01-01

    A number of different marker-assisted selection (MAS) approaches do exist for the improvement of polygenic traits. Results of a marker-assisted backcross (MABC) selection experiment aimed at improving grain yield under drought conditions in tropical maize are presented and compared with alternative MAS strategies. The introgression of favourable alleles at five target regions involved in the expression of yield components and flowering traits increased grain yield and reduced the asynchrony between male and female flowering under water-limited conditions. Eighty-five per cent of the recurrent parent's genotype at non-target loci was recovered in only four generations of MABC by screening large segregating populations (2200 individuals) for three of the four generations. Selected MABC-derived BC(2)F(3) families were crossed with two testers and evaluated under different water regimes. Mean grain yield of MABC-derived hybrids was consistently higher than that of control hybrids (crosses from the recurrent parent to the same two testers as the MABC-derived families) under severe water stress conditions. Under those conditions, the best five MABC-derived hybrids yielded, on average, at least 50% more than control hybrids. Under mild water stress, defined as resulting in <50% yield reduction, no difference was observed between MABC-derived hybrids and the control plants, thus confirming that the genetic regulation for drought tolerance is dependent on stress intensity. MABC conversions involving several target regions are likely to result in partial rather than complete line conversion. Simulations were conducted to assess the utility of such partial conversions, i.e. containing favourable donor alleles at non-target regions, for subsequent phenotypic selection. The results clearly showed that selecting several genotypes (10-20) at each MABC cycle was most efficient. In the light of these results, alternative approaches to MABC are discussed, including recurrent

  11. The impact of the rank of marker variance-covariance matrix in principal component evaluation for genomic selection applications.

    PubMed

    Dimauro, C; Cellesi, M; Pintus, M A; Macciotta, N P P

    2011-12-01

    In genomic selection (GS) programmes, direct genomic values (DGV) are evaluated using information provided by high-density SNP chip. Being DGV accuracy strictly dependent on SNP density, it is likely that an increase in the number of markers per chip will result in severe computational consequences. Aim of present work was to test the effectiveness of principal component analysis (PCA) carried out by chromosome in reducing the marker dimensionality for GS purposes. A simulated data set of 5700 individuals with an equal number of SNP distributed over six chromosomes was used. PCs were extracted both genome-wide (ALL) and separately by chromosome (CHR) and used to predict DGVs. In the ALL scenario, the SNP variance-covariance matrix (S) was singular, positive semi-definite and contained null information which introduces 'spuriousness' in the derived results. On the contrary, the S matrix for each chromosome (CHR scenario) had a full rank. Obtained DGV accuracies were always better for CHR than ALL. Moreover, in the latter scenario, DGV accuracies became soon unsettled as the number of animals decreases, whereas in CHR, they remain stable till 900-1000 individuals. In real applications where a 54k SNP chip is used, the largest number of markers per chromosome is approximately 2500. Thus, a number of around 3000 genotyped animals could lead to reliable results when the original SNP variables are replaced by a reduced number of PCs. © 2011 Blackwell Verlag GmbH.

  12. QTL mapping of soybean oil content for marker-assisted selection in plant breeding program.

    PubMed

    Leite, D C; Pinheiro, J B; Campos, J B; Di Mauro, A O; Unêda-Trevisoli, S H

    2016-03-18

    The present study was undertaken to detect and map the quantitative trait loci (QTL) related to soybean oil content. We used 244 progenies derived from a bi-parental cross of the Lineage 69 (from Universidade Estadual Paulista "Júlio de Mesquita Filho"/Faculdade de Ciências Agrárias e Veterinárias - Breeding Program) and Tucunaré cultivar. A total of 358 simple sequence repeat (SSR; microsatellite) markers were used to investigate the polymorphism between the parental lines, and for the polymorphic lines all the F2 individuals were tested. Evaluation of the oil content and phenotype was performed with the aid of a Tango equipment by near infra-red reflectance spectroscopy, using single F2 seeds and F2:3 progenies, in triplicate. The data were analyzed by QTL Cartographer program for 56 SSR polymorphic markers. Two oil-content related QTLs were detected on K and H linkage groups. The total phenotypic variation explained by QTLs ranged from 7.8 to 46.75% for oil content. New QTLs were identified for the oil content in addition to those previously identified in other studies. The results reported in this study show that regions different from those already known could be involved in the genetic control of soybean oil content.

  13. Characterization of Nestin, a Selective Marker for Bone Marrow Derived Mesenchymal Stem Cells

    PubMed Central

    Xie, Liang; Zeng, Xin; Hu, Jing; Chen, Qianming

    2015-01-01

    Mesenchymal stem cells (MSCs) are multipotent cells capable of differentiating into multiple cell lineages and contributing to tissue repair and regeneration. Characterization of the physiological function of MSCs has been largely hampered by lack of unique markers. Nestin, originally found in neuroepithelial stem cells, is an intermediate filament protein expressed in the early stages of development. Increasing studies have shown a particular association between Nestin and MSCs. Nestin could characterize a subset of bone marrow perivascular MSCs which contributed to bone development and closely contacted with hematopoietic stem cells (HSCs). Nestin expressing (Nes+) MSCs also play a role in the progression of various diseases. However, Nes+ cells were reported to participate in angiogenesis as MSCs or endothelial progenitor cells (EPCs) in several tissues and be a heterogeneous population comprising mesenchymal cells and endothelial cells in the developing bone marrow. In this review article, we will summarize the progress of the research on Nestin, particularly the function of Nes+ cells in bone marrow, and discuss the feasibility of using Nestin as a specific marker for MSCs. PMID:26236348

  14. Tissue culture specificity of the tobacco ASA2 promoter driving hpt as a selectable marker for soybean transformation selection.

    PubMed

    Zernova, Olga; Zhong, Wei; Zhang, Xing-Hai; Widholm, Jack

    2008-11-01

    This study was carried out to determine if the tobacco anthranilate synthase ASA2 2.3 kb promoter drives tissue culture specific expression and if it is strong enough to drive hpt (hygromycin phosphotransferase) gene expression at a level sufficient to allow selection of transformed soybean embryogenic culture lines. A number of transformed cell lines were selected showing that the promoter was strong enough. Northern blot analysis of plant tissues did not detect hpt mRNA in the untransformed control or in the ASA2-hpt plants except in developing seeds while hpt mRNA was detected in all tissues of the CaMV35S-hpt positive control line plants. However, when the more sensitive RT-PCR assay was used all tissues of the ASA2-hpt plants except roots and mature seeds were found to contain detectable hpt mRNA. Embryogenic tissue cultures initiated from the ASA2-hpt plants contained hpt mRNA detectable by both northern and RT-PCR analysis and the cultures were hygromycin resistant. Friable callus initiated from leaves of ASA2-hpt plants did in some cases contain hpt mRNA that was only barely detectable by northern hybridization even though the callus was very hygromycin resistant. Thus the ASA2 promoter is strong enough to drive sufficient hpt expression in soybean embryogenic cultures for hygromycin selection and only very low levels of expression were found in most plant tissues with none in mature seeds.

  15. A novel linkage map of sugarcane with evidence for clustering of retrotransposon-based markers

    PubMed Central

    2012-01-01

    Background The development of sugarcane as a sustainable crop has unlimited applications. The crop is one of the most economically viable for renewable energy production, and CO2 balance. Linkage maps are valuable tools for understanding genetic and genomic organization, particularly in sugarcane due to its complex polyploid genome of multispecific origins. The overall objective of our study was to construct a novel sugarcane linkage map, compiling AFLP and EST-SSR markers, and to generate data on the distribution of markers anchored to sequences of scIvana_1, a complete sugarcane transposable element, and member of the Copia superfamily. Results The mapping population parents (‘IAC66-6’ and ‘TUC71-7’) contributed equally to polymorphisms, independent of marker type, and generated markers that were distributed into nearly the same number of co-segregation groups (or CGs). Bi-parentally inherited alleles provided the integration of 19 CGs. The marker number per CG ranged from two to 39. The total map length was 4,843.19 cM, with a marker density of 8.87 cM. Markers were assembled into 92 CGs that ranged in length from 1.14 to 404.72 cM, with an estimated average length of 52.64 cM. The greatest distance between two adjacent markers was 48.25 cM. The scIvana_1-based markers (56) were positioned on 21 CGs, but were not regularly distributed. Interestingly, the distance between adjacent scIvana_1-based markers was less than 5 cM, and was observed on five CGs, suggesting a clustered organization. Conclusions Results indicated the use of a NBS-profiling technique was efficient to develop retrotransposon-based markers in sugarcane. The simultaneous maximum-likelihood estimates of linkage and linkage phase based strategies confirmed the suitability of its approach to estimate linkage, and construct the linkage map. Interestingly, using our genetic data it was possible to calculate the number of retrotransposon scIvana_1 (~60) copies in the sugarcane

  16. ERP markers of target selection discriminate children with high vs. low working memory capacity

    PubMed Central

    Shimi, Andria; Nobre, Anna Christina; Scerif, Gaia

    2015-01-01

    Selective attention enables enhancing a subset out of multiple competing items to maximize the capacity of our limited visual working memory (VWM) system. Multiple behavioral and electrophysiological studies have revealed the cognitive and neural mechanisms supporting adults’ selective attention of visual percepts for encoding in VWM. However, research on children is more limited. What are the neural mechanisms involved in children’s selection of incoming percepts in service of VWM? Do these differ from the ones subserving adults’ selection? Ten-year-olds and adults used a spatial arrow cue to select a colored item for later recognition from an array of four colored items. The temporal dynamics of selection were investigated through EEG signals locked to the onset of the memory array. Both children and adults elicited significantly more negative activity over posterior scalp locations contralateral to the item to-be-selected for encoding (N2pc). However, this activity was elicited later and for longer in children compared to adults. Furthermore, although children as a group did not elicit a significant N2pc during the time-window in which N2pc was elicited in adults, the magnitude of N2pc during the “adult time-window” related to their behavioral performance during the later recognition phase of the task. This in turn highlights how children’s neural activity subserving attention during encoding relates to better subsequent VWM performance. Significant differences were observed when children were divided into groups of high vs. low VWM capacity as a function of cueing benefit. Children with large cue benefits in VWM capacity elicited an adult-like contralateral negativity following attentional selection of the to-be-encoded item, whereas children with low VWM capacity did not. These results corroborate the close coupling between selective attention and VWM from childhood and elucidate further the attentional mechanisms constraining VWM performance in

  17. ERP markers of target selection discriminate children with high vs. low working memory capacity.

    PubMed

    Shimi, Andria; Nobre, Anna Christina; Scerif, Gaia

    2015-01-01

    Selective attention enables enhancing a subset out of multiple competing items to maximize the capacity of our limited visual working memory (VWM) system. Multiple behavioral and electrophysiological studies have revealed the cognitive and neural mechanisms supporting adults' selective attention of visual percepts for encoding in VWM. However, research on children is more limited. What are the neural mechanisms involved in children's selection of incoming percepts in service of VWM? Do these differ from the ones subserving adults' selection? Ten-year-olds and adults used a spatial arrow cue to select a colored item for later recognition from an array of four colored items. The temporal dynamics of selection were investigated through EEG signals locked to the onset of the memory array. Both children and adults elicited significantly more negative activity over posterior scalp locations contralateral to the item to-be-selected for encoding (N2pc). However, this activity was elicited later and for longer in children compared to adults. Furthermore, although children as a group did not elicit a significant N2pc during the time-window in which N2pc was elicited in adults, the magnitude of N2pc during the "adult time-window" related to their behavioral performance during the later recognition phase of the task. This in turn highlights how children's neural activity subserving attention during encoding relates to better subsequent VWM performance. Significant differences were observed when children were divided into groups of high vs. low VWM capacity as a function of cueing benefit. Children with large cue benefits in VWM capacity elicited an adult-like contralateral negativity following attentional selection of the to-be-encoded item, whereas children with low VWM capacity did not. These results corroborate the close coupling between selective attention and VWM from childhood and elucidate further the attentional mechanisms constraining VWM performance in children.

  18. Diversity Analysis in Cannabis sativa Based on Large-Scale Development of Expressed Sequence Tag-Derived Simple Sequence Repeat Markers

    PubMed Central

    Cheng, Chaohua; Tang, Qing; Chen, Ping; Wang, Changbiao; Zang, Gonggu; Zhao, Lining

    2014-01-01

    Cannabis sativa L. is an important economic plant for the production of food, fiber, oils, and intoxicants. However, lack of sufficient simple sequence repeat (SSR) markers has limited the development of cannabis genetic research. Here, large-scale development of expressed sequence tag simple sequence repeat (EST-SSR) markers was performed to obtain more informative genetic markers, and to assess genetic diversity in cannabis (Cannabis sativa L.). Based on the cannabis transcriptome, 4,577 SSRs were identified from 3,624 ESTs. From there, a total of 3,442 complementary primer pairs were designed as SSR markers. Among these markers, trinucleotide repeat motifs (50.99%) were the most abundant, followed by hexanucleotide (25.13%), dinucleotide (16.34%), tetranucloetide (3.8%), and pentanucleotide (3.74%) repeat motifs, respectively. The AAG/CTT trinucleotide repeat (17.96%) was the most abundant motif detected in the SSRs. One hundred and seventeen EST-SSR markers were randomly selected to evaluate primer quality in 24 cannabis varieties. Among these 117 markers, 108 (92.31%) were successfully amplified and 87 (74.36%) were polymorphic. Forty-five polymorphic primer pairs were selected to evaluate genetic diversity and relatedness among the 115 cannabis genotypes. The results showed that 115 varieties could be divided into 4 groups primarily based on geography: Northern China, Europe, Central China, and Southern China. Moreover, the coefficient of similarity when comparing cannabis from Northern China with the European group cannabis was higher than that when comparing with cannabis from the other two groups, owing to a similar climate. This study outlines the first large-scale development of SSR markers for cannabis. These data may serve as a foundation for the development of genetic linkage, quantitative trait loci mapping, and marker-assisted breeding of cannabis. PMID:25329551

  19. Diversity analysis in Cannabis sativa based on large-scale development of expressed sequence tag-derived simple sequence repeat markers.

    PubMed

    Gao, Chunsheng; Xin, Pengfei; Cheng, Chaohua; Tang, Qing; Chen, Ping; Wang, Changbiao; Zang, Gonggu; Zhao, Lining

    2014-01-01

    Cannabis sativa L. is an important economic plant for the production of food, fiber, oils, and intoxicants. However, lack of sufficient simple sequence repeat (SSR) markers has limited the development of cannabis genetic research. Here, large-scale development of expressed sequence tag simple sequence repeat (EST-SSR) markers was performed to obtain more informative genetic markers, and to assess genetic diversity in cannabis (Cannabis sativa L.). Based on the cannabis transcriptome, 4,577 SSRs were identified from 3,624 ESTs. From there, a total of 3,442 complementary primer pairs were designed as SSR markers. Among these markers, trinucleotide repeat motifs (50.99%) were the most abundant, followed by hexanucleotide (25.13%), dinucleotide (16.34%), tetranucloetide (3.8%), and pentanucleotide (3.74%) repeat motifs, respectively. The AAG/CTT trinucleotide repeat (17.96%) was the most abundant motif detected in the SSRs. One hundred and seventeen EST-SSR markers were randomly selected to evaluate primer quality in 24 cannabis varieties. Among these 117 markers, 108 (92.31%) were successfully amplified and 87 (74.36%) were polymorphic. Forty-five polymorphic primer pairs were selected to evaluate genetic diversity and relatedness among the 115 cannabis genotypes. The results showed that 115 varieties could be divided into 4 groups primarily based on geography: Northern China, Europe, Central China, and Southern China. Moreover, the coefficient of similarity when comparing cannabis from Northern China with the European group cannabis was higher than that when comparing with cannabis from the other two groups, owing to a similar climate. This study outlines the first large-scale development of SSR markers for cannabis. These data may serve as a foundation for the development of genetic linkage, quantitative trait loci mapping, and marker-assisted breeding of cannabis.

  20. Feature subset selection based on relevance

    NASA Astrophysics Data System (ADS)

    Wang, Hui; Bell, David; Murtagh, Fionn

    In this paper an axiomatic characterisation of feature subset selection is presented. Two axioms are presented: sufficiency axiom—preservation of learning information, and necessity axiom—minimising encoding length. The sufficiency axiom concerns the existing dataset and is derived based on the following understanding: any selected feature subset should be able to describe the training dataset without losing information, i.e. it is consistent with the training dataset. The necessity axiom concerns the predictability and is derived from Occam's razor, which states that the simplest among different alternatives is preferred for prediction. The two axioms are then restated in terms of relevance in a concise form: maximising both the r( X; Y) and r( Y; X) relevance. Based on the relevance characterisation, four feature subset selection algorithms are presented and analysed: one is exhaustive and the remaining three are heuristic. Experimentation is also presented and the results are encouraging. Comparison is also made with some well-known feature subset selection algorithms, in particular, with the built-in feature selection mechanism in C4.5.

  1. Quasar Selection Based on Photometric Variability

    NASA Astrophysics Data System (ADS)

    MacLeod, C. L.; Brooks, K.; Ivezić, Ž.; Kochanek, C. S.; Gibson, R.; Meisner, A.; Kozłowski, S.; Sesar, B.; Becker, A. C.; de Vries, W. H.

    2011-02-01

    We develop a method for separating quasars from other variable point sources using Sloan Digital Sky Survey (SDSS) Stripe 82 light-curve data for ~ 10,000 variable objects. To statistically describe quasar variability, we use a damped random walk model parametrized by a damping timescale, τ, and an asymptotic amplitude (structure function), SF∞. With the aid of an SDSS spectroscopically confirmed quasar sample, we demonstrate that variability selection in typical extragalactic fields with low stellar density can deliver complete samples with reasonable purity (or efficiency, E). Compared to a selection method based solely on the slope of the structure function, the inclusion of the τ information boosts E from 60% to 75% while maintaining a highly complete sample (98%) even in the absence of color information. For a completeness of C = 90%, E is boosted from 80% to 85%. Conversely, C improves from 90% to 97% while maintaining E = 80% when imposing a lower limit on τ. With the aid of color selection, the purity can be further boosted to 96%, with C = 93%. Hence, selection methods based on variability will play an important role in the selection of quasars with data provided by upcoming large sky surveys, such as Pan-STARRS and the Large Synoptic Survey Telescope (LSST). For a typical (simulated) LSST cadence over 10 years and a photometric accuracy of 0.03 mag (achieved at i ≈ 22), C is expected to be 88% for a simple sample selection criterion of τ>100 days. In summary, given an adequate survey cadence, photometric variability provides an even better method than color selection for separating quasars from stars.

