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Sample records for groupings usingbranched dna

  1. Directed assembly of discrete gold nanoparticle groupings usingbranched DNA scaffolds

    SciTech Connect

    Claridge, Shelley A.; Goh, Sarah L.; Frechet, Jean M.J.; Williams, Shara C.; Micheel, Christine M.; Alivisatos, A. Paul

    2004-09-14

    The concept of self-assembled dendrimers is explored for the creation of discrete nanoparticle assemblies. Hybridization of branched DNA trimers and nanoparticle-DNA conjugates results in the synthesis of nanoparticle trimer and tetramer complexes. Multiple tetramer architectures are investigated, utilizing Au-DNA conjugates with varying secondary structural motifs. Hybridization products are analyzed by gel electrophoresis, and discrete bands are observed corresponding to structures with increasing numbers of hybridization events. Samples extracted from each band are analyzed by transmission electron microscopy, and statistics compiled from micrographs are used to compare assembly characteristics for each architecture. Asymmetric structures are also produced in which both 5 and 10 nm Au particles are assembled on branched scaffolds.

  2. Finding human promoter groups based on DNA physical properties

    NASA Astrophysics Data System (ADS)

    Zeng, Jia; Cao, Xiao-Qin; Zhao, Hongya; Yan, Hong

    2009-10-01

    DNA rigidity is an important physical property originating from the DNA three-dimensional structure. Although the general DNA rigidity patterns in human promoters have been investigated, their distinct roles in transcription are largely unknown. In this paper, we discover four highly distinct human promoter groups based on similarity of their rigidity profiles. First, we find that all promoter groups conserve relatively rigid DNAs at the canonical TATA box [a consensus TATA(A/T)A(A/T) sequence] position, which are important physical signals in binding transcription factors. Second, we find that the genes activated by each group of promoters share significant biological functions based on their gene ontology annotations. Finally, we find that these human promoter groups correlate with the tissue-specific gene expression.

  3. Human xeroderma pigmentosum group G gene encodes a DNA endonuclease.

    PubMed Central

    Habraken, Y; Sung, P; Prakash, L; Prakash, S

    1994-01-01

    Because of defective nucleotide excision repair of ultraviolet damaged DNA, xeroderma pigmentosum (XP) patients suffer from a high incidence of skin cancers. Cell fusion studies have identified seven XP complementation groups, A to G. Previous studies have implicated the products of these seven XP genes in the recognition of ultraviolet-induced DNA damage and in incision of the damage-containing DNA strand. Here, we express the XPG-encoded protein in Sf9 insect cells and purify it to homogeneity. We demonstrate that XPG is a single-strand specific DNA endonuclease, thus identifying the catalytic role of the protein in nucleotide excision repair. We suggest that XPG nuclease acts on the single-stranded region created as a result of the combined action of the XPB helicase and XPD helicase at the DNA damage site. Images PMID:8078765

  4. Mitochondrial DNA evolution in the Anaxyrus boreas species group

    USGS Publications Warehouse

    Goebel, A.M.; Ranker, T.A.; Corn, P.S.; Olmstead, R.G.

    2009-01-01

    The Anaxyrus boreas species group currently comprises four species in western North America including the broadly distributed A. boreas, and three localized species, Anaxyrus nelsoni, Anaxyrus exsul and Anaxyrus canorus. Phylogenetic analyses of the mtDNA 12S rDNA, cytochrome oxidase I, control region, and restriction sites data, identified three major haplotype clades. The Northwest clade (NW) includes both subspecies of A. boreas and divergent minor clades in the middle Rocky Mountains, coastal, and central regions of the west and Pacific Northwest. The Southwest (SW) clade includes A. exsul, A. nelsoni, and minor clades in southern California. Anaxyrus canorus, previously identified as paraphyletic, has populations in both the NW and SW major clades. The Eastern major clade (E) includes three divergent lineages from southern Utah, the southern Rocky Mountains, and north of the Great Basin at the border of Utah and Nevada. These results identify new genetic variation in the eastern portion of the toad's range and are consistent with previous regional studies from the west coast. Low levels of control region sequence divergence between major clades (2.2-4.7% uncorrected pair-wise distances) are consistent with Pleistocene divergence and suggest that the phylogeographic history of the group was heavily influenced by dynamic Pleistocene glacial and climatic changes, and especially pluvial changes, in western North America. Results reported here may impact conservation plans in that the current taxonomy does not reflect the diversity in the group. ?? 2008 Elsevier Inc.

  5. Mitochondrial DNA evolution in the Anaxyrus boreas species group.

    PubMed

    Goebel, Anna M; Ranker, Tom A; Corn, Paul Stephen; Olmstead, Richard G

    2009-02-01

    The Anaxyrus boreas species group currently comprises four species in western North America including the broadly distributed A. boreas, and three localized species, Anaxyrus nelsoni, Anaxyrusexsul and Anaxyrus canorus. Phylogenetic analyses of the mtDNA 12S rDNA, cytochrome oxidase I, control region, and restriction sites data, identified three major haplotype clades. The Northwest clade (NW) includes both subspecies of A. boreas and divergent minor clades in the middle Rocky Mountains, coastal, and central regions of the west and Pacific Northwest. The Southwest (SW) clade includes A. exsul, A. nelsoni, and minor clades in southern California. Anaxyrus canorus, previously identified as paraphyletic, has populations in both the NW and SW major clades. The Eastern major clade (E) includes three divergent lineages from southern Utah, the southern Rocky Mountains, and north of the Great Basin at the border of Utah and Nevada. These results identify new genetic variation in the eastern portion of the toad's range and are consistent with previous regional studies from the west coast. Low levels of control region sequence divergence between major clades (2.2-4.7% uncorrected pair-wise distances) are consistent with Pleistocene divergence and suggest that the phylogeographic history of the group was heavily influenced by dynamic Pleistocene glacial and climatic changes, and especially pluvial changes, in western North America. Results reported here may impact conservation plans in that the current taxonomy does not reflect the diversity in the group. PMID:18662792