  2. A genetic linkage map for hazelnut (Corylus avellana L.) based on RAPD and SSR markers.

    PubMed

    Mehlenbacher, Shawn A; Brown, Rebecca N; Nouhra, Eduardo R; Gökirmak, Tufan; Bassil, Nahla V; Kubisiak, Thomas L

    2006-02-01

    A linkage map for European hazelnut (Corylus avellana L.) was constructed using random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) markers and the 2-way pseudotestcross approach. A full-sib population of 144 seedlings from the cross OSU 252.146 x OSU 414.062 was used. RAPD markers in testcross configuration, segregating 1:1, were used to construct separate maps for each parent. Fifty additional RAPD loci were assigned to linkage groups as accessory markers whose exact location could not be determined. Markers in intercross configuration, segregating 3:1, were used to pair groups in one parent with their homologues in the other. Eleven groups were identified for each parent, corresponding to the haploid chromosome number of hazelnut (n = x = 11). Thirty of the 31 SSR loci were able to be assigned to a linkage group. The maternal map included 249 RAPD and 20 SSR markers and spanned a distance of 661 cM. The paternal map included 271 RAPD and 28 SSR markers and spanned a distance of 812 cM. The maps are quite dense, with an average of 2.6 cM between adjacent markers. The S-locus, which controls pollen-stigma incompatibility, was placed on chromosome 5S where 6 markers linked within a distance of 10 cM were identified. A locus for resistance to eastern filbert blight, caused by Anisogramma anomala, was placed on chromosome 6R for which two additional markers tightly linked to the dominant allele were identified and sequenced. These maps will serve as a starting point for future studies of the hazelnut genome, including map-based cloning of important genes. The inclusion of SSR loci on the map will make it useful in other populations.

  3. Evaluation of genetic diversity in jackfruit (Artocarpus heterophyllus Lam.) based on amplified fragment length polymorphism markers.

    PubMed

    Shyamalamma, S; Chandra, S B C; Hegde, M; Naryanswamy, P

    2008-07-22

    Artocarpus heterophyllus Lam., commonly called jackfruit, is a medium-sized evergreen tree that bears high yields of the largest known edible fruit. Yet, it has been little explored commercially due to wide variation in fruit quality. The genetic diversity and genetic relatedness of 50 jackfruit accessions were studied using amplified fragment length polymorphism markers. Of 16 primer pairs evaluated, eight were selected for screening of genotypes based on the number and quality of polymorphic fragments produced. These primer combinations produced 5976 bands, 1267 (22%) of which were polymorphic. Among the jackfruit accessions, the similarity coefficient ranged from 0.137 to 0.978; the accessions also shared a large number of monomorphic fragments (78%). Cluster analysis and principal component analysis grouped all jackfruit genotypes into three major clusters. Cluster I included the genotypes grown in a jackfruit region of Karnataka, called Tamaka, with very dry conditions; cluster II contained the genotypes collected from locations having medium to heavy rainfall in Karnataka; cluster III grouped the genotypes in distant locations with different environmental conditions. Strong coincidence of these amplified fragment length polymorphism-based groupings with geographical localities as well as morphological characters was observed. We found moderate genetic diversity in these jackfruit accessions. This information should be useful for tree breeding programs, as part of our effort to popularize jackfruit as a commercial crop.

  4. A PCR-based forward genetics screening, using expression domain-specific markers, identifies mutants in endosperm transfer cell development

    PubMed Central

    Muñiz, Luis M.; Gómez, Elisa; Guyon, Virginie; López, Maribel; Khbaya, Bouchaib; Sellam, Olivier; Peréz, Pascual; Hueros, Gregorio

    2014-01-01

    Mutant collections are an invaluable source of material on which forward genetic approaches allow the identification of genes affecting a wide variety of biological processes. However, some particular developmental stages and morphological structures may resist analysis due to their physical inaccessibility or to deleterious effects associated to their modification. Furthermore, lethal mutations acting early in development may escape detection. We have approached the characterization of 101 maize seed mutants, selected from a collection of 27,500 visually screened Mu-insertion lines, using a molecular marker approach based on a set of genes previously ascribed to different tissue compartments within the early developing kernel. A streamlined combination of qRT-PCR assays has allowed us to preliminary pinpoint the affected compartment, establish developmental comparisons to WT siblings and select mutant lines with alterations in the different compartments. Furthermore, clusters of markers co-affected by the underlying mutation were identified. We have analyzed more extensively a set of lines presenting significant variation in transfer cell-associated expression markers, and have performed morphological observations, and immunolocalization experiments to confirm the results, validating this approach as an efficient mutant description tool. PMID:24808899

  5. Theoretical analysis of cell separation based on cell surface marker density.

    PubMed

    Chalmers, J J; Zborowski, M; Moore, L; Mandal, S; Fang, B B; Sun, L

    1998-07-05

    A theoretical analysis was performed to determine the number of fractions a multidisperse, immunomagnetically labeled cell population can be separated into based on the surface marker (antigen) density. A number of assumptions were made in this analysis: that there is a proportionality between the number of surface markers on the cell surface and the number of immunomagnetic labels bound; that this surface marker density is independent of the cell diameter; and that there is only the presence of magnetic and drag forces acting on the cell. Due to the normal distribution of cell diameters, a "randomizing" effect enters into the analysis, and an analogy between the "theoretical plate" analysis of distillation, adsorption, and chromatography can be made. Using the experimentally determined, normal distribution of cell diameters for human lymphocytes and a breast cancer cell line, and fluorescent activated cell screening data of specific surface marker distributions, examples of theoretical plate calculations were made and discussed.

  6. A genotypic and phenotypic information source for marker-assisted selection of cereals: the CEREALAB database

    PubMed Central

    Milc, Justyna; Sala, Antonio; Bergamaschi, Sonia; Pecchioni, Nicola

    2011-01-01

    The CEREALAB database aims to store genotypic and phenotypic data obtained by the CEREALAB project and to integrate them with already existing data sources in order to create a tool for plant breeders and geneticists. The database can help them in unravelling the genetics of economically important phenotypic traits; in identifying and choosing molecular markers associated to key traits; and in choosing the desired parentals for breeding programs. The database is divided into three sub-schemas corresponding to the species of interest: wheat, barley and rice; each sub-schema is then divided into two sub-ontologies, regarding genotypic and phenotypic data, respectively. Database URL: http://www.cerealab.unimore.it/jws/cerealab.jnlp PMID:21247929

  7. Localization Based on Magnetic Markers for an All-Wheel Steering Vehicle

    PubMed Central

    Byun, Yeun Sub; Kim, Young Chol

    2016-01-01

    Real-time continuous localization is a key technology in the development of intelligent transportation systems. In these systems, it is very important to have accurate information about the position and heading angle of the vehicle at all times. The most widely implemented methods for positioning are the global positioning system (GPS), vision-based system, and magnetic marker system. Among these methods, the magnetic marker system is less vulnerable to indoor and outdoor environment conditions; moreover, it requires minimal maintenance expenses. In this paper, we present a position estimation scheme based on magnetic markers and odometry sensors for an all-wheel-steering vehicle. The heading angle of the vehicle is determined by using the position coordinates of the last two detected magnetic markers and odometer data. The instant position and heading angle of the vehicle are integrated with an extended Kalman filter to estimate the continuous position. GPS data with the real-time kinematics mode was obtained to evaluate the performance of the proposed position estimation system. The test results show that the performance of the proposed localization algorithm is accurate (mean error: 3 cm; max error: 9 cm) and reliable under unexpected missing markers or incorrect markers. PMID:27916827

  8. Localization Based on Magnetic Markers for an All-Wheel Steering Vehicle.

    PubMed

    Byun, Yeun Sub; Kim, Young Chol

    2016-11-29

    Real-time continuous localization is a key technology in the development of intelligent transportation systems. In these systems, it is very important to have accurate information about the position and heading angle of the vehicle at all times. The most widely implemented methods for positioning are the global positioning system (GPS), vision-based system, and magnetic marker system. Among these methods, the magnetic marker system is less vulnerable to indoor and outdoor environment conditions; moreover, it requires minimal maintenance expenses. In this paper, we present a position estimation scheme based on magnetic markers and odometry sensors for an all-wheel-steering vehicle. The heading angle of the vehicle is determined by using the position coordinates of the last two detected magnetic markers and odometer data. The instant position and heading angle of the vehicle are integrated with an extended Kalman filter to estimate the continuous position. GPS data with the real-time kinematics mode was obtained to evaluate the performance of the proposed position estimation system. The test results show that the performance of the proposed localization algorithm is accurate (mean error: 3 cm; max error: 9 cm) and reliable under unexpected missing markers or incorrect markers.

  9. CAPs markers to assist selection for low vicine and convicine contents in faba bean (Vicia faba L.).

    PubMed

    Gutierrez, N; Avila, C M; Duc, G; Marget, P; Suso, M J; Moreno, M T; Torres, A M

    2006-12-01

    The antinutritional factors (ANFs) present in Vicia spp. seeds are a major constraint to the wider utilization of these crops as grain legumes. In the case of faba bean (Vicia faba L.), a breeding priority is the absence vicine and convicine (v-c); responsible for favism in humans and for the reduced animal performance or low egg production in laying hens. The discovery of a spontaneous mutant allele named vc-, which induces a 10-20 fold reduction of v-c contents, may facilitate the process. However, the high cost and difficulty of the chemical detection of v-c seriously restricts the advances in breeding-selection. To identify random amplified polymorphic DNA (RAPD) markers linked to this gene, we have analysed an F(2 )population derived from a cross between a line with high v-c content (Vf6) and the vc- genotype (line 1268). Quantification of v-c was done by spectrophotometry on the parents and the F(2 )population (n = 136). By using bulked segregant analysis (BSA), two RAPD markers linked in coupling and repulsion phase to the allele vc- were identified and further converted into sequence characterized amplified regions (SCARs). Amplification of SCARS was more consistent, although the initial polymorphism between pools was lost. To recover the polymorphisms several approaches were explored. Restriction digestion with HhaI (for SCAR SCH01(620)) and RsaI (for SCAR SCAB12(850)) revealed clear differences between the parental lines. The simultaneous use of the two cleavage amplified polymorphism (CAP) markers will allow the correct fingerprinting of faba bean plants and can be efficiently used in breeding selection to track the introgression of the vc- allele to develop cultivars with low v-c content and improved nutritional value.

  10. Recombineering with tolC as a Selectable/Counter-selectable Marker: remodeling the rRNA Operons of Escherichia coli

    PubMed Central

    DeVito, Joseph A.

    2008-01-01

    This work describes the novel use of tolC as a selectable/counter-selectable marker for the facile modification of DNA in Escherichia coli. Expression of TolC (an outer membrane protein) confers relative resistance to toxic small molecules, while its absence renders the cell tolerant to colicin E1. These features, coupled with the λredgam recombination system, allow for selection of tolC insertions/deletions anywhere on the E. coli chromosome or on plasmid DNA. This methodology obviates the need for minimal growth media, specialized wash protocols and the lengthy incubation times required by other published recombineering methods. As a rigorous test of the TolC selection system, six out of seven 23S rRNA genes were consecutively and seamlessly removed from the E. coli chromosome without affecting expression of neighboring genes within the complex rrn operons. The resulting plasmid-free strain retains one 23S rRNA gene (rrlC) in its natural location on the chromosome and is the first mutant of its kind. These new rRNA mutants will be useful in the study of rRNA gene regulation and ribosome function. Given its high efficiency, low background and facility in rich media, tolC selection is a broadly applicable method for the modification of DNA by recombineering. PMID:18084036

  11. AFLP genome scan in the black rat (Rattus rattus) from Madagascar: detecting genetic markers undergoing plague-mediated selection.

    PubMed

    Tollenaere, C; Duplantier, J-M; Rahalison, L; Ranjalahy, M; Brouat, C

    2011-03-01

    The black rat (Rattus rattus) is the main reservoir of plague (Yersinia pestis infection) in Madagascar's rural zones. Black rats are highly resistant to plague within the plague focus (central highland), whereas they are susceptible where the disease is absent (low altitude zone). To better understand plague wildlife circulation and host evolution in response to a highly virulent pathogen, we attempted to determine genetic markers associated with plague resistance in this species. To this purpose, we combined a population genomics approach and an association study, both performed on 249 AFLP markers, in Malagasy R. rattus. Simulated distributions of genetic differentiation were compared to observed data in four independent pairs, each consisting of one population from the plague focus and one from the plague-free zone. We found 22 loci (9% of 249) with higher differentiation in at least two independent population pairs or with combining P-values over the four pairs significant. Among the 22 outlier loci, 16 presented significant association with plague zone (plague focus vs. plague-free zone). Population genetic structure inferred from outlier loci was structured by plague zone, whereas the neutral loci dataset revealed structure by geography (eastern vs. western populations). A phenotype association study revealed that two of the 22 loci were significantly associated with differentiation between dying and surviving rats following experimental plague challenge. The 22 outlier loci identified in this study may undergo plague selective pressure either directly or more probably indirectly due to hitchhiking with selected loci. © 2010 Blackwell Publishing Ltd.

  12. [Bladder cancer screening with urine-based tumour markers - occupational medical experience].

    PubMed

    Nasterlack, M; Feil, G; Leng, G; Pesch, B; Huber, S; Sievert, K-D; Johnen, G; Taeger, D; Mayer, T; Kluckert, M; Brüning, T; Stenzl, A

    2011-03-01

    Bladder cancer responds favourably to treatment and has a good survival rate, provided it is diagnosed at an early stage. Established methods exist for the early detection, however, their specificity and positive predictive value are not yet satisfactory. Innovative markers have been proposed, but still require validation in prospective studies. We provide a literature-based short overview on the currently available and some proposed markers for the early detection of bladder cancer and evaluate the need for validation in further studies. We further provide some first results of such a recently finished study in an occupational setting. We conducted a prospective screening study over seven years in 1610 males with former occupational exposure to carcinogenic aromatic amines. Annual bladder cancer screening according to statutory requirements was offered. In addition to the regularly performed check for hematuria and urine cytology, the markers NMP22, UroVysion™ and survivin were performed in voided urine samples of the participants. Positive findings (not for survivin) were further followed through urethrocystoscopy. A total of 7219 urine samples were screened. During the study period 16 incidental and 4 recurrent bladder tumours, thereof three papillomas, occurred in a total of 19 participants. 14 out of twenty tumours were marker-positive, and all but two were early stage findings. Cell-based markers (cytology, UroVysion™) und molecular markers (NMP22, survivin) were largely complementary, thus acting as a "multi-marker panel". Eight of the tumours were identified by a positive cytology. Six tumours were not detected by any of the tumour markers. The results will be further evaluated through the inclusion of confounding factors, which have so far rarely been examined in other studies. This may lead to the development of tiered diagnostic strategies with the aim to reduce the number of invasive diagnostic procedures in the future. © Georg Thieme Verlag KG

  13. Deoxysphingoid bases as plasma markers in Diabetes mellitus

    PubMed Central

    2010-01-01

    Background Sphingoid bases are formed from the precursors L-serine and palmitoyl-CoA-a reaction which is catalyzed by the serine-palmitoyltransferase (SPT). SPT metabolizes, besides palmitoyl-CoA also other acyl-CoAs but shows also variability towards the use of other amino acid substrates. The enzyme is also able to metabolize alanine, which results in the formation of an atypical deoxy-sphingoid base (DSB). This promiscuous activity is greatly increased in the case of the sensory neuropathy HSAN1, and pathologically elevated DSB levels have been identified as the cause of this disease. Clinically, HSAN1 shows a pronounced similarity to the diabetic sensory neuropathy (DSN), which is the most common chronic complication of diabetes mellitus. Since serine and alanine metabolism is functionally linked to carbohydrate metabolism by their precursors 3-phosphoglycerate and pyruvate, we were interested to see whether the levels of certain sphingoid base metabolites are altered in patients with diabetes. Results In a case-control study we compared plasma sphingoid base levels between healthy and diabetic individuals. DSB levels were higher in the diabetic group whereas C16 and C18 sphingoid bases were not significantly different. Plasma serine, but not alanine levels were lower in the diabetic group. A subsequent lipoprotein fractionation showed that the DSBs are primarily present in the LDL and VLDL fraction. Conclusion Our results suggest that DSBs are a novel category of plasma biomarkers in diabetes which reflect functional impairments of carbohydrate metabolism. Furthermore, elevated DSB levels as we see them in diabetic patients might also contribute to the progression of the diabetic sensory neuropathy, the most frequent complication of diabetes. PMID:20712864

  14. Comparison between low-cost marker-less and high-end marker-based motion capture systems for the computer-aided assessment of working ergonomics.

    PubMed

    Patrizi, Alfredo; Pennestrì, Ettore; Valentini, Pier Paolo

    2016-01-01

    The paper deals with the comparison between a high-end marker-based acquisition system and a low-cost marker-less methodology for the assessment of the human posture during working tasks. The low-cost methodology is based on the use of a single Microsoft Kinect V1 device. The high-end acquisition system is the BTS SMART that requires the use of reflective markers to be placed on the subject's body. Three practical working activities involving object lifting and displacement have been investigated. The operational risk has been evaluated according to the lifting equation proposed by the American National Institute for Occupational Safety and Health. The results of the study show that the risk multipliers computed from the two acquisition methodologies are very close for all the analysed activities. In agreement to this outcome, the marker-less methodology based on the Microsoft Kinect V1 device seems very promising to promote the dissemination of computer-aided assessment of ergonomics while maintaining good accuracy and affordable costs. PRACTITIONER’S SUMMARY: The study is motivated by the increasing interest for on-site working ergonomics assessment. We compared a low-cost marker-less methodology with a high-end marker-based system. We tested them on three different working tasks, assessing the working risk of lifting loads. The two methodologies showed comparable precision in all the investigations.

  15. Effect of dietary Schizochytrium microalga oil on selected markers of low-grade inflammation in rats.

    PubMed

    Komprda, T; Sládek, Z; Škultéty, O; Křížková, S; Rozíková, V; Němcová, B; Šustrová, T; Valová, M

    2016-12-01

    The objective of the present study was to evaluate a potential of Schizochytrium microalga oil to alleviate possible negative effects of high-fat-high-energy diets. Forty adult male rats (Wistar Albino) were fed 7 weeks the diet containing beef tallow + evaporated sweetened milk (diet T) intended to cause mild obesity and low-grade systemic inflammation. Consequently, the animals were divided into four groups by 10 animals each and fed either the T-diet (control) or the diet containing 6% of safflower oil (S), 6% of fish oil (F) and 6% of Schizochytrium microalga oil (A), respectively, for another 7 weeks. The A-diet decreased (p < 0.05) live weight to 86% and glycaemia to 85% of control, respectively; an effect of the S- and F-diet on these markers was insignificant (p > 0.05). In comparison with control, higher (p < 0.05) deposition of eicosapentaenoic acid (EPA) + docosahexaenoic acid (DHA) in the epididymal adipose tissue (EAT) of the A-rats correlated with increased (p < 0.05) plasma adiponectin concentration, but it was without the effect (p > 0.05) on cellular adiponectin content in the EAT. Higher (p < 0.05) EPA+DHA deposition in the liver of the A-rats correlated with higher expression (149% of control; p < 0.05) of the gene coding for peroxisome proliferator-activated receptor gamma, and with lower expression (82% and 66%; p < 0.05) of the genes coding for adiponectin receptors AdipoR1 and AdipoR2; no relationship to the expression of receptor GPR120 was found. The A-diet did not affect amount of the nuclear fraction of the nuclear factor kappa B in the liver, but increased plasma level of anti-inflammatory cytokine TGF-β1 (p < 0.05). The presented data agree with results of other in vivo rodent and human studies, but not with literature data regarding in vitro experiments: it can be concluded that the effects of dietary oils on inflammatory markers need further investigation.

  16. Novel cloning vectors containing selectable genetic markers for use in streptomyces and related organisms

    SciTech Connect

    Fayerman, J.T.; Jones, M.D.; Malin, N.E.

    1987-05-19

    A recombinant DNA cloning vector is described comprising: a functional Streptomyces origin of replication-containing restriction fragment derived from the -- 8.8 kb BamHI restriction fragment of plasmid pFJ301, and one or more DNA segments that convey resistance to at least one antibiotic or that convey colorimetric selectivity when transformed into a sensitive restrictionless host cell.

  17. Sequence-Based Genotyping for Marker Discovery and Co-Dominant Scoring in Germplasm and Populations

    PubMed Central

    Truong, Hoa T.; Ramos, A. Marcos; Yalcin, Feyruz; de Ruiter, Marjo; van der Poel, Hein J. A.; Huvenaars, Koen H. J.; Hogers, René C. J.; van Enckevort, Leonora. J. G.; Janssen, Antoine; van Orsouw, Nathalie J.; van Eijk, Michiel J. T.

    2012-01-01

    Conventional marker-based genotyping platforms are widely available, but not without their limitations. In this context, we developed Sequence-Based Genotyping (SBG), a technology for simultaneous marker discovery and co-dominant scoring, using next-generation sequencing. SBG offers users several advantages including a generic sample preparation method, a highly robust genome complexity reduction strategy to facilitate de novo marker discovery across entire genomes, and a uniform bioinformatics workflow strategy to achieve genotyping goals tailored to individual species, regardless of the availability of a reference sequence. The most distinguishing features of this technology are the ability to genotype any population structure, regardless whether parental data is included, and the ability to co-dominantly score SNP markers segregating in populations. To demonstrate the capabilities of SBG, we performed marker discovery and genotyping in Arabidopsis thaliana and lettuce, two plant species of diverse genetic complexity and backgrounds. Initially we obtained 1,409 SNPs for arabidopsis, and 5,583 SNPs for lettuce. Further filtering of the SNP dataset produced over 1,000 high quality SNP markers for each species. We obtained a genotyping rate of 201.2 genotypes/SNP and 58.3 genotypes/SNP for arabidopsis (n = 222 samples) and lettuce (n = 87 samples), respectively. Linkage mapping using these SNPs resulted in stable map configurations. We have therefore shown that the SBG approach presented provides users with the utmost flexibility in garnering high quality markers that can be directly used for genotyping and downstream applications. Until advances and costs will allow for routine whole-genome sequencing of populations, we expect that sequence-based genotyping technologies such as SBG will be essential for genotyping of model and non-model genomes alike. PMID:22662172

  18. Selection of possible marker peptides for the detection of major ruminant milk proteins in food by liquid chromatography-tandem mass spectrometry.

    PubMed

    Ansari, Parisa; Stoppacher, Norbert; Rudolf, Judith; Schuhmacher, Rainer; Baumgartner, Sabine

    2011-01-01

    The aim of this work was the determination of peptides, which can function as markers for identification of milk allergens in food samples. Emphasis was placed on two casein proteins (α- and β-casein) and two whey proteins (α-lactalbumin and β-lactoglobulin). In silico tryptic digestion provided preliminary information about the expected peptides. After tryptic digestion of four milk allergens, the analytical data obtained by combination of reversed-phase high performance liquid chromatography and quadrupole tandem mass spectrometry (LC-MS/MS) led to the identification of 26 peptides. Seven of these peptides were synthesized and used for calibration of the LC-MS/MS system. Species specificity of the selected peptides was sought by BLAST search. Among the selected peptides, only LIVTQTMK from β-lactoglobulin (m/z 467.6, charge 2+) was found to be cow milk specific and could function as a marker. Two other peptides, FFVAPFPEVFGK from α-casein (m/z 693.3, charge 2+) and GPFPIIV from β-casein (m/z 742.5, charge 1+), occur in water buffalo milk too. The other four peptides appear in the milk of other species also and can be used as markers for ruminant species milk. Using these seven peptides, a multianalyte MS-based method was developed. For the establishment of the method, it was applied at first to different dairy samples, and then to chocolate and blank samples, and the peptides could be determined down to 1 ng/mL in food samples. At the end, spiked samples were measured, where the target peptides could be detected with a high recovery (over 50%).

  19. Marker-Based Estimation of Genetic Parameters in Genomics

    PubMed Central

    Hu, Zhiqiu; Yang, Rong-Cai

    2014-01-01

    Linear mixed model (LMM) analysis has been recently used extensively for estimating additive genetic variances and narrow-sense heritability in many genomic studies. While the LMM analysis is computationally less intensive than the Bayesian algorithms, it remains infeasible for large-scale genomic data sets. In this paper, we advocate the use of a statistical procedure known as symmetric differences squared (SDS) as it may serve as a viable alternative when the LMM methods have difficulty or fail to work with large datasets. The SDS procedure is a general and computationally simple method based only on the least squares regression analysis. We carry out computer simulations and empirical analyses to compare the SDS procedure with two commonly used LMM-based procedures. Our results show that the SDS method is not as good as the LMM methods for small data sets, but it becomes progressively better and can match well with the precision of estimation by the LMM methods for data sets with large sample sizes. Its major advantage is that with larger and larger samples, it continues to work with the increasing precision of estimation while the commonly used LMM methods are no longer able to work under our current typical computing capacity. Thus, these results suggest that the SDS method can serve as a viable alternative particularly when analyzing ‘big’ genomic data sets. PMID:25025305

  20. Development of Expressed Sequence Tag (EST)-based Cleaved Amplified Polymorphic Sequence (CAPS) markers of tea plant and their application to cultivar identification.

    PubMed

    Ujihara, Tomomi; Taniguchi, Fumiya; Tanaka, Jun-Ichi; Hayashi, Nobuyuki

    2011-03-09

    To develop cleaved amplified polymorphic sequence (CAPS) markers for cultivar identification of the tea leaf, 5 primer pairs designed on the basis of genes that encode proteins related to nitrogen assimilation and 26 primer pairs based on expressed sequence tag (EST) sequences of the root of tea plant were screened. From combinations of primer pair and restriction enzyme that showed polymorphism among tea plants, 16 markers were selected and applied to DNA fingerprinting of Japanese tea cultivars. Sixty-three cultivars, except for a bud sport (Kiraka) and its original cultivar (Yabukita) and a pair that was the progeny of the same crossing parent (Harumoegi and Sakimidori), were distinguished from one another. By combining the 16 markers with previously developed CAPS markers and observing the physical appearance, 67 cultivars were distinguishable. The cultivars involve approximately 95% of total tea cultivating area in Japan; therefore, about 95% of tea leaves produced in Japan can be authenticated by labeling their cultivars.

  1. An annotated genetic map of loblolly pine based on microsatellite and cDNA markers

    PubMed Central

    2011-01-01

    Background Previous loblolly pine (Pinus taeda L.) genetic linkage maps have been based on a variety of DNA polymorphisms, such as AFLPs, RAPDs, RFLPs, and ESTPs, but only a few SSRs (simple sequence repeats), also known as simple tandem repeats or microsatellites, have been mapped in P. taeda. The objective of this study was to integrate a large set of SSR markers from a variety of sources and published cDNA markers into a composite P. taeda genetic map constructed from two reference mapping pedigrees. A dense genetic map that incorporates SSR loci will benefit complete pine genome sequencing, pine population genetics studies, and pine breeding programs. Careful marker annotation using a variety of references further enhances the utility of the integrated SSR map. Results The updated P. taeda genetic map, with an estimated genome coverage of 1,515 cM(Kosambi) across 12 linkage groups, incorporated 170 new SSR markers and 290 previously reported SSR, RFLP, and ESTP markers. The average marker interval was 3.1 cM. Of 233 mapped SSR loci, 84 were from cDNA-derived sequences (EST-SSRs) and 149 were from non-transcribed genomic sequences (genomic-SSRs). Of all 311 mapped cDNA-derived markers, 77% were associated with NCBI Pta UniGene clusters, 67% with RefSeq proteins, and 62% with functional Gene Ontology (GO) terms. Duplicate (i.e., redundant accessory) and paralogous markers were tentatively identified by evaluating marker sequences by their UniGene cluster IDs, clone IDs, and relative map positions. The average gene diversity, He, among polymorphic SSR loci, including those that were not mapped, was 0.43 for 94 EST-SSRs and 0.72 for 83 genomic-SSRs. The genetic map can be viewed and queried at http://www.conifergdb.org/pinemap. Conclusions Many polymorphic and genetically mapped SSR markers are now available for use in P. taeda population genetics, studies of adaptive traits, and various germplasm management applications. Annotating mapped genes with Uni

  2. Updates on Immunohistochemical and Molecular Markers in Selected Head and Neck Diagnostic Problems.

    PubMed

    Yi, Chun-Hui; Jim Zhai, Qihui; Wang, Beverly Y

    2017-09-01

    - The head and neck regions have complex anatomic structures. They are not exempted from the rare occurrences of highly unusual, diagnostically challenging malignant neoplasms and mimickers. - To review and update the utility of immunohistochemistry and molecular biomarkers and to pursue diagnostic accuracy on selected rare neoplasms, especially some poorly differentiated malignancies. - Personal experience and information from the literature. - Head and neck tumors include neoplasms originating from heterogeneous tissue. Using the selected clinical cases, this review illustrates a continuous development of emerging molecular-genetic techniques to assist in the interpretation of uncommon, often poorly differentiated, highly malignant neoplasms. The diagnostic results are appropriately transmitted to the oncologists, radiation oncologists, and surgeons to create a coordinated plan of care for patients with these unusual disorders affecting the head and neck.

  3. The Swift Turbidity Marker

    ERIC Educational Resources Information Center

    Omar, Ahmad Fairuz; MatJafri, Mohd Zubir

    2011-01-01

    The Swift Turbidity Marker is an optical instrument developed to measure the level of water turbidity. The components and configuration selected for the system are based on common turbidity meter design concepts but use a simplified methodology to produce rapid turbidity measurements. This work is aimed at high school physics students and is the…

  4. The Swift Turbidity Marker

    ERIC Educational Resources Information Center

    Omar, Ahmad Fairuz; MatJafri, Mohd Zubir

    2011-01-01

    The Swift Turbidity Marker is an optical instrument developed to measure the level of water turbidity. The components and configuration selected for the system are based on common turbidity meter design concepts but use a simplified methodology to produce rapid turbidity measurements. This work is aimed at high school physics students and is the…

  5. Cytosine deaminase MX cassettes as positive/negative selectable markers in Saccharomyces cerevisiae.

    PubMed

    Hartzog, Phillip E; Nicholson, Bradly P; McCusker, John H

    2005-07-30

    We describe positive/negative selectable cytosine deaminase MX cassettes for use in Saccharomyces cerevisiae. The basis of positive selection for cytosine deaminase (Fcy1) activity is that (a) fcy1 strains are unable to grow on medium containing cytosine as a sole nitrogen source and (b) fcy1 ura3 strains are unable to grow on medium containing cytosine as the sole pyrimidine source. Conversely, as 5-fluorocytosine (5FC) is toxic to cytosine deaminase-producing cells, fcy1 strains are resistant to 5FC. FCY1MX and FCA1MX cassettes, containing open reading frames (ORFs) of S. cerevisiae FCY1 and Candida albicans FCA1, respectively, were constructed and used to disrupt targeted genes in S. cerevisiae fcy1 strains. In addition, new direct repeat cassettes, kanPR, FCA1PR, FCY1PR and CaURA3PR, were developed to allow efficient deletion of target genes in cells containing MX3 repeats. Finally, the FCY1- and FCA1MX3 or PR direct repeat cassettes can be readily recycled after 5FC counter-selection on both synthetic and rich media.

  6. Kinematical models to reduce the effect of skin artifacts on marker-based human motion estimation.

    PubMed

    Cerveri, P; Pedotti, A; Ferrigno, G

    2005-11-01

    The estimation of the skeletal motion obtained from marker-based motion capture systems is known to be affected by significant bias caused by skin movement artifacts, which affects joint center and rotation axis estimation. Among different techniques proposed in the literature, that based on rigid body model, still the most used by commercial motion capture systems, can smooth only part of the above effects without eliminating their main components. In order to sensibly improve the accuracy of the motion estimation, a novel technique, named local motion estimation (LME), is proposed. This rests on a recently described approach that, using virtual humans and extended Kalman filters, estimates the kinematical variables directly from 2D measurements without requiring the 3D marker reconstruction. In this paper, we show how such method can be extended to include the computation of the local marker displacement due to skin artifacts. The 3D marker coordinates, expressed in the corresponding local reference coordinate frames, are inserted into the state vector of the filter and their dynamics is automatically estimated, with adequate accuracy, without assuming any particular deformation function. Simulated experiments of lower limb motion, involving systematic mislocations (5, 10, 20 mm) and random errors of the marker coordinates and joint center locations (+/-5, +/-10, +/-15 mm), have shown that artifact motion can be substantially decoupled from the global skeletal motion with an effective increase of the accuracy wrt standard techniques. In particular, the comparison between the nominal kinematical variables and the one recovered from markers attached to the skin surface proved LME to be sensibly superior (50% in the worse condition) to the methods imposing marker-bone rigidity. In conclusion, while requiring further validation on real movement data, we argue that the proposed method can constitute an appropriate approach toward the improvement of the human motion

  7. The RNA binding protein RBPMS is a selective marker of ganglion cells in the mammalian retina

    PubMed Central

    Rodriguez, Allen R.; de Sevilla Müller, Luis Pérez; Brecha, Nicholas C.

    2014-01-01

    There are few neurochemical markers that reliably identify retinal ganglion cells (RGCs), which are a heterogeneous population of cells that integrate and transmit the visual signal from the retina to the central visual nuclei. We have developed and characterized a new set of affinity purified guinea pig and rabbit antibodies against RNA-binding protein with multiple splicing (RBPMS). On Western blots these antibodies recognize a single band at ~24 kDa, corresponding to RBPMS, and they strongly label RGC and displaced RGC (dRGC) somata in mouse, rat, guinea pig, rabbit and monkey retina. RBPMS immunoreactive cells and RGCs identified by other techniques have a similar range of somal diameters and areas. The density of RBPMS cells in mouse and rat retina is comparable to earlier semi-quantitative estimates of RGCs. RBPMS is mainly expressed in medium and large DAPI-, DRAQ5-, NeuroTrace- and NeuN-stained cells in the ganglion cell layer (GCL), and RBPMS is not expressed in syntaxin (HPC-1) immunoreactive cells in the inner nuclear layer (INL) and GCL, consistent with their identity as RGCs, and not displaced amacrine cells. In mouse and rat retina, most RBPMS cells are lost following optic nerve crush or transection at three weeks, and all Brn3a, SMI-32 and melanopsin immunoreactive RGCs also express RBPMS immunoreactivity. RBPMS immunoreactivity is localized to CFP-fluorescent RGCs in the B6.Cg-Tg(Thy1-CFP)23Jrs/J mouse line. These findings show that antibodies against RBPMS are robust reagents that exclusively identify RGCs and dRGCs in multiple mammalian species, and they will be especially useful for quantification of RGCs. PMID:24318667

  8. Systemic inflammation in peripheral arterial disease with or without coexistent chronic obstructive pulmonary disease: analysis of selected markers

    PubMed Central

    Sleszycka, Justyna; Safianowska, Aleksandra; Wiechno, Wieslaw; Domagala-Kulawik, Joanna

    2012-01-01

    Introduction Low-grade systemic inflammation plays an important role in the pathogenesis and natural history of chronic obstructive pulmonary disease (COPD) and peripheral arterial disease (PAD). The aim of the study was to analyze plasma concentrations of selected markers of inflammation in patients suffering from PAD with or without coexistent COPD. Material and methods Thirty patients (6 women) with advanced PAD (at least IIb stage according to Fontaine scale) hospitalized due to critical limb ischemia were examined. In all patients spirometry was performed to confirm or exclude COPD. Plasma concentration of IL-6, IL-8 and TNF-α was measured using ELISA method. Statistical analysis was performed according to COPD status and according to smoking status independently. Results In the whole group of patients with PAD, COPD was recognized in 14 cases (for the first time in 10 cases). All patients were smokers (46.7% current, 53.3% ex-smokers). We found a significant correlation between FEV1%N (percent of norm of first second expiratory volume) and the number of years of smoking (r = –0.39; p < 0.05). We found similar concentrations of IL-6 (2.54 pg/ml vs. 2.31 pg/ml), IL-8 (8.55 pg/ml vs. 8.14 pg/ml, TNF-α (0.72 pg/ml vs. 1.75 pg/ml) in the COPD(+) group in comparison to the COPD(–) group (differences were not significant). We observed significant positive correlations (p < 0.05) between concentrations of measured markers and significant negative correlations between pain free walking distance and these markers. Conclusions Our study confirmed coexistence of PAD with COPD. The character of inflammation is similar in these smoking-related diseases PMID:22852003

  9. Fluorescence-based resource for semiautomated genomic analyses using microsatellite markers

    SciTech Connect

    Levitt, R.C.; Kiser, M.B.; Dragwa, C.