  6. Genetic Kinship Investigation from Blood Groups to DNA Markers

    PubMed Central

    Geserick, Gunther; Wirth, Ingo

    2012-01-01

    The forensic application of hereditary characteristics became possible after the discovery of human blood groups by Karl Landsteiner in 1901. The foundation for their use in kinship investigation was laid by Emil von Dungern and Ludwig Hirschfeld in 1910 by clarification of the inheritance of the ABO groups. Up to the middle of the 20th century further red cell membrane systems were discovered. From the 1920s Fritz Schiff and Georg Strassmann fought for the introduction of blood groups into forensic kinship investigation. A new era of hemogenetics was opened from 1955 as genetic polymorphisms were described in serum proteins. Starting in 1958 there followed the complex HLA system of white blood cells, which from 1963 was joined by polymophisms in erythrocyte enzymes. Therefore, from the 1980s, it was possible to clarify the majority of kinship cases with a combination of conventional markers. From 1990 to 2000 the conventional markers were gradually replaced by the more effective DNA markers. Simultaneously typing shifted from the phenotype level to the genotype level. The genomic structure of conventional genetic markers could also now be explained. As a reflection of scientific progress the legal situation also changed, particularly in the form of the official guidelines for kinship investigation. PMID:22851931

  7. Short-step chemical synthesis of DNA by use of MMTrS group for protection of 5'-hydroxyl group.

    PubMed

    Shiraishi, Miyuki; Utagawa, Eri; Ohkubo, Akihiro; Sekine, Mitsuo; Seio, Kohji

    2007-01-01

    4-methoxytrithylthio (MMTrS) group was applied for the appropriately protected four canonical nucleosides. We prepared the phosphoroamidite units by use of these nucleosides and developed the synthesis of oligodeoxynucleotides without any acidic treatment. Moreover, the new DNA synthesis protocol was applied to an automated DNA synthesizer for the synthesis of longer oligodeoxynucleotides. PMID:18029620

  8. Hands on Group Work Paper Model for Teaching DNA Structure, Central Dogma and Recombinant DNA

    ERIC Educational Resources Information Center

    Altiparmak, Melek; Nakiboglu Tezer, Mahmure

    2009-01-01

    Understanding life on a molecular level is greatly enhanced when students are given the opportunity to visualize the molecules. Especially understanding DNA structure and function is essential for understanding key concepts of molecular biology such as DNA, central dogma and the manipulation of DNA. Researches have shown that undergraduate…

  9. Sequence of figwort mosaic virus DNA (caulimovirus group).

    PubMed Central

    Richins, R D; Scholthof, H B; Shepherd, R J

    1987-01-01

    The nucleotide sequence of an infectious clone of figwort mosaic virus (FMV) was determined using the dideoxynucleotide chain termination method. The double-stranded DNA genome (7743 base pairs) contained eight open reading frames (ORFs), seven of which corresponded approximately in size and location to the ORFs found in the genome of cauliflower mosaic virus (CaMV) and carnation etched ring virus (CERV). ORFs I and V of FMV demonstrated the highest degrees of nucleotide and amino acid sequence homology with the equivalent coding regions of CaMV and CERV. Regions II, III and IV showed somewhat less homology with the analogous regions of CaMV and CERV, and ORF VI showed homology with the corresponding gene of CaMV and CERV in only a short segment near the middle of the putative gene product. A 16 nucleotide sequence, complementary to the 3' terminus of methionine initiator tRNA (tRNAimet) and presumed to be the primer binding site for initiation of reverse transcription to produce minus strand DNA, was found in the FMV genome near the discontinuity in the minus strand. Sequences near the three interruptions in the plus strand of FMV DNA bear strong resemblance to similarly located sequences of 3 other caulimoviruses and are inferred to be initiation sites for second strand DNA synthesis. Additional conserved sequences in the small and large intergenic regions are pointed out including a highly conserved 35 bp sequence that occurs in the latter region. PMID:3671088

  10. Sequence of figwort mosaic virus DNA (caulimovirus group).

    PubMed

    Richins, R D; Scholthof, H B; Shepherd, R J

    1987-10-26

    The nucleotide sequence of an infectious clone of figwort mosaic virus (FMV) was determined using the dideoxynucleotide chain termination method. The double-stranded DNA genome (7743 base pairs) contained eight open reading frames (ORFs), seven of which corresponded approximately in size and location to the ORFs found in the genome of cauliflower mosaic virus (CaMV) and carnation etched ring virus (CERV). ORFs I and V of FMV demonstrated the highest degrees of nucleotide and amino acid sequence homology with the equivalent coding regions of CaMV and CERV. Regions II, III and IV showed somewhat less homology with the analogous regions of CaMV and CERV, and ORF VI showed homology with the corresponding gene of CaMV and CERV in only a short segment near the middle of the putative gene product. A 16 nucleotide sequence, complementary to the 3' terminus of methionine initiator tRNA (tRNAimet) and presumed to be the primer binding site for initiation of reverse transcription to produce minus strand DNA, was found in the FMV genome near the discontinuity in the minus strand. Sequences near the three interruptions in the plus strand of FMV DNA bear strong resemblance to similarly located sequences of 3 other caulimoviruses and are inferred to be initiation sites for second strand DNA synthesis. Additional conserved sequences in the small and large intergenic regions are pointed out including a highly conserved 35 bp sequence that occurs in the latter region. PMID:3671088

  11. Two high-mobility group box domains act together to underwind and kink DNA

    SciTech Connect

    Sánchez-Giraldo, R.; Acosta-Reyes, F. J.; Malarkey, C. S.; Saperas, N.; Churchill, M. E. A.; Campos, J. L.