    1994-11-15

    To facilitate the practical application of highly-efficient semiautomated methods for general application in genomic analyses, the authors have developed a fluorescence-based microsatellite marker resource. Ninety highly polymorphic microsatellite markers were combined to provide a rapid, accurate, and highly efficient initial genome-wide screening system. These markers are spaced on average every 33 cM, with a mean heterozygosity of 81% (range 65-94%), covering 22 autosomes and the X and Y chromosomes. Less than 10% of the genome lies beyond 20 cM of the nearest marker. Since this genomic analysis system is fully compatible with automated fragment analyzers using simultaneous four-color fluorescence-based detection systems, the 5 groups of 18 markers can be detected concurrently. This multiplex detection provides a throughput of 1944 genotypes daily per instrument. This system will be highly beneficial in a number of clinical and research applications including linkage, cancer genetics, forensics, and cytogenetics. 16 refs., 1 fig., 2 tabs.

  10. Stimulator Selection in SSVEP-Based Spatial Selective Attention Study.

    PubMed

    Xie, Songyun; Liu, Chang; Obermayer, Klaus; Zhu, Fangshi; Wang, Linan; Xie, Xinzhou; Wang, Wei

    2016-01-01

    Steady-State Visual Evoked Potentials (SSVEPs) are widely used in spatial selective attention. In this process the two kinds of visual simulators, Light Emitting Diode (LED) and Liquid Crystal Display (LCD), are commonly used to evoke SSVEP. In this paper, the differences of SSVEP caused by these two stimulators in the study of spatial selective attention were investigated. Results indicated that LED could stimulate strong SSVEP component on occipital lobe, and the frequency of evoked SSVEP had high precision and wide range as compared to LCD. Moreover a significant difference between noticed and unnoticed frequencies in spectrum was observed whereas in LCD mode this difference was limited and selectable frequencies were also limited. Our experimental finding suggested that average classification accuracies among all the test subjects in our experiments were 0.938 and 0.853 in LED and LCD mode, respectively. These results indicate that LED simulator is appropriate for evoking the SSVEP for the study of spatial selective attention.

  11. Heat shock induced excision of selectable marker genes in transgenic banana by the Cre-lox site-specific recombination system.

    PubMed

    Chong-Pérez, Borys; Kosky, Rafael G; Reyes, Maritza; Rojas, Luis; Ocaña, Bárbara; Tejeda, Marisol; Pérez, Blanca; Angenon, Geert

    2012-06-30

    Selectable marker genes are indispensable for efficient production of transgenic events, but are no longer needed after the selection process and may cause public concern and technological problems. Although several gene excision systems exist, few have been optimized for vegetatively propagated crops. Using a Cre-loxP auto-excision strategy, we obtained transgenic banana plants cv. Grande Naine (Musa AAA) devoid of the marker gene used for selection. We used T-DNA vectors with the cre recombinase gene under control of a heat shock promoter and selectable marker gene cassettes placed between two loxP sites in direct orientation, and a gene of interest inserted outside of the loxP sites. Heat shock promoters pGmHSP17.6-L and pHSP18.2, from soybean and Arabidopsis respectively, were tested. A transient heat shock treatment of primary transgenic embryos was sufficient for inducing cre and excising cre and the marker genes. Excision efficiency, as determined by PCR and Southern hybridization was 59.7 and 40.0% for the GmHSP17.6-L and HSP18.2 promoters, respectively. Spontaneous excision was not observed in 50 plants derived from untreated transgenic embryos. To our knowledge this is the first report describing an efficient marker gene removal system for banana. The method described is simple and might be generally applicable for the production of marker-free transgenic plants of many crop species.

  12. Genetic diversity in South African Nguni cattle ecotypes based on microsatellite markers.

    PubMed

    Sanarana, Yandisiwe; Visser, Carina; Bosman, Lydia; Nephawe, Khathutshelo; Maiwashe, Azwihangwisi; van Marle-Köster, Este

    2016-02-01

    The Nguni cattle breed is a landrace breed adapted to different ecological regions of South Africa. A number of ecotypes are recognised based on phenotype within the breed, but it is not known if they are genetically distinct. In this study, molecular characterisation was performed on Makhathini (MAK), Pedi (PED), Shangaan (SHA) and Venda (VEN) Nguni cattle ecotypes. Two Nguni cattle populations, not kept as separate ecotypes, from the University of Fort Hare (UFH) and Agricultural Research Council Loskop South farm (LOS) were also included. Genotypic data was generated for 189 unrelated Nguni cattle selected based on pedigree records using 22 microsatellite markers. The expected heterozygosity values varied from 69 % (UFH) to 72 % (PED) with a mean number of alleles ranging from 6.0 to 6.9. The F ST estimate demonstrated that 4.8 % of the total genetic variation was due to the genetic differentiation between the populations and 92.2 % accounted for differences within the populations. The genetic distances and structure analysis revealed the closest relationship between MAK, PEDI and SHA ecotypes, followed by SHA and VEN. The UFH population clustered with the MAK ecotype, indicating that they are more genetically similar, while the LOS cattle grouped as a distinct cluster. Results suggest that the genetic differentiation between the PED and SHA ecotypes is low and can be regarded as one ecotype based on limited genetic differences. The results of this study can be applied as a point of reference for further genetic studies towards conservation of Nguni cattle ecotypes.

  13. Marker recycling via 5-fluoroorotic acid and 5-fluorocytosine counter-selection in the white-rot agaricomycete Pleurotus ostreatus.

    PubMed

    Nakazawa, Takehito; Tsuzuki, Masami; Irie, Toshikazu; Sakamoto, Masahiro; Honda, Yoichi

    2016-09-01

    Of all of the natural polymers, lignin, an aromatic heteropolymer in plant secondary cell walls, is the most resistant to biological degradation. White-rot fungi are the only known organisms that can depolymerize or modify wood lignin. Investigating the mechanisms underlying lignin biodegradation by white-rot fungi would contribute to the ecofriendly utilization of woody biomass as renewable resources in the future. Efficient gene disruption, which is generally very challenging in the white-rot fungi, was established in Pleurotus ostreatus (the oyster mushroom). Some of the genes encoding manganese peroxidases, enzymes that are considered to be involved in lignin biodegradation, were disrupted separately, and the phenotype of each single-gene disruptant was analysed. However, it remains difficult to generate multi-gene disruptants in this fungus. Here we developed a new genetic transformation marker in P. ostreatus and demonstrated two marker recycling methods that use counter-selection to generate a multigene disruptant. This study will enable future genetic studies of white-rot fungi, and it will increase our understanding of the complicated mechanisms, which involve various enzymes, including lignin-degrading enzymes, underlying lignin biodegradation by these fungi. Copyright © 2016 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  14. A non-destructive genotyping system from a single seed for marker-assisted selection in watermelon.

    PubMed

    Meru, G; McDowell, D; Waters, V; Seibel, A; Davis, J; McGregor, C

    2013-03-11

    Genomic tools for watermelon breeding are becoming increasingly available. A high throughput genotyping system would facilitate the use of DNA markers in marker-assisted selection. DNA extraction from leaf material requires prior seed germination and is often time-consuming and cost prohibitive. In an effort to develop a more efficient system, watermelon seeds of several genotypes and various seed sizes were sampled by removing ⅓ or ½ sections from the distal ends for DNA extraction, while germinating the remaining proximal parts of the seed. Removing ⅓ of the seed from the distal end had no effect on seed germination percentage or seedling vigor. Different DNA extraction protocols were tested to identify a method that could yield DNA of sufficient quality for amplification by polymerase chain reaction. A sodium dodecyl sulfate extraction protocol with 1% polyvinylpyrrolidone yielded DNA that could be amplified with microsatellite primers and was free of pericarp contamination. In this study, an efficient, non-destructive genotyping protocol for watermelon seed was developed.

  15. Comparison of genomic, marker-assisted, and pedigree-BLUP selection methods to increase beta-glucan concentration in elite oat germplasm

    USDA-ARS?s Scientific Manuscript database

    Beta-glucan, a soluble fiber found in oat grain, is good for human health, and selection for higher levels of this compound is regarded as an important breeding objective. Recent advances in oat DNA markers present an opportunity to investigate new selection methods for polygenic traits such as beta...

  16. Height, selected genetic markers and prostate cancer risk: results from the PRACTICAL consortium.

    PubMed

    Lophatananon, Artitaya; Stewart-Brown, Sarah; Kote-Jarai, Zsofia; Olama, Ali Amin Al; Garcia, Sara Benlloch; Neal, David E; Hamdy, Freddie C; Donovan, Jenny L; Giles, Graham G; Fitzgerald, Liesel M; Southey, Melissa C; Pharoah, Paul; Pashayan, Nora; Gronberg, Henrik; Wiklund, Fredrik; Aly, Markus; Stanford, Janet L; Brenner, Hermann; Dieffenbach, Aida K; Arndt, Volker; Park, Jong Y; Lin, Hui-Yi; Sellers, Thomas; Slavov, Chavdar; Kaneva, Radka; Mitev, Vanio; Batra, Jyotsna; Spurdle, Amanda; Clements, Judith A; Easton, Douglas; Eeles, Rosalind A; Muir, Kenneth

    2017-08-22

    Evidence on height and prostate cancer risk is mixed, however, recent studies with large data sets support a possible role for its association with the risk of aggressive prostate cancer. We analysed data from the PRACTICAL consortium consisting of 6207 prostate cancer cases and 6016 controls and a subset of high grade cases (2480 cases). We explored height, polymorphisms in genes related to growth processes as main effects and their possible interactions. The results suggest that height is associated with high-grade prostate cancer risk. Men with height >180 cm are at a 22% increased risk as compared to men with height <173 cm (OR 1.22, 95% CI 1.01-1.48). Genetic variants in the growth pathway gene showed an association with prostate cancer risk. The aggregate scores of the selected variants identified a significantly increased risk of overall prostate cancer and high-grade prostate cancer by 13% and 15%, respectively, in the highest score group as compared to lowest score group. There was no evidence of gene-environment interaction between height and the selected candidate SNPs.Our findings suggest a role of height in high-grade prostate cancer. The effect of genetic variants in the genes related to growth is seen in all cases and high-grade prostate cancer. There is no interaction between these two exposures.

  17. Height, selected genetic markers and prostate cancer risk: results from the PRACTICAL consortium

    PubMed Central

    Lophatananon, Artitaya; Stewart-Brown, Sarah; Kote-Jarai, Zsofia; Olama, Ali Amin Al; Garcia, Sara Benlloch; Neal, David E; Hamdy, Freddie C; Donovan, Jenny L; Giles, Graham G; Fitzgerald, Liesel M; Southey, Melissa C; Pharoah, Paul; Pashayan, Nora; Gronberg, Henrik; Wiklund, Fredrik; Aly, Markus; Stanford, Janet L; Brenner, Hermann; Dieffenbach, Aida K; Arndt, Volker; Park, Jong Y; Lin, Hui-Yi; Sellers, Thomas; Slavov, Chavdar; Kaneva, Radka; Mitev, Vanio; Batra, Jyotsna; Spurdle, Amanda; Clements, Judith A; Schleutker, Johanna; Nordestgaard, BØrge G; Wiklund, Fredrik; Travis, Ruth C; Haiman, Christopher A; Thibodeau, Stephen N; Maier, Christiane; Walther, Vogel; Blot, William J; Kibel, Adam S; Cybulski, Cezary; Cannon-Albright, Lisa; Pandha, Hardev; Teixeira, Manuel R; Cook, Margaret; Govindasami, Koveela; Guy, Michelle; Leongamornlert, Daniel; Sawyer, Emma J; Wilkinson, Rosemary; Morgan, Angela; Fisher, Cyril; Saunders, Edward J; Tymrakiewicz, Malgorzata; Livni, Naomi; Hazel, Steve; Dadaev, Tokhir; Cox, Angela; George, Anne; Lane, Athene; Marsden, Gemma; Davis, Michael; Brown, Paul; Pedersen, John; Hopper, John L; Karlsson, Ami; Cavalli-Bjoerkman, Carin; Adolfson, Jan; Johansson, Jan-Erik; Broms, Michael; Stattin, Paer; Kolb, Suzanne; Stegmaier, Christa; Zachariah, Babu; Park, Hyun; Haley, James; Pow-Sang, Julio; Rincon, Maria; Radlein, Selina; Vlahova, Aleksandrina; Mitkova, Atanaska; Kachakova, Darina; Popov, Elenko; Christova, Svetlana; Dikov, Tihomir; Eckert, Allison; Collins, Angus; Wood, Glenn; Malone, Greg; Alexander, Kimberly; Kerr, Kris; Kedda, Mary-Anne; Turner, Megan; Saunders, Pamela; Heathcote, Peter; Srinivasan, Srilakshmi; Omara, Tracy; Yeadon, Trina; Lose, Felicity; Easton, Douglas; Eeles, Rosalind A; Muir, Kenneth

    2017-01-01

    Background: Evidence on height and prostate cancer risk is mixed, however, recent studies with large data sets support a possible role for its association with the risk of aggressive prostate cancer. Methods: We analysed data from the PRACTICAL consortium consisting of 6207 prostate cancer cases and 6016 controls and a subset of high grade cases (2480 cases). We explored height, polymorphisms in genes related to growth processes as main effects and their possible interactions. Results: The results suggest that height is associated with high-grade prostate cancer risk. Men with height >180 cm are at a 22% increased risk as compared to men with height <173 cm (OR 1.22, 95% CI 1.01–1.48). Genetic variants in the growth pathway gene showed an association with prostate cancer risk. The aggregate scores of the selected variants identified a significantly increased risk of overall prostate cancer and high-grade prostate cancer by 13% and 15%, respectively, in the highest score group as compared to lowest score group. Conclusions: There was no evidence of gene-environment interaction between height and the selected candidate SNPs. Our findings suggest a role of height in high-grade prostate cancer. The effect of genetic variants in the genes related to growth is seen in all cases and high-grade prostate cancer. There is no interaction between these two exposures. PMID:28765617

  18. Microsatellite markers reveal the potential for kin selection on black grouse leks

    PubMed Central

    glund, J. H; Alatalo, R. V.; Lundberg, A.; ki, P. T. Rintam; Lindell, J.

    1999-01-01

    The evolution of social behaviour has puzzled biologists since Darwin. Since Hamilton's theoretical work in the 1960s it has been realized that social behaviour may evolve through the effects of kinship. By helping relatives, an individual may pass on its genes despite negative effects on its own reproduction. Leks are groups of males that females visit primarily to mate. The selective advantage for males to join such social groups has been given much recent attention, but no clear picture has yet emerged. Here we show, using microsatellite analysis, that males but not females of a lekking bird (the black grouse, Tetrao tetrix) are genetically structured at the lek level. We interpret this structuring to be the effects of strong natal philopatry in males. This has the consequence that males on any specific lek should be more related than expected by chance as indicated by our genetic data. Our results thus suggest that kin selection is a factor that needs to be considered in the evolution and maintenance of the lek mating system in black grouse and sheds new light on models of lek evolution.

  19. A Genome-Wide Scan of Selective Sweeps and Association Mapping of Fruit Traits Using Microsatellite Markers in Watermelon

    PubMed Central

    Reddy, Umesh K.; Abburi, Lavanya; Abburi, Venkata Lakshmi; Saminathan, Thangasamy; Cantrell, Robert; Vajja, Venkata Gopinath; Reddy, Rishi; Tomason, Yan R.; Levi, Amnon; Wehner, Todd C.; Nimmakayala, Padma

    2015-01-01

    Our genetic diversity study uses microsatellites of known map position to estimate genome level population structure and linkage disequilibrium, and to identify genomic regions that have undergone selection during watermelon domestication and improvement. Thirty regions that showed evidence of selective sweep were scanned for the presence of candidate genes using the watermelon genome browser (www.icugi.org). We localized selective sweeps in intergenic regions, close to the promoters, and within the exons and introns of various genes. This study provided an evidence of convergent evolution for the presence of diverse ecotypes with special reference to American and European ecotypes. Our search for location of linked markers in the whole-genome draft sequence revealed that BVWS00358, a GA repeat microsatellite, is the GAGA type transcription factor located in the 5′ untranslated regions of a structure and insertion element that expresses a Cys2His2 Zinc finger motif, with presumed biological processes related to chitin response and transcriptional regulation. In addition, BVWS01708, an ATT repeat microsatellite, located in the promoter of a DTW domain-containing protein (Cla002761); and 2 other simple sequence repeats that association mapping link to fruit length and rind thickness. PMID:25425675

  20. Agrobacterium tumefaciens-mediated transformation of Beauveria bassiana using an herbicide resistance gene as a selection marker.

    PubMed

    Fang, Weiguo; Zhang, Yongjun; Yang, Xingyong; Zheng, Xuelian; Duan, Hui; Li, Yi; Pei, Yan

    2004-01-01

    Beauveria bassiana has been investigated for use in the biological control of several insects in agricultural practice. To understand the molecular basis of virulence and host specificity and to improve the entomopathogenicity of B. bassiana, we have developed a simple, highly efficient and reliable Agrobacterium-mediated transformation method for B. bassiana using a phosphinothricin acetyltransferase (bar) gene as a selectable marker. Most transformants contained single copies of T-DNA and the T-DNA inserts were stably inherited after five generations. With this highly efficient transformation method for B. bassiana, we also obtained two putative T-DNA-tagged mutants that may have altered growth habits or virulence. Thus, the described protocol could provide a useful tool to manipulate the genetic make-up and to tag genes that may be important for virulence or growth and development of B. bassiana.

  1. Functional molecular markers (EST-SSR) in the full-sib reciprocal recurrent selection program of maize (Zea mays L.).

    PubMed

    Galvão, K S C; Ramos, H C C; Santos, P H A D; Entringer, G C; Vettorazzi, J C F; Pereira, M G

    2015-07-03

    This study aimed to improve grain yield in the full-sib reciprocal recurrent selection program of maize from the North Fluminense State University. In the current phase of the program, the goal is to maintain, or even increase, the genetic variability within and among populations, in order to increase heterosis of the 13th cycle of reciprocal recurrent selection. Microsatellite expressed sequence tags (EST-SSRs) were used as a tool to assist the maximization step of genetic variability, targeting the functional genome. Eighty S1 progenies of the 13th recur-rent selection cycle, 40 from each population (CIMMYT and Piranão), were analyzed using 20 EST-SSR loci. Genetic diversity, observed heterozygosity, information content of polymorphism, and inbreeding co-efficient were estimated. Subsequently, analysis of genetic dissimilarity, molecular variance, and a graphical dispersion of genotypes were conducted. The number of alleles in the CIMMYT population ranged from 1 to 6, while in the Piranão population the range was from 2 to 8, with a mean of 3.65 and 4.35, respectively. As evidenced by the number of alleles, the Shannon index showed greater diversity for the Piranão population (1.04) in relation to the CIMMYT population (0.89). The genic SSR markers were effective in clustering genotypes into their respective populations before selection and an increase in the variation between populations after selection was observed. The results indicate that the study populations have expressive genetic diversity, which cor-responds to the functional genome, indicating that this strategy may contribute to genetic gain, especially in association with the grain yield of future hybrids.

  2. Genomic selection and complex trait prediction using a fast EM algorithm applied to genome-wide markers.

    PubMed

    Shepherd, Ross K; Meuwissen, Theo H E; Woolliams, John A

    2010-10-22

    The information provided by dense genome-wide markers using high throughput technology is of considerable potential in human disease studies and livestock breeding programs. Genome-wide association studies relate individual single nucleotide polymorphisms (SNP) from dense SNP panels to individual measurements of complex traits, with the underlying assumption being that any association is caused by linkage disequilibrium (LD) between SNP and quantitative trait loci (QTL) affecting the trait. Often SNP are in genomic regions of no trait variation. Whole genome Bayesian models are an effective way of incorporating this and other important prior information into modelling. However a full Bayesian analysis is often not feasible due to the large computational time involved. This article proposes an expectation-maximization (EM) algorithm called emBayesB which allows only a proportion of SNP to be in LD with QTL and incorporates prior information about the distribution of SNP effects. The posterior probability of being in LD with at least one QTL is calculated for each SNP along with estimates of the hyperparameters for the mixture prior. A simulated example of genomic selection from an international workshop is used to demonstrate the features of the EM algorithm. The accuracy of prediction is comparable to a full Bayesian analysis but the EM algorithm is considerably faster. The EM algorithm was accurate in locating QTL which explained more than 1% of the total genetic variation. A computational algorithm for very large SNP panels is described. emBayesB is a fast and accurate EM algorithm for implementing genomic selection and predicting complex traits by mapping QTL in genome-wide dense SNP marker data. Its accuracy is similar to Bayesian methods but it takes only a fraction of the time.

  3. Selective Inducible Nitric Oxide Synthase Inhibitor Reversed Zinc Chloride-Induced Spatial Memory Impairment via Increasing Cholinergic Marker Expression.

    PubMed

    Tabrizian, Kaveh; Azami, Kian; Belaran, Maryam; Soodi, Maliheh; Abdi, Khosrou; Fanoudi, Sahar; Sanati, Mehdi; Mottaghi Dastjerdi, Negar; Soltany Rezaee-Rad, Mohammad; Sharifzadeh, Mohammad

    2016-10-01

    Zinc, an essential micronutrient and biochemical element of the human body, plays structural, catalytic, and regulatory roles in numerous physiological functions. In the current study, the effects of a pretraining oral administration of zinc chloride (10, 25, and 50 mg/kg) for 14 consecutive days and post-training bilateral intra-hippocampal infusion of 1400W as a selective inducible nitric oxide synthase (iNOS) inhibitor (10, 50, and 100 μM/side), alone and in combination, on the spatial memory retention in Morris water maze (MWM) were investigated. Animals were trained for 4 days and tested 48 h after completion of training. Also, the molecular effects of these compounds on the expression of choline acetyltransferase (ChAT), as a cholinergic marker in the CA1 region of the hippocampus and medial septal area (MSA), were evaluated. Behavioral and molecular findings of this study showed that a 2-week oral administration of zinc chloride (50 mg/kg) impaired spatial memory retention in MWM and decreased ChAT expression. Immunohistochemical analysis of post-training bilateral intra-hippocampal infusion of 1400W revealed a significant increase in ChAT immunoreactivity. Furthermore, post-training bilateral intra-hippocampal infusion of 1400W into the CA1 region of the hippocampus reversed zinc chloride-induced spatial memory impairment in MWM and significantly increased ChAT expression in comparison with zinc chloride-treated animals. Taken together, these results emphasize the role of selective iNOS inhibitors in reversing zinc chloride-induced spatial memory deficits via modulation of cholinergic marker expression.

  4. An improved consensus linkage map of barley based on flow-sorted chromosomes and SNP markers

    USDA-ARS?s Scientific Manuscript database

    Recent advances in high-throughput genotyping have made it easier to combine information from different mapping populations into consensus genetic maps, which provide increased marker density and genome coverage compared to individual maps. Previously, a SNP-based genotyping platform was developed a...

  5. A Method of Rating Assimilation in Psychotherapy Based on Markers of Change

    ERIC Educational Resources Information Center

    Honos-Webb, Lara; Stiles, William B.; Greenberg, Leslie S

    2003-01-01

    This article reports the development of a marker-based method for rating assimilation in psychotherapy with a case study of a woman, Sarah, who was treated for depression with process-experiential psychotherapy. To increase the rigor of the qualitative case study methodology, independent judges rated passages--taken out of context and randomly…

  6. An annotated genetic map of loblolly pine based on microsatellite and cDNA markers

    Treesearch

    Craig S. Echt; Surya Saha; Konstantin V. Krutovsky; Kokulapalan Wimalanathan; John E. Erpelding; Chun Liang; C Dana Nelson

    2011-01-01

    Previous loblolly pine (Pinus taeda L.) genetic linkage maps have been based on a variety of DNA polymorphisms, such as AFLPs, RAPDs, RFLPs, and ESTPs, but only a few SSRs (simple sequence repeats), also known as simple tandem repeats or microsatellites, have been mapped in P. taeda. The objective of this study was to integrate a large set of SSR markers from a variety...

  7. How biomarkers will change psychiatry. Part II: Biomarker selection and potential inflammatory markers of depression.

    PubMed

    Macaluso, Matthew; Drevets, Wayne C; Preskorn, Sheldon H

    2012-07-01

    Part I of this series defined biomarkers and discussed their current use in general medicine and their potential research and clinical utility in psychiatry. In this second column in the series, the authors first discuss the rationale for selecting a biomarker. The second half of the column discusses the potential use of inflammatory biomarkers in depression, with a specific focus on derivatives of the inflammatory biomarker, thromboxane, to illustrate how biomarkers can be developed for use in clinical practice. In the future, biomarkers are likely to become an integral component of psychiatric treatment, providing information about a patient's odds of developing an illness, the severity of illness, and level of response to therapeutic interventions.

  8. Transformation of diatom Phaeodactylum tricornutum by electroporation and establishment of inducible selection marker.

    PubMed

    Niu, Ying-Fang; Yang, Zhi-Kai; Zhang, Meng-Han; Zhu, Cong-Cong; Yang, Wei-Dong; Liu, Jie-Sheng; Li, Hong-Ye

    2012-06-01

    Diatoms are important primary producers in the marine ecosystem. Currently it is difficult to genetically transform diatoms due to the technical limitations of existing methods. The promoter/terminator of the nitrate reductase gene of the model diatom Phaeodactylum tricornutum was cloned and used to drive chloramphenicol acetyltransferase (CAT) reporter gene expression. The construct was transferred by electroporation into P. tricornutum grown in medium lacking silicon. CAT expression was induced in transformed diatoms in the presence of nitrate, enabling growth in selective medium, and was repressed when ammonium was the only nitrogen source. Expression of CAT transcript and protein were demonstrated by RT-PCR and Western blot analysis, respectively. Our study is the first to report a successful genetic transformation of diatom by electroporation in an economical and efficient manner and provides a tightly regulated inducible gene expression system for diatom.

  9. Selection and management of DNA markers for use in genomic evaluation.

    PubMed

    Wiggans, G R; VanRaden, P M; Bacheller, L R; Tooker, M E; Hutchison, J L; Cooper, T A; Sonstegard, T S

    2010-05-01

    To facilitate routine genomic evaluation, a database was constructed to store genotypes for 50,972 single nucleotide polymorphisms (SNP) from the Illumina BovineSNP50 BeadChip (Illumina Inc., San Diego, CA). Multiple samples per animal are allowed. All SNP genotypes for a sample are stored in a single row. An indicator specifies whether the genotype for a sample was selected for use in genomic evaluation. Samples with low call rates or pedigree conflicts are designated as unusable. Among multiple samples that qualify for use in genomic evaluation, the one with the highest call rate is designated as usable. When multiple samples are stored for an animal, a composite is formed during extraction by using SNP genotypes from other samples to replace missing genotypes. To increase the number of SNP available, scanner output for approximately 19,000 samples was reprocessed. Any SNP with a minor allele frequency of > or = 1% for Holsteins, Jerseys, or Brown Swiss was selected, which was the primary reason that the number of SNP used for USDA genomic evaluations increased. Few parent-progeny conflicts (< or = 1%) and a high call rate (> or = 90%) were additional requirements that eliminated 2,378 SNP. Because monomorphic SNP did not degrade convergence during estimation of SNP effects, a single set of 43,385 SNP was adopted for all breeds. The use of a database for genotypes, detection of conflicts as genotypes are stored, online access for problem resolution, and use of a single set of SNP for genomic evaluations have simplified tracking of genotypes and genomic evaluation as a routine and official process. Copyright 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  10. Genetic diversity in wild species of passion fruit (Passiflora trintae) based on molecular markers.

    PubMed

    Cerqueira-Silva, C B M; Cardoso-Silva, C B; Santos, E S L; Conceição, L D H C S; Pereira, A S; Oliveira, A C; Corrêa, R X

    2010-10-26

    In spite of the importance of and the considerable variability observed in Passiflora (Passifloraceae), little is known about the genetic diversity of most of the species of this genus. We evaluated the genetic diversity by RAPD markers in 18 genotypes of Passiflora trintae. The 15 primers generated 112 markers, 84% of which were polymorphic. The genetic distance estimated by the complement of the Dice index (average dissimilarity = 0.30) and genotype grouping based on the UPGMA algorithm showed low variability among genotypes. More attention should be given to the study and conservation of the biodiversity of this economically important genus.

  11. Analysis of genetic diversity of certain species of Piper using RAPD-based molecular markers.

    PubMed

    Chowdhury, Utpal; Tanti, Bhaben; Rethy, Parakkal; Gajurel, Padma Raj

    2014-09-01

    The utility of RAPD markers in assessing genetic diversity and phenetic relationships of six different species of Piper from Northeast India was investigated. Polymerase chain reaction (PCR) with four arbitrary 10-mer oligonucleotide primers applied to the six species produced a total of 195 marker bands, of which, 159 were polymorphic. On average, six RAPD fragments were amplified per reaction. In the UPGMA phenetic dendrogram based on Jaccard's coefficient, the different accessions of Piper showed a high level of genetic variation. This study may be useful in identifying diverse genetic stocks of Piper, which may then be conserved on a priority basis.

  12. Fingerprint signatures based on nanomagnets as markers in materials for tracing and counterfeit protection

    NASA Astrophysics Data System (ADS)

    Szielasko, Klaus; Youssef, Amir; Sporn, Dieter; Mandel, Karl

    2016-05-01

    Identification and tracing of objects is a challenge of great current importance. Ideally, this can be achieved via small markers, which equip an object with a fingerprint identity. Herein, we communicate an approach that is based on structural arrangements of magnetic nano-entities to generate a kind of security key in an object. The nano-markers, which are applied as inks, can remotely be read out in a fast and simple way with a GMR sensor and yield an unmistakable signal pattern.

  13. Genetic characterization of Red Junglefowl (Gallus gallus), Thai indigenous chicken (Gallus domesticus), and two commercial lines using selective functional genes compared to microsatellite markers.

    PubMed

    Akaboot, P; Duangjinda, M; Phasuk, Y; Kaenchan, C; Chinchiyanond, W

    2012-07-19

    Genetic characterization among Red Junglefowl (GS, Gallus gallus spadiceus), Thai indigenous chicken (TIC, Gallus domesticus) and commercial lines has been widely used for studies of genealogical origin, genetic diversity, and effects of selection. We compared the efficiency of genetic characterization of chicken populations that had been under different intensities of selection using selective functional gene versus microsatellite marker analyses. We genotyped 151 chickens from five populations: Red Junglefowl, TIC and commercial lines (BR, broiler and WL, White Leghorn). Genetic structure analyses using six loci of five functional genes - corresponding to heat tolerance (heat shock protein 70, HSP70/C, HSP70/M), broodiness (vasoactive intestinal peptide receptor-1, VIPR-1), egg production-[24-bp indel (insertion or deletion) prolactin, 24bpPRL], ovulation rate (growth hormone receptor, GHR), and growth (insulin-like growth factor-1, IGF-1) - were compared with 18 microsatellite markers. PCR-RFLP and allele specific PCR were used for functional gene typing. A neighbor-joining tree from Nei's genetic distance was constructed to show genetic relationships. A similar pattern was found with both functional genes and microsatellites. Three groups consisting of BR, WL and TIC-GS-GG were formed. A principal component plot based on individual similarity using Dice's coefficient was also constructed to confirm the relationship. Different patterns were found when using functional genes versus microsatellites. A principal component plot with functional genes also gave three clusters consisting of BR, WL and TIC-GS-GG. A principal component plot using microsatellites gave four clusters, consisting of WL, GG, TIC, and BR-GS. Characterization of BR and GS differs from previous studies. We concluded that genetic characterization with appropriate functional genes is more accurate when differences in genetic make-up among populations are known. Genetic characterization using

  14. A Bio-Economic Case Study of Canadian Honey Bee (Hymenoptera: Apidae) Colonies: Marker-Assisted Selection (MAS) in Queen Breeding Affects Beekeeper Profits

    PubMed Central

    Baylis, Kathy; Hoover, Shelley E.; Currie, Rob W.; Melathopoulos, Andony P.; Pernal, Stephen F.; Foster, Leonard J.; Guarna, M. Marta

    2017-01-01

    Abstract Over the past decade in North America and Europe, winter losses of honey bee (Hymenoptera: Apidae) colonies have increased dramatically. Scientific consensus attributes these losses to multifactorial causes including altered parasite and pathogen profiles, lack of proper nutrition due to agricultural monocultures, exposure to pesticides, management, and weather. One method to reduce colony loss and increase productivity is through selective breeding of queens to produce disease-, pathogen-, and mite-resistant stock. Historically, the only method for identifying desirable traits in honey bees to improve breeding was through observation of bee behavior. A team of Canadian scientists have recently identified markers in bee antennae that correspond to behavioral traits in bees and can be tested for in a laboratory. These scientists have demonstrated that this marker-assisted selection (MAS) can be used to produce hygienic, pathogen-resistant honey bee colonies. Based on this research, we present a beekeeping case study where a beekeeper’s profit function is used to evaluate the economic impact of adopting colonies selected for hygienic behavior using MAS into an apiary. Our results show a net profit gain from an MAS colony of between 2% and 5% when Varroa mites are effectively treated. In the case of ineffective treatment, MAS generates a net profit benefit of between 9% and 96% depending on the Varroa load. When a Varroa mite population has developed some treatment resistance, we show that MAS colonies generate a net profit gain of between 8% and 112% depending on the Varroa load and degree of treatment resistance. PMID:28334400

  15. A Multi-Marker Genetic Association Test Based on the Rasch Model Applied to Alzheimer’s Disease

    PubMed Central

    Wang, Wenjia; Mandel, Jonas; Bouaziz, Jan; Commenges, Daniel; Nabirotchkine, Serguei; Chumakov, Ilya; Cohen, Daniel; Guedj, Mickaël

    2015-01-01

    Results from Genome-Wide Association Studies (GWAS) have shown that the genetic basis of complex traits often include many genetic variants with small to moderate effects whose identification remains a challenging problem. In this context multi-marker analysis at the gene and pathway level can complement traditional point-wise approaches that treat the genetic markers individually. In this paper we propose a novel statistical approach for multi-marker analysis based on the Rasch model. The method summarizes the categorical genotypes of SNPs by a generalized logistic function into a genetic score that can be used for association analysis. Through different sets of simulations, the false-positive rate and power of the proposed approach are compared to a set of existing methods, and shows good performances. The application of the Rasch model on Alzheimer’s Disease (AD) ADNI GWAS dataset also allows a coherent interpretation of the results. Our analysis supports the idea that APOE is a major susceptibility gene for AD. In the top genes selected by proposed method, several could be functionally linked to AD. In particular, a pathway analysis of these genes also highlights the metabolism of cholesterol, that is known to play a key role in AD pathogenesis. Interestingly, many of these top genes can be integrated in a hypothetic signalling network. PMID:26379234

  16. Covariance based outlier detection with feature selection.

    PubMed

    Zwilling, Chris E; Wang, Michelle Y

    2016-08-01

    The present covariance based outlier detection algorithm selects from a candidate set of feature vectors that are best at identifying outliers. Features extracted from biomedical and health informatics data can be more informative in disease assessment and there are no restrictions on the nature and number of features that can be tested. But an important challenge for an algorithm operating on a set of features is for it to winnow the effective features from the ineffective ones. The powerful algorithm described in this paper leverages covariance information from the time series data to identify features with the highest sensitivity for outlier identification. Empirical results demonstrate the efficacy of the method.