    2015-06-30

    The crystal structure of HMGB1 box A bound to an unmodified AT-rich DNA fragment is reported at a resolution of 2 Å. A new mode of DNA recognition for HMG box proteins is found in which two box A domains bind in an unusual configuration generating a highly kinked DNA structure. High-mobility group protein 1 (HMGB1) is an essential and ubiquitous DNA architectural factor that influences a myriad of cellular processes. HMGB1 contains two DNA-binding domains, box A and box B, which have little sequence specificity but have remarkable abilities to underwind and bend DNA. Although HMGB1 box A is thought to be responsible for the majority of HMGB1–DNA interactions with pre-bent or kinked DNA, little is known about how it recognizes unmodified DNA. Here, the crystal structure of HMGB1 box A bound to an AT-rich DNA fragment is reported at a resolution of 2 Å. Two box A domains of HMGB1 collaborate in an unusual configuration in which the Phe37 residues of both domains stack together and intercalate the same CG base pair, generating highly kinked DNA. This represents a novel mode of DNA recognition for HMGB proteins and reveals a mechanism by which structure-specific HMG boxes kink linear DNA.

  12. Hematopoietic gene promoters subjected to a group-combinatorial study of DNA samples: identification of a megakaryocytic selective DNA signature

    PubMed Central

    Hazony, Yehonathan; Lu, Jun; St. Hilaire, Cynthia; Ravid, Katya

    2006-01-01

    Identification of common sub-sequences for a group of functionally related DNA sequences can shed light on the role of such elements in cell-specific gene expression. In the megakaryocytic lineage, no one single unique transcription factor was described as linage specific, raising the possibility that a cluster of gene promoter sequences presents a unique signature. Here, the megakaryocytic gene promoter group, which consists of both human and mouse 5′ non-coding regions, served as a case study. A methodology for group-combinatorial search has been implemented as a customized software platform. It extracts the longest common sequences for a group of related DNA sequences and allows for single gaps of varying length, as well as double- and multiple-gap sequences. The results point to common DNA sequences in a group of genes that is selectively expressed in megakaryocytes, and which does not appear in a large group of control, random and specific sequences. This suggests a role for a combination of these sequences in cell-specific gene expression in the megakaryocytic lineage. The data also point to an intrinsic cross-species difference in the organization of 5′ non-coding sequences within the mammalian genomes. This methodology may be used for the identification of regulatory sequences in other lineages. PMID:16936310

  13. Complementation of DNA repair in xeroderma pigmentosum group A cell extracts by a protein with affinity for damaged DNA.

    PubMed

    Robins, P; Jones, C J; Biggerstaff, M; Lindahl, T; Wood, R D

    1991-12-01

    Complementation group A of xeroderma pigmentosum (XP) represents one of the most prevalent and serious forms of this cancer-prone disorder. Because of a marked defect in DNA excision repair, cells from individuals with XP-A are hypersensitive to the toxic and mutagenic effects of ultraviolet light and many chemical agents. We report here the isolation of the XP-A DNA repair protein by complementation of cell extracts from a repair-defective human XP-A cell line. XP-A protein purified from calf thymus migrates on denaturing gel electrophoresis as a doublet of 40 and 42 kilodaltons. The XP-A protein binds preferentially to ultraviolet light-irradiated DNA, with a preference for damaged over nondamaged nucleotides of approximately 10(3). This strongly suggests that the XP-A protein plays a direct role in the recognition of and incision at lesions in DNA. We further show that this protein corresponds to the product encoded by a recently isolated gene that can restore excision repair to XP-A cells. Thus, excision repair of plasmid DNA by cell extracts sufficiently resembles genomic repair in cells to reveal accurately the repair defect in an inherited disease. The general approach described here can be extended to the identification and isolation of other human DNA repair proteins. PMID:1935910

  14. Complementation of DNA repair in xeroderma pigmentosum group A cell extracts by a protein with affinity for damaged DNA.

    PubMed Central

    Robins, P; Jones, C J; Biggerstaff, M; Lindahl, T; Wood, R D

    1991-01-01

    Complementation group A of xeroderma pigmentosum (XP) represents one of the most prevalent and serious forms of this cancer-prone disorder. Because of a marked defect in DNA excision repair, cells from individuals with XP-A are hypersensitive to the toxic and mutagenic effects of ultraviolet light and many chemical agents. We report here the isolation of the XP-A DNA repair protein by complementation of cell extracts from a repair-defective human XP-A cell line. XP-A protein purified from calf thymus migrates on denaturing gel electrophoresis as a doublet of 40 and 42 kilodaltons. The XP-A protein binds preferentially to ultraviolet light-irradiated DNA, with a preference for damaged over nondamaged nucleotides of approximately 10(3). This strongly suggests that the XP-A protein plays a direct role in the recognition of and incision at lesions in DNA. We further show that this protein corresponds to the product encoded by a recently isolated gene that can restore excision repair to XP-A cells. Thus, excision repair of plasmid DNA by cell extracts sufficiently resembles genomic repair in cells to reveal accurately the repair defect in an inherited disease. The general approach described here can be extended to the identification and isolation of other human DNA repair proteins. Images PMID:1935910