  17. Selective isolation of nitrogen bases from petroleum

    SciTech Connect

    Schmitter, J.M.; Ignatiadis, I.; Arpino, P.; Guiochon, G.

    1983-09-01

    A method has been designed for the rapid separation of nitrogen bases from petroleum by trapping their quaternary ammonium salts on hydrochloric acid treated silica. A chromatographic column filled with this adsorbent and equipped with a solvent recycling system allows a selective and quantitative extraction of basic compounds from large amounts (>100 g) of crude oil. The procedure has been tested with reference compounds, a coker gas-oil, and a heavy biodegraded crude and applied to 15 different oil samples. 22 references, 3 figures, 4 tables.

  18. Genetic variability in wild genotypes of Passiflora cincinnata based on RAPD markers.

    PubMed

    Cerqueira-Silva, C B M; Conceição, L D H C S; Santos, E S L; Cardoso-Silva, C B; Pereira, A S; Oliveira, A C; Corrêa, R X

    2010-12-21

    The genetic diversity and characteristics of commercial interest of Passiflora species make it useful to characterize wild germplasm, because of their potential use for fruit, ornamental and medicinal purposes. We evaluated genetic diversity, using RAPD markers, of 32 genotypes of Passiflora cincinnata collected from the wild in the region of Vitória da Conquista, Bahia, Brazil. Thirteen primers generated 95 polymorphic markers and only one monomorphic marker. The mean genetic distance between the genotypes estimated by the complement of the Dice index was 0.51 (ranging from 0.20-0.85), and genotype grouping based on the UPGMA algorithm showed wide variability among the genotypes. This type of information contributes to identification and conservation of the biodiversity of this species and for the identification of pairs of divergent individuals for maximum exploitation of existing variability.

  19. Mach-zehnder based optical marker/comb generator for streak camera calibration

    DOEpatents

    Miller, Edward Kirk

    2015-03-03

    This disclosure is directed to a method and apparatus for generating marker and comb indicia in an optical environment using a Mach-Zehnder (M-Z) modulator. High speed recording devices are configured to record image or other data defining a high speed event. To calibrate and establish time reference, the markers or combs are indicia which serve as timing pulses (markers) or a constant-frequency train of optical pulses (comb) to be imaged on a streak camera for accurate time based calibration and time reference. The system includes a camera, an optic signal generator which provides an optic signal to an M-Z modulator and biasing and modulation signal generators configured to provide input to the M-Z modulator. An optical reference signal is provided to the M-Z modulator. The M-Z modulator modulates the reference signal to a higher frequency optical signal which is output through a fiber coupled link to the streak camera.

  20. The first genetic linkage map of Primulina eburnea (Gesneriaceae) based on EST-derived SNP markers.

    PubMed

    Feng, Chen; Feng, Chao; Kang, Ming

    2016-06-01

    Primulina eburnea is a promising candidate for domestication and floriculture, since it is easy to culture and has beautiful flowers. An F₂ population of 189 individuals was established for the construction of first-generation linkage maps based on expressed sequence tags-derived single-nucleotide polymorphism markers using the massARRAY genotyping platform. Of the 232 screened markers, 215 were assigned to 18 LG according to the haploid number of chromosomes in the species. The linkage map spanned a total of 3774.7 cM with an average distance of 17.6 cM between adjacent markers. This linkage map provides a framework for identification of important genes in breeding programmes.

  1. Population structure and admixture in Cerro Largo, Uruguay, based on blood markers and mitochondrial DNA polymorphisms.

    PubMed

    Sans, Mónica; Merriwether, D Andrew; Hidalgo, Pedro C; Bentancor, Nilo; Weimer, Tania A; Franco, Maria Helena L P; Alvarez, Inés; Kemp, Brian M; Salzano, Francisco M

    2006-01-01

    Recent studies of the Uruguayan population revealed different amounts of Amerindian and African genetic contributions. Our previous analysis of Afro-Uruguayans from the capital city of the Department of Cerro Largo showed a high proportion of African genes, and the effects of directional mating involving Amerindian women. In this paper, we extended the analysis to a sample of more than 100 individuals representing a random sample of the population of the whole Department. Based on 18 autosomal markers and one X-linked marker, we estimated 82% European, 8% Amerindian, and 10% African contributions to their ancestry, while from seven mitochondrial DNA site-specific polymorphic markers and sequences of hypervariable segment I, we determined 49% European, 30% Amerindian, and 21% African maternal contributions. Directional matings between Amerindian women and European men were detected, but differences involving Africans were not significant. Data about the specific origins of maternal lineages were also provided, and placed in a historical context.

  2. Marker based watershed to segment dim infrared target through image enhancement

    NASA Astrophysics Data System (ADS)

    Zhou, Fugen; Bai, Xiangzhi; Xie, Yongchun; Jin, Ting

    2008-10-01

    A novel marker based watershed through image enhancement is proposed to segment the dim infrared target. The dim infrared target is firstly enhanced by CB top-hat transformation and image quantization. Then, the accurate marker of the target can be easily obtained through image binarisation and marker filtering. To calculate an efficient gradient image of the dim target for the watershed segmentation, the gradient image is firstly calculated through Sobel operator and then efficiently enhanced through pseudo top-hat transformation and gradient quantization. Because of the enhancement of the dim target and the gradient image, the watershed can efficiently segment the dim infrared image. Experimental results show that the proposed algorithm is much efficient for dim infrared target segmentation.

  3. Stimulator Selection in SSVEP-Based Spatial Selective Attention Study

    PubMed Central

    Liu, Chang; Zhu, Fangshi; Wang, Linan; Xie, Xinzhou; Wang, Wei

    2016-01-01

    Steady-State Visual Evoked Potentials (SSVEPs) are widely used in spatial selective attention. In this process the two kinds of visual simulators, Light Emitting Diode (LED) and Liquid Crystal Display (LCD), are commonly used to evoke SSVEP. In this paper, the differences of SSVEP caused by these two stimulators in the study of spatial selective attention were investigated. Results indicated that LED could stimulate strong SSVEP component on occipital lobe, and the frequency of evoked SSVEP had high precision and wide range as compared to LCD. Moreover a significant difference between noticed and unnoticed frequencies in spectrum was observed whereas in LCD mode this difference was limited and selectable frequencies were also limited. Our experimental finding suggested that average classification accuracies among all the test subjects in our experiments were 0.938 and 0.853 in LED and LCD mode, respectively. These results indicate that LED simulator is appropriate for evoking the SSVEP for the study of spatial selective attention. PMID:28044073

  4. Single Marker and Haplotype-Based Association Analysis of Semolina and Pasta Colour in Elite Durum Wheat Breeding Lines Using a High-Density Consensus Map

    PubMed Central

    Haile, Jemanesh K.; Cory, Aron T.; Clarke, Fran R.; Clarke, John M.; Knox, Ron E.; Pozniak, Curtis J.

    2017-01-01

    Association mapping is usually performed by testing the correlation between a single marker and phenotypes. However, because patterns of variation within genomes are inherited as blocks, clustering markers into haplotypes for genome-wide scans could be a worthwhile approach to improve statistical power to detect associations. The availability of high-density molecular data allows the possibility to assess the potential of both approaches to identify marker-trait associations in durum wheat. In the present study, we used single marker- and haplotype-based approaches to identify loci associated with semolina and pasta colour in durum wheat, the main objective being to evaluate the potential benefits of haplotype-based analysis for identifying quantitative trait loci. One hundred sixty-nine durum lines were genotyped using the Illumina 90K Infinium iSelect assay, and 12,234 polymorphic single nucleotide polymorphism (SNP) markers were generated and used to assess the population structure and the linkage disequilibrium (LD) patterns. A total of 8,581 SNPs previously localized to a high-density consensus map were clustered into 406 haplotype blocks based on the average LD distance of 5.3 cM. Combining multiple SNPs into haplotype blocks increased the average polymorphism information content (PIC) from 0.27 per SNP to 0.50 per haplotype. The haplotype-based analysis identified 12 loci associated with grain pigment colour traits, including the five loci identified by the single marker-based analysis. Furthermore, the haplotype-based analysis resulted in an increase of the phenotypic variance explained (50.4% on average) and the allelic effect (33.7% on average) when compared to single marker analysis. The presence of multiple allelic combinations within each haplotype locus offers potential for screening the most favorable haplotype series and may facilitate marker-assisted selection of grain pigment colour in durum wheat. These results suggest a benefit of haplotype-based

  5. Analysis of genetic diversity among selected grasspea (Lathyrus sativus L.) genotypes using RAPD markers.

    PubMed

    Barik, Durga P; Acharya, Laxmikanta; Mukherjee, Arup K; Chand, Pradeep K

    2007-01-01

    Randomly amplified polymorphic DNA (RAPD) technique was applied to assess the genetic variability among five selected genotypes of grasspea. Out of 30 random decamer primers tested for the present investigation 20 showed reproducible DNA amplification. A total of 257 loci were amplified of which 159 were polymorphic including 57 genotype-specific unique bands. Amplicons had molecular weights ranging from 3.0 kb to 0.1 kb. Majority amplicons were shared by most of the genotypes which indicated a very narrow genetic gap between them. The dendrogram constructed on the basis of RAPD data showed two clusters. The local genotype collected from Nayagarh was grouped along with IC-120451 and IC-120453, sharing a common node at an 82% similarity level. The other genotypes, IC-120478 and IC-120487, were located in the second clade having a common node at 84% similarity level. The investigation showed that though all the genotypes of grasspea were of apparently similar morphology there exists polymorphism at the molecular level, which can be exploited in breeding programmes aimed at crop improvement.

  6. Selective Nitration of Tyr(99) in Calmodulin as a Marker of Cellular Conditions of Oxidative Stress

    SciTech Connect

    Smallwood, Heather S. ); Galeva, Nadezhda A.; Bartlett, Ryan K.; Urbauer, Ramona J.; Williams, Todd D.; Urbauer, Jeffrey L.; Squier, Thomas C. )

    2003-01-01

    We examined the possible role of methionines as oxidant scavengers that prevent the peroxynitrite-induced nitration of tyrosines within calmodulin (CaM). We used mass spectrometry to investigate the reactivity of peroxynitrite with CaM at physiological pH. The possible role of methionines in scavenging peroxynitrite(ONOO-)was assessed in wild-type CaM and following substitution of all nine methionines in CaM with leucines. We find that peroxynitrite selectively nitrates Tyr-99 at physiological pH resulting in the formation of between 0.05 and 0.25 mol of nitrotyrosine/mol of CaM when the added molar ratio of peroxynitrite per CaM was varied between 2.5 and 15. In wild-type CaM there is a corresponding oxidation of between 0.8 and 2.8 mol of methionine to form methionine sulfoxide. However, following site-directed substitution of all nine methionines in wild-type CaM with leucines, the extent of nitration by peroxynitrite was unchanged. These results indicate that Tyr-99 is readily nitrated by perioxynitrite and that methionine side chains do not function as an antioxidant in scavenging perioxynitrite. Thus, separate reactive species are involved in the oxidation of

  7. Innate Obesity, Revealed by Selection Markers, Confers Significant Imprint of Hypothalamic Genes Controlling Energy Expenditure.

    PubMed

    Henry, Belinda A

    2017-01-01

    The incidence of obesity is rapidly escalating and has reached epidemic proportions. In all species, including rodents, humans, and sheep, there is large variation in the degree of weight gain across individuals in response to an obesogenic environment. This individual variation is, at least in part, determined by innate differences in energy expenditure, of which adaptive thermogenesis is a key component. The hypothalamus is essential to the control of body weight and adiposity. Appetite-regulating peptides within the hypothalamus exert reciprocal effects on food intake and energy expenditure, such that neuropeptides that stimulate food intake inhibit thermogenesis and vice versa. This review discusses the role of the hypothalamic neuropeptides in determining innate predisposition to obesity in 3 animal models being obesity-prone and obesity-resistant rodents, genetically lean and obese sheep, and animals selected for high/low cortisol responsiveness. In rodents, leptin resistance is a primary feature of the propensity to become obese. This contrasts that of larger mammals, such as sheep, where altered susceptibility to obesity manifests within the melanocortin and/or orexin pathways. This review highlights fundamental species differences within the hypothalamus that lead to altered susceptibility to weight gain and increased propensity to become obese. © 2016 S. Karger AG, Basel.

  8. Recombineering reagents for improved inducible expression and selection marker re-use in Schizosaccharomyces pombe.

    PubMed

    Erler, Axel; Maresca, Marcello; Fu, Jun; Stewart, A Francis

    2006-08-01

    The fission yeast Schizosaccharomyces pombe is an excellent model organism for cell biology. However, its genetic toolbox is less developed than that of Saccharomyces cerevisiae. In the first part of this study we describe an improved inducible expression vector based on tetracycline regulation of the CaMV35S promoter, which is also capable of chromosomal integration and therefore works in minimal and in rich media. We found that anhydrotetracycline is a superior ligand for induction. Maximum expression levels were observed after 12 h in minimal media (EMM) and after 9 h in rich media (YES), which is faster than the nmt1 promoter system. The system was combined with a convenient recombineering-based subcloning strategy for ease of cloning. In the second part we present four template plasmids, pSVEM-bsd, pSVEM-nat, pSVEM-kan and pSVEM-hph, which harbour four recyclable disruption cassettes based on the Cre recombinase lox71/66 strategy for use in PCR targeting methods. Cre-mediated excision leaves a non-functional mutant lox site in the genome, allowing the reiterative usage of these cassettes for multiple targetings. These cassettes are also configured with dual eukaryotic/prokaryotic promoters so that they can be used for recombineering in E. coli. Amongst other purposes, this permits the rapid and convenient creation of targeting constructs with much longer homology arms for difficult and complex targetings in the Sz. pombe genome.

  9. Use of theragnostic markers to select drugs for phase II/III trials for Alzheimer disease

    PubMed Central

    2010-01-01

    In a slowly progressive disorder like Alzheimer disease, evaluation of the clinical effect of novel drug candidates requires large numbers of patients and extended treatment periods. Current cell- and animal-based disease models of Alzheimer disease are poor at predicting a positive treatment response in patients. To help bridge the gap between disease models and large and costly clinical trials with high failure rates, biomarkers for the intended biochemical drug effect may be of value. Such biomarkers may be called 'theragnostic'. Here, we review the literature addressing the prospective value of these biomarkers. PMID:21122172

  10. Genetic diversity analysis of sweet kernel apricot in China based on SSR and ISSR markers.

    PubMed

    Liu, M P; Du, H Y; Zhu, G P; Fu, D L; Tana, W Y

    2015-08-19

    Simple sequence repeat (SSR) and inter-simple sequence repeat (ISSR) markers were used to evaluate genetic diversity among 22 sweet kernel apricot accessions and 12 cultivars in China to provide information on how to improve the utilization of kernel apricot germplasms. The results showed that 10 pairs of SSR primers screened from 40 primer pairs amplified 43 allelic variants, all of which were polymorphic (100%), and 9 ISSR primers selected from 100 primers amplified 67 allelic variants with 50 polymorphic bands (74.63%). There was a relatively distant genetic relationship between the 34 samples, where their genetic similarity coefficient was between 0.62 and 0.99. The UPGMA dendrogram constructed using combined data of the two marker systems separated the genotypes into three main clusters.

  11. Identification of Phoenix dactylifera L. varieties based on amplified fragment length polymorphism (AFLP) markers.

    PubMed

    Diaz, Susana; Pire, Carmen; Ferrer, Juan; Bonete, Maria José

    2003-01-01

    The amplified fragment length polymorphism (AFLP) technique was applied to identify palm varieties. Fluorescence labelled primers were used in selective amplifications and the amplified fragments were detected on capillary gel electrophoresis using an automated DNA sequencer with the analysis fragment option. This is a rapid and efficient technique for detecting a large number of DNA markers on the date palm. Phoenix dactylifera L. varieties Bou-Fegous, Medjool, and E-528 from Estación Phoenix (Elche), Spain, were analysed, yielding a total of 310 AFLP fragments derived from five primer combinations. The process for regenerating the date palm cultivars from in vitro tissue culture should yield individuals phenotypically and genetically identical to the explant they are derived from. The AFLP markers obtained were successfully used for comparing and identifying vitroplants of palm.

  12. Expression analysis onto microarrays of randomly selected cDNA clones highlights HOXB13 as a marker of human prostate cancer

    PubMed Central

    Edwards, S; Campbell, C; Flohr, P; Shipley, J; Giddings, I; te-Poele, R; Dodson, A; Foster, C; Clark, J; Jhavar, S; Kovacs, G; Cooper, C S

    2004-01-01

    In a strategy aimed at identifying novel markers of human prostate cancer, we performed expression analysis using microarrays of clones randomly selected from a cDNA library prepared from the LNCaP prostate cancer cell line. Comparisons of expression profiles in primary human prostate cancer, adjacent normal prostate tissue, and a selection of other (nonprostate) normal human tissues, led to the identification of a set of clones that were judged as the best candidate markers of normal and/or malignant prostate tissue. DNA sequencing of the selected clones revealed that they included 10 genes that had previously been established as prostate markers: NKX3.1, KLK2, KLK3 (PSA), FOLH1 (PSMA), STEAP2, PSGR, PRAC, RDH11, Prostein and FASN. Following analysis of the expression patterns of all selected and sequenced genes through interrogation of SAGE databases, a further three genes from our clone set, HOXB13, SPON2 and NCAM2, emerged as additional candidate markers of human prostate cancer. Quantitative RT–PCR demonstrated the specificity of expression of HOXB13 in prostate tissue and revealed its ubiquitous expression in a series of 37 primary prostate cancers and 20 normal prostates. These results demonstrate the utility of this expression-microarray approach in hunting for new markers of individual human cancer types. PMID:15583692

  13. Expression analysis onto microarrays of randomly selected cDNA clones highlights HOXB13 as a marker of human prostate cancer.

    PubMed

    Edwards, S; Campbell, C; Flohr, P; Shipley, J; Giddings, I; Te-Poele, R; Dodson, A; Foster, C; Clark, J; Jhavar, S; Kovacs, G; Cooper, C S

    2005-01-31

    In a strategy aimed at identifying novel markers of human prostate cancer, we performed expression analysis using microarrays of clones randomly selected from a cDNA library prepared from the LNCaP prostate cancer cell line. Comparisons of expression profiles in primary human prostate cancer, adjacent normal prostate tissue, and a selection of other (nonprostate) normal human tissues, led to the identification of a set of clones that were judged as the best candidate markers of normal and/or malignant prostate tissue. DNA sequencing of the selected clones revealed that they included 10 genes that had previously been established as prostate markers: NKX3.1, KLK2, KLK3 (PSA), FOLH1 (PSMA), STEAP2, PSGR, PRAC, RDH11, Prostein and FASN. Following analysis of the expression patterns of all selected and sequenced genes through interrogation of SAGE databases, a further three genes from our clone set, HOXB13, SPON2 and NCAM2, emerged as additional candidate markers of human prostate cancer. Quantitative RT-PCR demonstrated the specificity of expression of HOXB13 in prostate tissue and revealed its ubiquitous expression in a series of 37 primary prostate cancers and 20 normal prostates. These results demonstrate the utility of this expression-microarray approach in hunting for new markers of individual human cancer types.

  14. A new metabolomics-based strategy for identification of endogenous markers of urine adulteration attempts exemplified for potassium nitrite.

    PubMed

    Steuer, Andrea E; Arnold, Kim; Schneider, Tom D; Poetzsch, Michael; Kraemer, Thomas

    2017-08-16

    Urine adulteration to circumvent positive drug testing represents a problem for toxicological laboratories. While creatinine is a suitable marker for dilution, detection of chemicals is often performed by dipstick tests associated with high rates of false positives. Several methods would be necessary to check for all possible adulterants. Untargeted mass spectrometry (MS) methods used in metabolomics should theoretically allow detecting concentration changes of any endogenous urinary metabolite or presence of new biomarkers produced by chemical adulteration. As a proof of concept study, urine samples from 10 volunteers were treated with KNO2 and analyzed by high-resolution MS. For statistical data evaluation, XCMS(plus) and MetaboAnalyst were used. Compound identification was performed by database searches using an in-house database, Chemspider, METLIN, HMDB, and NIST. Principle component analysis revealed clear separation between treated and untreated urine samples. In detail, 307 features showed significant concentration changes with fold changes greater than 2 (79 decreased; 228 increased). Mainly amino acids (e.g., histidine, methylhistidine, di- and trimethyllysine) and purines (uric acid) were detected in lower amounts. 5-HO-isourate was found to be formed as a new compound from uric acid and, e.g., imidazole lactate concentrations increased due to the breakdown of histidine. This metabolomics-based strategy allowed for a broad identification range of markers of urinary adulteration. More studies will be needed to investigate routine applicability of identified potential markers exploring urinary conditions of their formation and stability. Selected markers might then be integrated into routine MS screening procedures allowing for detection of adulteration within routine MS analysis. Graphical Abstract ᅟ.

  15. Codominant PCR-based markers and candidate genes for powdery mildew resistance in melon (Cucumis melo L.).

    PubMed

    Yuste-Lisbona, Fernando J; Capel, Carmen; Gómez-Guillamón, María L; Capel, Juan; López-Sesé, Ana I; Lozano, Rafael

    2011-03-01

    Powdery mildew caused by Podosphaera xanthii is a major disease in melon crops, and races 1, 2, and 5 of this fungus are those that occur most frequently in southern Europe. The genotype TGR-1551 bears a dominant gene that provides resistance to these three races of P. xanthii. By combining bulked segregant analysis and amplified fragment length polymorphisms (AFLP), we identified eight markers linked to this dominant gene. Cloning and sequencing of the selected AFLP fragments allowed the development of six codominant PCR-based markers which mapped on the linkage group (LG) V. Sequence analysis of these markers led to the identification of two resistance-like genes, MRGH5 and MRGH63, belonging to the nucleotide binding site (NBS)-leucine-rich repeat (LRR) gene family. Quantitative trait loci (QTL) analysis detected two QTLs, Pm-R1-2 and Pm-R5, the former significantly associated with the resistance to races 1 and 2 (LOD score of 26.5 and 33.3; 53.6 and 61.9% of phenotypic variation, respectively), and the latter with resistance to race 5 (LOD score of 36.8; 65.5% of phenotypic variation), which have been found to be colocalized with the MRGH5 and MRGH63 genes, respectively. The results suggest that the cluster of NBS-LRR genes identified in LG V harbours candidate genes for resistance to races 1, 2, and 5 of P. xanthii. The evaluation of other resistant germplasm showed that the codominant markers here reported are also linked to the Pm-w resistance gene carried by the accession 'WMR-29' proving their usefulness as genotyping tools in melon breeding programmes.

  16. Predictive ability of direct genomic values for lifetime net merit of Holstein sires using selected subsets of single nucleotide polymorphism markers.

    PubMed

    Weigel, K A; de los Campos, G; González-Recio, O; Naya, H; Wu, X L; Long, N; Rosa, G J M; Gianola, D

    2009-10-01

    The objective of the present study was to assess the predictive ability of subsets of single nucleotide polymorphism (SNP) markers for development of low-cost, low-density genotyping assays in dairy cattle. Dense SNP genotypes of 4,703 Holstein bulls were provided by the USDA Agricultural Research Service. A subset of 3,305 bulls born from 1952 to 1998 was used to fit various models (training set), and a subset of 1,398 bulls born from 1999 to 2002 was used to evaluate their predictive ability (testing set). After editing, data included genotypes for 32,518 SNP and August 2003 and April 2008 predicted transmitting abilities (PTA) for lifetime net merit (LNM$), the latter resulting from progeny testing. The Bayesian least absolute shrinkage and selection operator method was used to regress August 2003 PTA on marker covariates in the training set to arrive at estimates of marker effects and direct genomic PTA. The coefficient of determination (R(2)) from regressing the April 2008 progeny test PTA of bulls in the testing set on their August 2003 direct genomic PTA was 0.375. Subsets of 300, 500, 750, 1,000, 1,250, 1,500, and 2,000 SNP were created by choosing equally spaced and highly ranked SNP, with the latter based on the absolute value of their estimated effects obtained from the training set. The SNP effects were re-estimated from the training set for each subset of SNP, and the 2008 progeny test PTA of bulls in the testing set were regressed on corresponding direct genomic PTA. The R(2) values for subsets of 300, 500, 750, 1,000, 1,250, 1,500, and 2,000 SNP with largest effects (evenly spaced SNP) were 0.184 (0.064), 0.236 (0.111), 0.269 (0.190), 0.289 (0.179), 0.307 (0.228), 0.313 (0.268), and 0.322 (0.291), respectively. These results indicate that a low-density assay comprising selected SNP could be a cost-effective alternative for selection decisions and that significant gains in predictive ability may be achieved by increasing the number of SNP allocated to

  17. Linkage map of the honey bee, Apis mellifera, based on RAPD markers

    SciTech Connect

    Hunt, G.J.; Page, R.E. Jr.

    1995-03-01

    A linkage map was constructed for the honey bee based on the segregation of 365 random amplified polymorphic DNA (RAPD) markers in haploid male progeny of a single female bee. The X locus for sex determination and genes for black body color and malate dehydrogenase were mapped to separate linkage groups. RAPD markers were very efficient for mapping, with an average of about 2.8 loci mapped for each 10-nucleotide primer that was used in polymerase chain reactions. The mean interval size between markers on the map was 9.1 cM. The map covered 3110 cM of linked markers on 26 linkage groups. We estimate the total genome size to be {approximately}3450 cM. The size of the map indicated a very high recombination rate for the honey bee. The relationship of physical to genetic distance was estimated at 52 kb/cM, suggesting that map-based cloning of genes will be feasible for this species. 71 refs., 6 figs., 1 tab.

  18. Linkage Map of the Honey Bee, Apis Mellifera, Based on Rapd Markers

    PubMed Central

    Hunt, G. J.; Page-Jr, R. E.

    1995-01-01

    A linkage map was constructed for the honey bee based on the segregation of 365 random amplified polymorphic DNA (RAPD) markers in haploid male progeny of a single female bee. The X locus for sex determination and genes for black body color and malate dehydrogenase were mapped to separate linkage groups. RAPD markers were very efficient for mapping, with an average of about 2.8 loci mapped for each 10-nucleotide primer that was used in polymerase chain reactions. The mean interval size between markers on the map was 9.1 cM. The map covered 3110 cM of linked markers on 26 linkage groups. We estimate the total genome size to be ~3450 cM. The size of the map indicated a very high recombination rate for the honey bee. The relationship of physical to genetic distance was estimated at 52 kb/cM, suggesting that map-based cloning of genes will be feasible for this species. PMID:7768445

  19. Ultrasound-based measurement of molecular marker concentration in large blood vessels: a feasibility study.

    PubMed

    Wang, Shiying; Mauldin, F William; Klibanov, Alexander L; Hossack, John A

    2015-01-01

    Ultrasound molecular imaging has demonstrated efficacy in pre-clinical studies for cancer and cardiovascular inflammation. However, these techniques often require lengthy protocols because of waiting periods or additional control microbubble injections. Moreover, they are not capable of quantifying molecular marker concentration in human tissue environments that exhibit variable attenuation and propagation path lengths. Our group recently investigated a modulated acoustic radiation force-based imaging sequence, which was found to detect targeted adhesion independent of control measurements. In the present study, this sequence was tested against various experimental parameters to determine its feasibility for quantitative measurements of molecular marker concentration. Results indicated that measurements obtained from the sequence (residual-to-saturation ratio, Rresid) were independent of acoustic pressure and attenuation (p > 0.13, n = 10) when acoustic pressures were sufficiently low. The Rresid parameter exhibited a linear relationship with measured molecular marker concentration (R(2) > 0.94). Consequently, feasibility was illustrated in vitro, for quantification of molecular marker concentration in large vessels using a modulated acoustic radiation force-based sequence. Moreover, these measurements were independent of absolute acoustic reflection amplitude and used short imaging protocols (3 min) without control measurements. Copyright © 2015 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

  20. Assessment of genetic diversity among Indian potato (Solanum tuberosum L.) collection using microsatellite and retrotransposon based marker systems.

    PubMed

    Sharma, Vishakha; Nandineni, Madhusudan R

    2014-04-01

    Potato (Solanum tuberosum) is an important non-cereal crop throughout the world and is highly recommended for ensuring global food security. Owing to the complexities in genetics and inheritance pattern of potato, the conventional method of cross breeding for developing improved varieties has been difficult. Identification and tagging of desirable traits with informative molecular markers would aid in the development of improved varieties. Insertional polymorphism of copia-like and gypsy-like long terminal repeat retrotransposons (RTN) were investigated among 47 potato varieties from India using Inter-Retrotransposon Amplified Polymorphism (IRAP) and Retrotransposon Microsatellite Amplified Polymorphism (REMAP) marker techniques and were compared with the DNA profiles obtained with simple sequence repeats (SSRs). The genetic polymorphism, efficiency of polymorphism and effectiveness of marker systems were evaluated to assess the extent of genetic diversity among Indian potato varieties. A total of 139 polymorphic SSR alleles, 270 IRAP and 98 REMAP polymorphic bands, showing polymorphism of 100%, 87.9% and 68.5%, respectively, were used for detailed characterization of the genetic relationships among potato varieties by using cluster analysis and principal coordinate analysis (PCoA). IRAP analysis resulted in the highest number of polymorphic bands with an average of 15 polymorphic bands per assay unit when compared to the other two marker systems. Based on pair-wise comparison, the genetic similarity was calculated using Dice similarity coefficient. The SSRs showed a wide range in genetic similarity values (0.485-0.971) as compared to IRAP (0.69-0.911) and REMAP (0.713-0.947). A Mantel's matrix correspondence test showed a high positive correlation (r=0.6) between IRAP and REMAP, an intermediate value (r=0.58) for IRAP and SSR and the lowest value (r=0.17) for SSR and REMAP. Statistically significant cophenetic correlation coefficient values, of 0.961, 0.941 and 0

  1. SNP markers-based map construction and genome-wide linkage analysis in Brassica napus.

    PubMed

    Raman, Harsh; Dalton-Morgan, Jessica; Diffey, Simon; Raman, Rosy; Alamery, Salman; Edwards, David; Batley, Jacqueline

    2014-09-01

    An Illumina Infinium array comprising 5306 single nucleotide polymorphism (SNP) markers was used to genotype 175 individuals of a doubled haploid population derived from a cross between Skipton and Ag-Spectrum, two Australian cultivars of rapeseed (Brassica napus L.). A genetic linkage map based on 613 SNP and 228 non-SNP (DArT, SSR, SRAP and candidate gene markers) covering 2514.8 cM was constructed and further utilized to identify loci associated with flowering time and resistance to blackleg, a disease caused by the fungus Leptosphaeria maculans. Comparison between genetic map positions of SNP markers and the sequenced Brassica rapa (A) and Brassica oleracea (C) genome scaffolds showed several genomic rearrangements in the B. napus genome. A major locus controlling resistance to L. maculans was identified at both seedling and adult plant stages on chromosome A07. QTL analyses revealed that up to 40.2% of genetic variation for flowering time was accounted for by loci having quantitative effects. Comparative mapping showed Arabidopsis and Brassica flowering genes such as Phytochrome A/D, Flowering Locus C and agamous-Like MADS box gene AGL1 map within marker intervals associated with flowering time in a DH population from Skipton/Ag-Spectrum. Genomic regions associated with flowering time and resistance to L. maculans had several SNP markers mapped within 10 cM. Our results suggest that SNP markers will be suitable for various applications such as trait introgression, comparative mapping and high-resolution mapping of loci in B. napus. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  2. Review of selected national surgical curricula: quantity is not the sole marker of quality.

    PubMed

    Singh, Pritam; Aggarwal, Rajesh; Darzi, Ara

    2014-01-01

    Despite marked global variations in length and structure of surgical training programs, their common end product is a trained surgeon capable of independent practice. If variations exist, yet the end product is similar, modifications to curricula could potentially enhance the quality and efficiency of surgical training. This review evaluates global general surgery training programs and compares their curricula against the established standards for assessment of curricula. A convenience sampling method was employed during an online search for nationally recognized general surgery curricula. Curricula of Australia, Canada, Hong Kong, the United Kingdom, and the United States of America were individually reviewed and subsequently evaluated against the General Medical Council's "Standards for curricula and assessment systems." Postgraduate surgical training is completed in 5 years in Canada and the United States, whereas this takes a minimum of 7, 7, and 10 years in Australia, Hong Kong, and the United Kingdom, respectively. However, when their general surgery curricula are objectively compared, they are remarkably similar. The principle disparities noted were in documentation and standardization of the structured in-training assessment system. This review highlights variations in the structure of general surgery training programs globally. There is a need for an objective method to assess training quality, not reliant upon quantity alone. An evidence-based approach is the gold standard in patient care; it is essential to invest resources into developing an evidence-based curricular approach to ensure surgical training quality can be accurately evaluated to maintain and enhance the standards. Copyright © 2014 Association of Program Directors in Surgery. Published by Elsevier Inc. All rights reserved.

  3. RAPD-based SCAR marker SCA 12 linked to recessive gene conferring resistance to anthracnose in sorghum [Sorghum bicolor (L.) Moench].

    PubMed

    Singh, Monika; Chaudhary, K; Boora, K S

    2006-12-01

    Anthracnose, caused by Colletotrichum graminicola, infects all aerial parts of sorghum, Sorghum bicolor (L.) Moench, plants and causes loss of as much as 70%. F(1) and F(2) plants inoculated with local isolates of C. graminicola indicated that resistance to anthracnose in sorghum accession G 73 segregated as a recessive trait in a cross with susceptible cultivar HC 136. To facilitate the use of marker-assisted selection in sorghum breeding programs, a PCR-based specific sequence characterized amplified region (SCAR) marker was developed. A total of 29 resistant and 20 susceptible recombinant inbred lines (RILs) derived from a HC 136 x G 73 cross was used for bulked segregant analysis to identify a RAPD marker closely linked to a gene for resistance to anthracnose. The polymorphism between the parents HC 136 and G 73 was evaluated using 84 random sequence decamer primers. Among these, only 24 primers generated polymorphism. On bulked segregant analysis, primer OPA 12 amplified a unique band of 383 bp only in the resistant parent G 73 and resistant bulk. Segregation analysis of individual RILs showed the marker OPA 12(383) was 6.03 cM from the locus governing resistance to anthracnose. The marker OPA 12(383) was cloned and sequenced. Based on the sequence of cloned RAPD product, a pair of SCAR markers SCA 12-1 and SCA 12-2 was designed using the MacVector program, which specifically amplified this RAPD fragment in resistant parent G 73, resistant bulk and respective RILs. Therefore, it was confirmed that SCAR marker SCA 12 is at the same locus as RAPD marker OPA 12(383) and hence, is linked to the gene for resistance to anthracnose.

  4. Persimmon breeding in Japan for pollination-constant non-astringent (PCNA) type with marker-assisted selection

    PubMed Central

    Sato, Akihiko; Yamada, Masahiko

    2016-01-01

    Oriental persimmon (Diospyros kaki) originated in Eastern Asia, and many indigenous cultivars have been developed in China, Japan, and Korea. These cultivars are classified into four groups based on their natural astringency loss on the tree and seed formation: pollination-constant non-astringent (PCNA), pollination-variant non-astringent (PVNA), pollination-constant astringent (PCA), and pollination-variant astringent (PVA). PCNA is the most desirable type because the fruit can be eaten without any postharvest treatment; therefore, one of the goals of our persimmon breeding programs is to release superior PCNA cultivars. The PCNA genotype is recessive to the other three non-PCNA genotypes, and PCNA-type F1 offspring are obtained exclusively from crosses among PCNA genotypes. Moreover, the number of superior PCNA cross-parents have been limited. In the late 1980s, inbreeding depression became obvious, especially in terms of fruit size, tree vigor, and productivity. To mitigate the inbreeding, a backcross program using PCNA [(non-PCNA × PCNA) × PCNA] was started in 1990. This process, however, was inefficient because only 15% of the offspring were PCNA, and all offspring had to be grown to the fruiting stage. Therefore, molecular markers linked to the PCNA locus were developed for discriminating PCNA offspring. A molecular marker linked to Chinese PCNA has also been developed. PMID:27069391

  5. Assessment of MRI-Based Marker of Dopaminergic Integrity as a Biological Indicator of Gulf War Illness

    DTIC Science & Technology

    2015-10-01

    AWARD NUMBER: W81XWH-14-1-0622 TITLE: Assessment of MRI -Based Marker of Dopaminergic Integrity as a Biological Indicator of Gulf War...COVERED 29 Sep 2014 – 28 Sept 2015 4. TITLE AND SUBTITLE Illness 5a. CONTRACT NUMBER Assessment of MRI -Based Marker of Dopaminergic Integrity as a

  6. An International Reference Consensus Genetic Map with 897 Marker Loci Based on 11 Mapping Populations for Tetraploid Groundnut (Arachis hypogaea L.)