  15. Mitochondrial DNA Haplogroup A may confer a genetic susceptibility to AIDS group from Southwest China.

    PubMed

    Wang, Hua-Wei; Xu, Yu; Miao, Ying-Lei; Luo, Hua-You; Wang, Kun-Hua

    2016-05-01

    The acquired immunodeficiency syndrome (AIDS) in humans was one of the chronic infections caused by human immunodeficiency virus (HIV), and the interactions between viral infection and mitochondrial energetic implicated that mitochondrial DNA (mtDNA) variation(s) may effect genetic susceptibility to AIDS. Thus, to illustrate the maternal genetic structure and further identify whether mtDNA variation(s) can effect HIV infection among southwest Chinese AIDS group, the whole mtDNA control region sequences of 70 AIDS patients and 480 health individuals from southwest China were analyzed here. Our results indicated the plausible recent genetic admixture results of AIDS group; comparison of matrilineal components between AIDS and matched Han groups showed that mtDNA haplogroup A (p = 0.048, OR = 3.006, 95% CI = 1.109-8.145) has a significant higher difference between the two groups; further comparison illustrated that mtDNA mutations 16,209 (p = 0.046, OR = 2.607, 95% CI = 0.988-6.876) and 16,319 (p = 0.009, OR = 2.965, 95% CI = 1.278-6.876) have significant differences between AIDS and matched control groups, and both of which were the defining variations of mtDNA haplogroup A, they further confirmed that mtDNA haplogroup A may confer genetic susceptibility to AIDS. Our results suggested that haplogroup A may confer a genetic susceptibility to AIDS group from Southwest China. PMID:25431816

  16. Mitochondrial DNA genetic diversity among four ethnic groups in Sierra Leone.

    PubMed

    Jackson, Bruce A; Wilson, Jamie Lee; Kirbah, Salwa; Sidney, Sheree S; Rosenberger, Joshua; Bassie, Larry; Alie, Joe A D; McLean, David C; Garvey, W Timothy; Ely, Bert

    2005-09-01

    Although there are numerous ethnic groups in Sierra Leone, the Mende and Temne together account for approximately 60% of the total population. To see if genetic differences could be observed among ethnic groups in Sierra Leone, the nucleotide sequence of the hypervariable 1 (HV1) region of mitochondrial DNA (mtDNA) was determined from samples of the two major ethnic groups, the Mende (n=59) and Temne (n=121), and of two minor ethnic groups, the Loko (n=29) and Limba (n=67). Among these 276 HV1 sequences, 164 individual haplotypes were observed. An analysis of molecular variance indicated that the distribution of these haplotypes within the Limba sample was significantly different from that of the other ethnic groups. No significant genetic variation was seen between the Mende, Temne, and Loko. These results indicate that distinguishing genetic differences can be observed among ethnic groups residing in historically close proximity to one another. Furthermore, we observed some mitochondrial DNA haplotypes that are common among the Sierra Leone ethnic groups but that have not been observed in other published studies of West African ethnic groups. Therefore, we may have evidence for mtDNA lineages that are unique to this region of West Africa. PMID:15761855

  17. Exocyclic groups in the minor groove influence the backbone conformation of DNA

    PubMed Central

    Wellenzohn, Bernd; Flader, Wolfgang; Winger, Rudolf H.; Hallbrucker, Andreas; Mayer, Erwin; Liedl, Klaus R.

    2001-01-01

    Exocyclic groups in the minor groove of DNA modulate the affinity and positioning of nucleic acids to the histone protein. The addition of exocyclic groups decreases the formation of this protein–DNA complex, while their removal increases nucleosome formation. On the other hand, recent theoretical results show a strong correlation between the BI/BII phosphate backbone conformation and the hydration of the grooves of the DNA. We performed a simulation of the d(CGCGAATTCGCG)2 Drew Dickerson dodecamer and one simulation of the d(CGCIAATTCGCG)2 dodecamer in order to investigate the influence of the exocyclic amino group of guanine. The removal of the amino group introduces a higher intrinsic flexibility to DNA supporting the suggestions that make the enhanced flexibility responsible for the enlarged histone complexation affinity. This effect is attributed to changes in the destacking interactions of both strands of the DNA. The differences in the hydration of the minor groove could be the explanation of this flexibility. The changed hydration of the minor groove also leads to a different BI/BII substate pattern. Due to the fact that the histone preferentially builds contacts with the backbone of the DNA, we propose an influence of these BI/BII changes on the nucleosome formation process. Thus, we provide an additional explanation for the enhanced affinity to the histone due to removal of exocyclic groups. In terms of BI/BII we are also able to explain how minor groove binding ligands could affect the nucleosome assembly without disrupting the structure of DNA. PMID:11812834

  18. Xeroderma Pigmentosum Group A Suppresses Mutagenesis Caused by Clustered Oxidative DNA Adducts in the Human Genome

    PubMed Central

    Sassa, Akira; Kamoshita, Nagisa; Kanemaru, Yuki; Honma, Masamitsu; Yasui, Manabu

    2015-01-01

    Clustered DNA damage is defined as multiple sites of DNA damage within one or two helical turns of the duplex DNA. This complex damage is often formed by exposure of the genome to ionizing radiation and is difficult to repair. The mutagenic potential and repair mechanisms of clustered DNA damage in human cells remain to be elucidated. In this study, we investigated the involvement of nucleotide excision repair (NER) in clustered oxidative DNA adducts. To identify the in vivo protective roles of NER, we established a human cell line lacking the NER gene xeroderma pigmentosum group A (XPA). XPA knockout (KO) cells were generated from TSCER122 cells derived from the human lymphoblastoid TK6 cell line. To analyze the mutagenic events in DNA adducts in vivo, we previously employed a system of tracing DNA adducts in the targeted mutagenesis (TATAM), in which DNA adducts were site-specifically introduced into intron 4 of thymidine kinase genes. Using the TATAM system, one or two tandem 7,8-dihydro-8-oxoguanine (8-oxoG) adducts were introduced into the genomes of TSCER122 or XPA KO cells. In XPA KO cells, the proportion of mutants induced by a single 8-oxoG (7.6%) was comparable with that in TSCER122 cells (8.1%). In contrast, the lack of XPA significantly enhanced the mutant proportion of tandem 8-oxoG in the transcribed strand (12%) compared with that in TSCER122 cells (7.4%) but not in the non-transcribed strand (12% and 11% in XPA KO and TSCER122 cells, respectively). By sequencing the tandem 8-oxoG-integrated loci in the transcribed strand, we found that the proportion of tandem mutations was markedly increased in XPA KO cells. These results indicate that NER is involved in repairing clustered DNA adducts in the transcribed strand in vivo. PMID:26559182

  19. Effects of trehalose polycation end-group functionalization on plasmid DNA uptake and transfection.

    PubMed

    Anderson, Kevin; Sizovs, Antons; Cortez, Mallory; Waldron, Chris; Haddleton, D M; Reineke, Theresa M

    2012-08-13

    In this study, we have synthesized six analogs of a trehalose-pentaethylenehexamine glycopolymer (Tr4) that contain (1A) adamantane, (1B) carboxy, (1C) alkynyl-oligoethyleneamine, (1D) azido trehalose, (1E) octyl, or (1F) oligoethyleneamine end groups and evaluated the effects of polymer end group chemistry on the ability of these systems to bind, compact, and deliver pDNA to cultured HeLa cells. The polymers were synthesized in one-pot azide-alkyne cycloaddition reactions with an adaptation of the Carothers equation for step-growth polymerization to produce a series of polymers with similar degrees of polymerization. An excess of end-capping monomer was added at the end of the polymerizations to maximize functionalization efficiency, which was evaluated with GPC, NMR, and MALDI-TOF. The polymers were all found to bind and compact pDNA at similarly low N/P ratios and form polyplexes with plasmid DNA. The effects of the different end group structures were most evident in the polyplex internalization and transfection assays in the presence of serum as determined by flow cytometry and luciferase gene expression, respectively. The Tr4 polymers end-capped with carboxyl groups (1B) (N/P = 7), octyne (1E) (N/P = 7), and oligoethyleneamine (1F) (N/P = 7), were taken into cells as polyplex and exhibited the highest levels of fluorescence, resulting from labeled plasmid. Similarly, the polymers end-functionalized with carboxyl groups (1E at N/P = 7), octyl groups (1E at N/P = 15), and in particular oligoethyleneamine groups (1F at N/P = 15) yielded dramatically higher reporter gene expression in the presence of serum. This study yields insight into how very subtle structural changes in polymer chemistry, such as end groups can yield very significant differences in the biological delivery efficiency and transgene expression of polymers used for pDNA delivery. PMID:22616977

  20. Maternal Methyl-Group Donor Intake and Global DNA (Hydroxy)Methylation before and during Pregnancy

    PubMed Central

    Pauwels, Sara; Duca, Radu Corneliu; Devlieger, Roland; Freson, Kathleen; Straetmans, Dany; Van Herck, Erik; Huybrechts, Inge; Koppen, Gurdun; Godderis, Lode

    2016-01-01

    It is still unclear to which extent methyl-group intake during pregnancy can affect maternal global DNA (hydroxyl)methylation. Pregnancy methylation profiling and its link with methyl-group intake in a healthy population could enhance our understanding of the development of pregnancy related disorders. One hundred forty-eight women were enrolled in the MANOE (MAternal Nutrition and Offspring’s Epigenome) study. Thiry-four women were enrolled before pregnancy and 116 during the first trimester of pregnancy. Global DNA (hydroxy)methylation in blood using LC-MS/MS and dietary methyl-group intake (methionine, folate, betaine, and choline) using a food-frequency questionnaire were estimated pre-pregnancy, during each trimester, and at delivery. Global DNA (hydroxy)methylation levels were highest pre-pregnancy and at weeks 18–22 of pregnancy. We observed a positive relation between folic acid and global DNA methylation (p = 0.04) and hydroxymethylation (p = 0.04). A high intake of methionine pre-pregnancy and in the first trimester showed lower (hydroxy)methylation percentage in weeks 11–13 and weeks 18–22, respectively. Choline and betaine intake in the first weeks was negatively associated with hydroxymethylation. Women with a high intake of these three methyl groups in the second and third trimester showed higher hyrdoxymethylation/methylation levels in the third trimester. To conclude, a time trend in DNA (hydroxy)methylation was found and women with higher methyl-group intake showed higher methylation in the third trimester, and not in earlier phases of pregnancy. PMID:27509522

  1. Maternal Methyl-Group Donor Intake and Global DNA (Hydroxy)Methylation before and during Pregnancy.

    PubMed

    Pauwels, Sara; Duca, Radu Corneliu; Devlieger, Roland; Freson, Kathleen; Straetmans, Dany; Van Herck, Erik; Huybrechts, Inge; Koppen, Gurdun; Godderis, Lode