    PubMed Central

    Pandey, Manish K.; Moretzsohn, Márcio C.; Sujay, Venkataswamy; Qin, Hongde; Hong, Yanbin; Faye, Issa; Chen, Xiaoping; BhanuPrakash, Amindala; Shah, Trushar M.; Gowda, Makanahally V. C.; Nigam, Shyam N.; Liang, Xuanqiang; Hoisington, Dave A.; Guo, Baozhu; Bertioli, David J.; Rami, Jean-Francois; Varshney, Rajeev K.

    2012-01-01

    Only a few genetic maps based on recombinant inbred line (RIL) and backcross (BC) populations have been developed for tetraploid groundnut. The marker density, however, is not very satisfactory especially in the context of large genome size (2800 Mb/1C) and 20 linkage groups (LGs). Therefore, using marker segregation data for 10 RILs and one BC population from the international groundnut community, with the help of common markers across different populations, a reference consensus genetic map has been developed. This map is comprised of 897 marker loci including 895 simple sequence repeat (SSR) and 2 cleaved amplified polymorphic sequence (CAPS) loci distributed on 20 LGs (a01–a10 and b01–b10) spanning a map distance of 3, 863.6 cM with an average map density of 4.4 cM. The highest numbers of markers (70) were integrated on a01 and the least number of markers (21) on b09. The marker density, however, was lowest (6.4 cM) on a08 and highest (2.5 cM) on a01. The reference consensus map has been divided into 20 cM long 203 BINs. These BINs carry 1 (a10_02, a10_08 and a10_09) to 20 (a10_04) loci with an average of 4 marker loci per BIN. Although the polymorphism information content (PIC) value was available for 526 markers in 190 BINs, 36 and 111 BINs have at least one marker with >0.70 and >0.50 PIC values, respectively. This information will be useful for selecting highly informative and uniformly distributed markers for developing new genetic maps, background selection and diversity analysis. Most importantly, this reference consensus map will serve as a reliable reference for aligning new genetic and physical maps, performing QTL analysis in a multi-populations design, evaluating the genetic background effect on QTL expression, and serving other genetic and molecular breeding activities in groundnut. PMID:22815973

  7. A wheat intervarietal genetic linkage map based on microsatellite and target region amplified polymorphism markers and its utility for detecting quantitative trait loci.

    PubMed

    Liu, Z H; Anderson, J A; Hu, J; Friesen, T L; Rasmussen, J B; Faris, J D

    2005-08-01

    Efficient user-friendly methods for mapping plant genomes are highly desirable for the identification of quantitative trait loci (QTLs), genotypic profiling, genomic studies, and marker-assisted selection. SSR (microsatellite) markers are user-friendly and efficient in detecting polymorphism, but they detect few loci. Target region amplification polymorphism (TRAP) is a relatively new PCR-based technique that detects a large number of loci from a single reaction without extensive pre-PCR processing of samples. In the investigation reported here, we used both SSRs and TRAPs to generate over 700 markers for the construction of a genetic linkage map in a hard red spring wheat intervarietal recombinant inbred population. A framework map consisting of 352 markers accounted for 3,045 cM with an average density of one marker per 8.7 cM. On average, SSRs detected 1.9 polymorphic loci per reaction, while TRAPs detected 24. Both marker systems were suitable for assigning linkage groups to chromosomes using wheat aneuploid stocks. We demonstrated the utility of the maps by identifying major QTLs for days to heading and reduced plant height on chromosomes 5A and 4B, respectively. Our results indicate that TRAPs are highly efficient for genetic mapping in wheat. The maps developed will be useful for the identification of quality and disease resistance QTLs that segregate in this population.

  8. Transcriptome sequencing of different narrow-leafed lupin tissue types provides a comprehensive uni-gene assembly and extensive gene-based molecular markers

    PubMed Central

    Kamphuis, Lars G; Hane, James K; Nelson, Matthew N; Gao, Lingling; Atkins, Craig A; Singh, Karam B

    2015-01-01

    Narrow-leafed lupin (NLL; Lupinus angustifolius L.) is an important grain legume crop that is valuable for sustainable farming and is becoming recognized as a human health food. NLL breeding is directed at improving grain production, disease resistance, drought tolerance and health benefits. However, genetic and genomic studies have been hindered by a lack of extensive genomic resources for the species. Here, the generation, de novo assembly and annotation of transcriptome datasets derived from five different NLL tissue types of the reference accession cv. Tanjil are described. The Tanjil transcriptome was compared to transcriptomes of an early domesticated cv. Unicrop, a wild accession P27255, as well as accession 83A:476, together being the founding parents of two recombinant inbred line (RIL) populations. In silico predictions for transcriptome-derived gene-based length and SNP polymorphic markers were conducted and corroborated using a survey assembly sequence for NLL cv. Tanjil. This yielded extensive indel and SNP polymorphic markers for the two RIL populations. A total of 335 transcriptome-derived markers and 66 BAC-end sequence-derived markers were evaluated, and 275 polymorphic markers were selected to genotype the reference NLL 83A:476 × P27255 RIL population. This significantly improved the completeness, marker density and quality of the reference NLL genetic map. PMID:25060816

  9. Transcriptome sequencing of different narrow-leafed lupin tissue types provides a comprehensive uni-gene assembly and extensive gene-based molecular markers.

    PubMed

    Kamphuis, Lars G; Hane, James K; Nelson, Matthew N; Gao, Lingling; Atkins, Craig A; Singh, Karam B

    2015-01-01

    Narrow-leafed lupin (NLL; Lupinus angustifolius L.) is an important grain legume crop that is valuable for sustainable farming and is becoming recognized as a human health food. NLL breeding is directed at improving grain production, disease resistance, drought tolerance and health benefits. However, genetic and genomic studies have been hindered by a lack of extensive genomic resources for the species. Here, the generation, de novo assembly and annotation of transcriptome datasets derived from five different NLL tissue types of the reference accession cv. Tanjil are described. The Tanjil transcriptome was compared to transcriptomes of an early domesticated cv. Unicrop, a wild accession P27255, as well as accession 83A:476, together being the founding parents of two recombinant inbred line (RIL) populations. In silico predictions for transcriptome-derived gene-based length and SNP polymorphic markers were conducted and corroborated using a survey assembly sequence for NLL cv. Tanjil. This yielded extensive indel and SNP polymorphic markers for the two RIL populations. A total of 335 transcriptome-derived markers and 66 BAC-end sequence-derived markers were evaluated, and 275 polymorphic markers were selected to genotype the reference NLL 83A:476 × P27255 RIL population. This significantly improved the completeness, marker density and quality of the reference NLL genetic map. © 2014 CSIRO. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  10. Marker peptide selection for the determination of hazelnut by LC-MS/MS and occurrence in other nuts.

    PubMed

    Ansari, Parisa; Stoppacher, Norbert; Baumgartner, Sabine

    2012-03-01

    The aim of this work was identifying and selecting hazelnut marker peptides and subsequently developing a complementary method of common immunoassay for the detection of hazelnut. For this purpose, at first, an in silico digestion of three major hazelnut allergens (Cor a 8, Cor a 9 and Cor a 11) was performed to get information about expected peptides. After extraction and trypsin digestion of hazelnut proteins, the samples were measured with tandem mass spectrometry (MS/MS) by direct infusion, which led to identification of 14 peptides. Eight of them with the highest MS signal were synthesized and used as standards for developing a liquid chromatography (LC)-MS/MS method in selected reaction monitoring (SRM) mode. Since almost all food allergens derived from nuts belong to the seed storage protein family and have homologue structure, a Basic Local Alignment Search Tool (BLAST) search was performed to identify the hazelnut specificity of the developed method. According to BLAST, only one peptide occurs in three other nuts, and the remaining seven selected peptides are hazelnut specific. Additionally to hazelnut, the eight other listed nuts in Directive 2003/89/EC as allergen were extracted, digested and measured with the developed method to prove the BLAST results. The analytical data confirmed that six peptides are hazelnut specific, on the contrary to anti-hazelnut antibodies, which showed cross-reactivities to all other nut extracts. Comparing these results, it could be shown that with this LC-MS/MS method in SRM mode, the specific detection of hazelnut is possible.

  11. Phylogeny reconstruction in the Caesalpinieae grade (Leguminosae) based on duplicated copies of the sucrose synthase gene and plastid markers.

    PubMed

    Manzanilla, Vincent; Bruneau, Anne

    2012-10-01

    The Caesalpinieae grade (Leguminosae) forms a morphologically and ecologically diverse group of mostly tropical tree species with a complex evolutionary history. This grade comprises several distinct lineages, but the exact delimitation of the group relative to subfamily Mimosoideae and other members of subfamily Caesalpinioideae, as well as phylogenetic relationships among the lineages are uncertain. With the aim of better resolving phylogenetic relationships within the Caesalpinieae grade, we investigated the utility of several nuclear markers developed from genomic studies in the Papilionoideae. We cloned and sequenced the low copy nuclear gene sucrose synthase (SUSY) and combined the data with plastid trnL and matK sequences. SUSY has two paralogs in the Caesalpinieae grade and in the Mimosoideae, but occurs as a single copy in all other legumes tested. Bayesian and maximum likelihood phylogenetic analyses suggest the two nuclear markers are congruent with plastid DNA data. The Caesalpinieae grade is divided into four well-supported clades (Cassia, Caesalpinia, Tachigali and Peltophorum clades), a poorly supported clade of Dimorphandra Group genera, and two paraphyletic groups, one with other Dimorphandra Group genera and the other comprising genera previously recognized as the Umtiza clade. A selection analysis of the paralogs, using selection models from PAML, suggests that SUSY genes are subjected to a purifying selection. One of the SUSY paralogs, under slightly stronger positive selection, may be undergoing subfunctionalization. The low copy SUSY gene is useful for phylogeny reconstruction in the Caesalpinieae despite the presence of duplicate copies. This study confirms that the Caesalpinieae grade is an artificial group, and highlights the need for further analyses of lineages at the base of the Mimosoideae.

  12. An approach of selecting appropriate markers from the primary tumor to enable detection of circulating tumor cells in patients with non-small cell lung cancer.

    PubMed

    Warawdekar, Ujjwala M; Sirajuddin, Mohamed M; Pramesh, Conjeevaram S; Mistry, Rajesh C

    2015-01-01

    Circulating tumor cells (CTCs) are rare and difficult to isolate, and require selecting minimal but appropriate markers. The aim of this study was to identify markers in the primary non small cell lung cancer (NSCLC) tissue to guide isolation of CTCs from the peripheral blood of patients with lung cancer. The expression of CK-19, EGFR and MUC-1 was evaluated by RT-PCR in the NSCLC tumor and paired adjacent normal tissues from 27 patients. The normal cytology, and the neoplastic and fibrotic pathology of the tissue were analyzed by histochemistry. The expression of the markers was analyzed in relation to the stage and grade of disease. Expression analysis showed that 42% of the tumors were positive for CK-19, whereas 85% for both EGFR and MUC-1. Ninety two percent of the tumors expressed any one marker. All (100%) adjacent normal tissues were CK-19 negative, 52% EGFR negative and 44% MUC-1 negative. CK-19 expression was specific to the tumor tissue but it was expressed by only 42% of them, manifesting a need for at least three markers to guide the detection of CTCs isolated from the peripheral blood of NSCLC patients. Histopathology demonstrated that 58% were adenocarcinomas, 35% squamous cell carcinomas and 7% had mixed pathology. This data serves as a prelude and emphasizes the importance of selecting markers expressed in the primary tumor tissue to facilitate and enable enumeration of CTCs.

  13. A marker-based watershed method for X-ray image segmentation.

    PubMed

    Zhang, Xiaodong; Jia, Fucang; Luo, Suhuai; Liu, Guiying; Hu, Qingmao

    2014-03-01

    Digital X-ray images are the most frequent modality for both screening and diagnosis in hospitals. To facilitate subsequent analysis such as quantification and computer aided diagnosis (CAD), it is desirable to exclude image background. A marker-based watershed segmentation method was proposed to segment background of X-ray images. The method consisted of six modules: image preprocessing, gradient computation, marker extraction, watershed segmentation from markers, region merging and background extraction. One hundred clinical direct radiograph X-ray images were used to validate the method. Manual thresholding and multiscale gradient based watershed method were implemented for comparison. The proposed method yielded a dice coefficient of 0.964±0.069, which was better than that of the manual thresholding (0.937±0.119) and that of multiscale gradient based watershed method (0.942±0.098). Special means were adopted to decrease the computational cost, including getting rid of few pixels with highest grayscale via percentile, calculation of gradient magnitude through simple operations, decreasing the number of markers by appropriate thresholding, and merging regions based on simple grayscale statistics. As a result, the processing time was at most 6s even for a 3072×3072 image on a Pentium 4 PC with 2.4GHz CPU (4 cores) and 2G RAM, which was more than one time faster than that of the multiscale gradient based watershed method. The proposed method could be a potential tool for diagnosis and quantification of X-ray images.

  14. Personalized Clinical Trials in Hepatocellular Carcinoma Based on Biomarker Selection

    PubMed Central

    Zhang, Bingnan; Finn, Richard S.

    2016-01-01

    Background Since the approval of sorafenib there have been numerous failures of new agents in Phase III studies for treatment of advanced hepatocellular carcinoma (HCC). These studies have generally ignored the molecular heterogeneity of HCC and they have not enrolled patients based on predictive markers of response. The development of molecular targeted therapeutics in HCC needs to model the approach that has been taken with great success in other solid tumors, to decrease the likelihood of failure in future studies. Summary Here we review the paradigm taken with novel targeted agents in other solid tumors and highlight ongoing studies in HCC that are incorporating biomarkers in clinical development. Key Messages With the appreciation of the molecular diversity of HCC, clinical development of new agents in HCC will need to be targeted towards those patients who are most likely to benefit. This strategy, based on biomarkers for patient selection, is more likely to yield positive results and mitigate the risk of continued negative Phase III studies. PMID:27493897

  15. ESAP plus: a web-based server for EST-SSR marker development.

    PubMed

    Ponyared, Piyarat; Ponsawat, Jiradej; Tongsima, Sissades; Seresangtakul, Pusadee; Akkasaeng, Chutipong; Tantisuwichwong, Nathpapat

    2016-12-22

    Simple sequence repeats (SSRs) have become widely used as molecular markers in plant genetic studies due to their abundance, high allelic variation at each locus and simplicity to analyze using conventional PCR amplification. To study plants with unknown genome sequence, SSR markers from Expressed Sequence Tags (ESTs), which can be obtained from the plant mRNA (converted to cDNA), must be utilized. With the advent of high-throughput sequencing technology, huge EST sequence data have been generated and are now accessible from many public databases. However, SSR marker identification from a large in-house or public EST collection requires a computational pipeline that makes use of several standard bioinformatic tools to design high quality EST-SSR primers. Some of these computational tools are not users friendly and must be tightly integrated with reference genomic databases. A web-based bioinformatic pipeline, called EST Analysis Pipeline Plus (ESAP Plus), was constructed for assisting researchers to develop SSR markers from a large EST collection. ESAP Plus incorporates several bioinformatic scripts and some useful standard software tools necessary for the four main procedures of EST-SSR marker development, namely 1) pre-processing, 2) clustering and assembly, 3) SSR mining and 4) SSR primer design. The proposed pipeline also provides two alternative steps for reducing EST redundancy and identifying SSR loci. Using public sugarcane ESTs, ESAP Plus automatically executed the aforementioned computational pipeline via a simple web user interface, which was implemented using standard PHP, HTML, CSS and Java scripts. With ESAP Plus, users can upload raw EST data and choose various filtering options and parameters to analyze each of the four main procedures through this web interface. All input EST data and their predicted SSR results will be stored in the ESAP Plus MySQL database. Users will be notified via e-mail when the automatic process is completed and they can

  16. Potential coeliac disease markers and autoimmunity in olmesartan induced enteropathy: A population-based study.

    PubMed

    Esteve, Maria; Temiño, Rocío; Carrasco, Anna; Batista, Lissette; Del Val, Adolfo; Blé, Michel; Santaolaria, Santos; Molina-Infante, Javier; Soriano, Germán; Agudo, Sandra; Zabana, Yamile; Andújar, Xavier; Aceituno, Montserrat; Ribes, Josepa; Madridejos, Rosa; Fernández-Bañares, Fernando

    2016-02-01

    (1) Assess the population-based incidence of severe olmesartan-associated enteropathy. (2) To describe patients of the Spanish registry. (3) Evaluate markers of potential coeliac disease and associated autoimmunity. Crude incidence rates in the area of Terrassa (Catalonia) were calculated. Clinical characteristics of patients in the Spanish registry were collected. Duodenal lymphocyte subpopulations and anti-TG2 IgA deposits were assessed in a subset of patients. Annual incidence rates (2011-2014) ranged from 0 to 22 cases per 10(4) treated patients. Twenty patients were included in the Spanish registry. Nineteen (95%) exhibited villous atrophy and 16 (80%) had severe enteropathy. Lupus-like disease occurred during olmesartan treatment in 3 patients. HLA-DQ2/DQ8 was positive in 64%. Markers of potential coeliac disease were present in 4 out of 8 patients (positive anti-TG2 deposits and/or increased CD3+gammadelta+ intraepithelial lymphocytes and reduced CD3-). Histopathological changes and clinical manifestations including autoimmune disorders improved after olmesartan discontinuation but not after gluten-free diet, irrespective of the presence or absence of coeliac markers. Incidence of severe olmesartan-associated enteropathy was low. Autoimmune phenomena were present in a subset of cases and reversed after olmesartan removal. A genetic coeliac disease background and the presence of potential coeliac markers might uncover predisposing factors. Copyright © 2015 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.