    2016-01-01

    It is still unclear to which extent methyl-group intake during pregnancy can affect maternal global DNA (hydroxyl)methylation. Pregnancy methylation profiling and its link with methyl-group intake in a healthy population could enhance our understanding of the development of pregnancy related disorders. One hundred forty-eight women were enrolled in the MANOE (MAternal Nutrition and Offspring's Epigenome) study. Thiry-four women were enrolled before pregnancy and 116 during the first trimester of pregnancy. Global DNA (hydroxy)methylation in blood using LC-MS/MS and dietary methyl-group intake (methionine, folate, betaine, and choline) using a food-frequency questionnaire were estimated pre-pregnancy, during each trimester, and at delivery. Global DNA (hydroxy)methylation levels were highest pre-pregnancy and at weeks 18-22 of pregnancy. We observed a positive relation between folic acid and global DNA methylation (p = 0.04) and hydroxymethylation (p = 0.04). A high intake of methionine pre-pregnancy and in the first trimester showed lower (hydroxy)methylation percentage in weeks 11-13 and weeks 18-22, respectively. Choline and betaine intake in the first weeks was negatively associated with hydroxymethylation. Women with a high intake of these three methyl groups in the second and third trimester showed higher hyrdoxymethylation/methylation levels in the third trimester. To conclude, a time trend in DNA (hydroxy)methylation was found and women with higher methyl-group intake showed higher methylation in the third trimester, and not in earlier phases of pregnancy. PMID:27509522

  2. ssDNA Pairing Accuracy Increases When Abasic Sites Divide Nucleotides into Small Groups

    PubMed Central

    Peacock-Villada, Alexandra; Coljee, Vincent; Danilowicz, Claudia; Prentiss, Mara

    2015-01-01

    Accurate sequence dependent pairing of single-stranded DNA (ssDNA) molecules plays an important role in gene chips, DNA origami, and polymerase chain reactions. In many assays accurate pairing depends on mismatched sequences melting at lower temperatures than matched sequences; however, for sequences longer than ~10 nucleotides, single mismatches and correct matches have melting temperature differences of less than 3°C. We demonstrate that appropriately grouping of 35 bases in ssDNA using abasic sites increases the difference between the melting temperature of correct bases and the melting temperature of mismatched base pairings. Importantly, in the presence of appropriately spaced abasic sites mismatches near one end of a long dsDNA destabilize the annealing at the other end much more effectively than in systems without the abasic sites, suggesting that the dsDNA melts more uniformly in the presence of appropriately spaced abasic sites. In sum, the presence of appropriately spaced abasic sites allows temperature to more accurately discriminate correct base pairings from incorrect ones. PMID:26115175

  3. New reagents for the introduction of reactive functional groups into chemically synthesized DNA probes.

    PubMed

    Skrzypczynski, Zbigniew; Wayland, Sarah

    2003-01-01

    An efficient and versatile preparative approach is described, allowing for the preparation of DNA probes modified with an aldehyde group at the 3'- or 5'-end. The developed synthetic strategy allows for the preparation of a new family of phosphoramidites and solid supports compatible with the automated synthesis of modified oligonucleotide probes. These new reagents were prepared from intermediates 3 and 3a, obtained from the commercially available aleuritic acid 1. It was demonstrated that the new phosphoramidite reagents also could be used as new types of cleavable linkers. A new and efficient method for the production of 5' aldehyde-labeled DNA probes was developed. PMID:12757390

  4. Chimeric viruses blur the borders between the major groups of eukaryotic single-stranded DNA viruses.

    PubMed

    Roux, Simon; Enault, François; Bronner, Gisèle; Vaulot, Daniel; Forterre, Patrick; Krupovic, Mart

    2013-01-01

    Metagenomic studies have uncovered an astonishing diversity of ssDNA viruses encoding replication proteins (Reps) related to those of eukaryotic Circoviridae, Geminiviridae or Nanoviridae; however, exact evolutionary relationships among these viruses remain obscure. Recently, a unique chimeric virus (CHIV) genome, which has apparently emerged via recombination between ssRNA and ssDNA viruses, has been discovered. Here we report on the assembly of 13 new CHIV genomes recovered from various environments. Our results indicate a single event of capsid protein (CP) gene capture from an RNA virus in the history of this virus group. The domestication of the CP gene was followed by an unprecedented recurrent replacement of the Rep genes in CHIVs with distant counterparts from diverse ssDNA viruses. We suggest that parasitic and symbiotic interactions between unicellular eukaryotes were central for the emergence of CHIVs and that such turbulent evolution was primarily dictated by incongruence between the CP and Rep proteins. PMID:24193254

  5. Distinct Structural Features of the Peroxide Response Regulator from Group A Streptococcus Drive DNA Binding

    PubMed Central

    Hammel, Michal; Nix, Jay C.; Tseng, Hsiao-Ling; Tsou, Chih-Cheng; Fei, Chun-Hsien; Chiou, Huo-Sheng; Jeng, U-Ser; Lin, Yee-Shin; Chuang, Woei-Jer; Wu, Jiunn-Jong; Wang, Shuying

    2014-01-01

    Group A streptococcus (GAS, Streptococcus pyogenes) is a strict human pathogen that causes severe, invasive diseases. GAS does not produce catalase, but has an ability to resist killing by reactive oxygen species (ROS) through novel mechanisms. The peroxide response regulator (PerR), a member of ferric uptake regulator (Fur) family, plays a key role for GAS to cope with oxidative stress by regulating the expression of multiple genes. Our previous studies have found that expression of an iron-binding protein, Dpr, is under the direct control of PerR. To elucidate the molecular interactions of PerR with its cognate promoter, we have carried out structural studies on PerR and PerR-DNA complex. By combining crystallography and small-angle X-ray scattering (SAXS), we confirmed that the determined PerR crystal structure reflects its conformation in solution. Through mutagenesis and biochemical analysis, we have identified DNA-binding residues suggesting that PerR binds to the dpr promoter at the per box through a winged-helix motif. Furthermore, we have performed SAXS analysis and resolved the molecular architecture of PerR-DNA complex, in which two 30 bp DNA fragments wrap around two PerR homodimers by interacting with the adjacent positively-charged winged-helix motifs. Overall, we provide structural insights into molecular recognition of DNA by PerR and define the hollow structural arrangement of PerR-30bpDNA complex, which displays a unique topology distinct from currently proposed DNA-binding models for Fur family regulators. PMID:24586487

  6. Ternary DNA chip based on a novel thymine spacer group chemistry.

    PubMed

    Yang, Yanli; Yildiz, Umit Hakan; Peh, Jaime; Liedberg, Bo

    2015-01-01

    A novel thymine-based surface chemistry suitable for label-free electrochemical DNA detection is described. It involves a simple two-step sequential process: immobilization of 9-mer thymine-terminated probe DNAs followed by backfilling with 9-mer thymine-based spacers (T9). As compared to commonly used organic spacer groups like 2-mercaptoethanol, 3-mercapto-1-propanol and 6-mercapto-1-hexanol, the 9-mer thymine-based spacers offer a 10-fold improvement in discriminating between complementary and non-complementary target hybridization, which is due mainly to facilitated transport of the redox probes through the probe-DNA/T9 layers. Electrochemical measurements, complemented with Surface Plasmon Resonance (SPR) and Quartz Crystal Microbalance (QCM-D) binding analyses, reveal that optimum selectivity between complementary and non-complementary hybridization is obtained for a sensing surface prepared using probe-DNA and backfiller T9 at equimolar concentration (1:1). At this particular ratio, the probe-DNAs are preferentially oriented and easily accessible to yield a sensing surface with favorable hybridization and electron transfer characteristics. Our findings suggest that oligonucleotide-based spacer groups offer an attractive alternative to short organic thiol spacers in the design of future DNA biochips. PMID:25465760

  7. Mitochondrial DNA control region analysis of three ethnic groups in the Republic of Macedonia.

    PubMed

    Jankova-Ajanovska, Renata; Zimmermann, Bettina; Huber, Gabriela; Röck, Alexander W; Bodner, Martin; Jakovski, Zlatko; Janeska, Biljana; Duma, Aleksej; Parson, Walther

    2014-11-01

    A total of 444 individuals representing three ethnic groups (Albanians, Turks and Romanies) in the Republic of Macedonia were sequenced in the mitochondrial control region. The mtDNA haplogroup composition differed between the three groups. Our results showed relatively high frequencies of haplogroup H12 in Albanians (8.8%) and less in Turks (3.3%), while haplogroups M5a1 and H7a1a were dominant in Romanies (13.7% and 10.3%, respectively) but rare in the former two. This highlights the importance of regional sampling for forensic mtDNA databasing purposes. These population data will be available on EMPOP under accession numbers EMP00644 (Albanians), EMP00645 (Romanies) and EMP00646 (Turks). PMID:25051224

  8. Mitochondrial DNA control region analysis of three ethnic groups in the Republic of Macedonia

    PubMed Central

    Jankova-Ajanovska, Renata; Zimmermann, Bettina; Huber, Gabriela; Röck, Alexander W.; Bodner, Martin; Jakovski, Zlatko; Janeska, Biljana; Duma, Aleksej; Parson, Walther

    2014-01-01

    A total of 444 individuals representing three ethnic groups (Albanians, Turks and Romanies) in the Republic of Macedonia were sequenced in the mitochondrial control region. The mtDNA haplogroup composition differed between the three groups. Our results showed relatively high frequencies of haplogroup H12 in Albanians (8.8%) and less in Turks (3.3%), while haplogroups M5a1 and H7a1a were dominant in Romanies (13.7% and 10.3%, respectively) but rare in the former two. This highlights the importance of regional sampling for forensic mtDNA databasing purposes. These population data will be available on EMPOP under accession numbers EMP00644 (Albanians), EMP00645 (Romanies) and EMP00646 (Turks). PMID:25051224

  9. Targeting radiosensitizers to DNA by attachment of an intercalating group: Nitroimidazole-linked phenanthridines

    SciTech Connect

    Cowan, D.S.; Panicucci, R.; McClelland, R.A.; Rauth, A.M. )

    1991-07-01

    The nitroimidazole-linked phenanthridine series of compounds (NLP-1, 2, and 3) were synthesized under the assumption that it should be possible to enhance the molar efficiency of 2-nitroimidazoles as hypoxic cell radiosensitizers and cytotoxins by targeting them to their likely site of action, DNA. The targeting group chosen was the phenanthridine moiety, the major component of the classical DNA intercalating compound, ethidium bromide. The sole difference between the compounds is the length of the hydrocarbon chain linking the nitroimidazole to the phenanthridine. The phenanthridine group with a three-carbon side chain, P-1, was also synthesized to allow studies on the effect of the targeting group by itself. The ability of the compounds to bind to DNA is inversely proportional to their linker chain length with binding constant values ranging from approximately 1 {times} 10(5) mol-1 for NLP-2 to 6 {times} 10(5) mol-1 for NLP-3. The NLP compounds show selective toxicity to hypoxic cells at 37 degrees C at external drug concentrations 10-40 times lower than would be required for untargeted 2-nitroimidazoles such as misonidazole in vitro. Toxicity to both hypoxic and aerobic cells is dependent on the linker chain: the shorter the chain, the greater the toxicity. In addition, the NLP compounds radiosensitize hypoxic cells at external drug concentrations as low as 0.05 mM with almost the full oxygen effect being observed at a concentration of 0.5 mM. These concentrations are 10-100 times lower than would be required for similar radiosensitization using misonidazole. Radiosensitizing ability is independent of linker chain length. The present compounds represent prototypes for further studies of the efficacy and mechanism of action of 2-nitroimidazoles targeted to DNA by linkage to an intercalating group.

  10. DNA barcodes from four loci provide poor resolution of taxonomic groups in the genus Crataegus.

    PubMed

    Zarrei, Mehdi; Talent, Nadia; Kuzmina, Maria; Lee, Jeanette; Lund, Jensen; Shipley, Paul R; Stefanović, Saša; Dickinson, Timothy A

    2015-01-01

    DNA barcodes can facilitate identification of organisms especially when morphological characters are limited or unobservable. To what extent this potential is realized in specific groups of plants remains to be determined. Libraries of barcode sequences from well-studied authoritatively identified plants represented by herbarium voucher specimens are needed in order for DNA barcodes to serve their intended purpose, where this is possible, and to understand the reasons behind their failure to do so, when this occurs. We evaluated four loci, widely regarded as universal DNA barcodes for plants, for their utility in hawthorn species identification. Three plastid regions, matK, rbcLa and psbA-trnH, and the internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA discriminate only some of the species of Crataegus that can be recognized on the basis of their morphology etc. This is, in part, because in Rosaceae tribe Maleae most individual plastid loci yield relatively little taxonomic resolution and, in part, because the effects of allopolyploidization have not been eliminated by concerted evolution of the ITS regions. Although individual plastid markers provided generally poor resolution of taxonomic groups in Crataegus, a few species were notable exceptions. In contrast, analyses of concatenated sequences of the 3 plastid barcode loci plus 11 additional plastid loci gave a well-resolved maternal phylogeny. In the ITS2 tree, different individuals of some species formed groups with taxonomically unrelated species. This is a sign of lineage sorting due to incomplete concerted evolution in ITS2. Incongruence between the ITS2 and plastid trees is best explained by hybridization between different lineages within the genus. In aggregate, limited between-species variation in plastid loci, hybridization and a lack of concerted evolution in ITS2 all combine to limit the utility of standard barcoding markers in Crataegus. These results have implications for authentication

  11. DNA barcodes from four loci provide poor resolution of taxonomic groups in the genus Crataegus

    PubMed Central

    Zarrei, Mehdi; Talent, Nadia; Kuzmina, Maria; Lee, Jeanette; Lund, Jensen; Shipley, Paul R.; Stefanović, Saša; Dickinson, Timothy A.

    2015-01-01

    DNA barcodes can facilitate identification of organisms especially when morphological characters are limited or unobservable. To what extent this potential is realized in specific groups of plants remains to be determined. Libraries of barcode sequences from well-studied authoritatively identified plants represented by herbarium voucher specimens are needed in order for DNA barcodes to serve their intended purpose, where this is possible, and to understand the reasons behind their failure to do so, when this occurs. We evaluated four loci, widely regarded as universal DNA barcodes for plants, for their utility in hawthorn species identification. Three plastid regions, matK, rbcLa and psbA-trnH, and the internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA discriminate only some of the species of Crataegus that can be recognized on the basis of their morphology etc. This is, in part, because in Rosaceae tribe Maleae most individual plastid loci yield relatively little taxonomic resolution and, in part, because the effects of allopolyploidization have not been eliminated by concerted evolution of the ITS regions. Although individual plastid markers provided generally poor resolution of taxonomic groups in Crataegus, a few species were notable exceptions. In contrast, analyses of concatenated sequences of the 3 plastid barcode loci plus 11 additional plastid loci gave a well-resolved maternal phylogeny. In the ITS2 tree, different individuals of some species formed groups with taxonomically unrelated species. This is a sign of lineage sorting due to incomplete concerted evolution in ITS2. Incongruence between the ITS2 and plastid trees is best explained by hybridization between different lineages within the genus. In aggregate, limited between-species variation in plastid loci, hybridization and a lack of concerted evolution in ITS2 all combine to limit the utility of standard barcoding markers in Crataegus. These results have implications for authentication

  12. Specialization of the DNA-Cleaving Activity of a Group I Ribozyme Through In Vitro Evolution

    NASA Technical Reports Server (NTRS)

    Tsang, Joyce; Joyce, Gerald F.

    1996-01-01

    In an earlier study, an in vitro evolution procedure was applied to a large population of variants of the Tetrahymena group 1 ribozyme to obtain individuals with a 10(exp 5)-fold improved ability to cleave a target single-stranded DNA substrate under simulated physiological conditions. The evolved ribozymes also showed a twofold improvement, compared to the wild-type, in their ability to cleave a single-stranded RNA substrate. Here, we report continuation of the in vitro evolution process using a new selection strategy to achieve both enhanced DNA and diminished RNA-cleavage activity. Our strategy combines a positive selection for DNA cleavage with a negative selection against RNA binding. After 36 "generations" of in vitro evolution, the evolved population showed an approx. 100-fold increase in the ratio of DNA to RNA-cleavage activity. Site-directed mutagenesis experiment confirmed the selective advantage of two covarying mutations within the catalytic core of ribozyme that are largely responsible for this modified behavior. The population of ribozymes has now undergone a total of 63 successive generations of evolution, resulting in an average 28 mutations relative to the wild-type that are responsible for the altered phenotype.

  13. Xeroderma Pigmentosum Group A Protein Loads as a Separate Factor onto DNA Lesions

    PubMed Central

    Rademakers, Suzanne; Volker, Marcel; Hoogstraten, Deborah; Nigg, Alex L.; Moné, Martijn J.; van Zeeland, Albert A.; Hoeijmakers, Jan H. J.; Houtsmuller, Adriaan B.; Vermeulen, Wim

    2003-01-01

    Nucleotide excision repair (NER) is the main DNA repair pathway in mammals for removal of UV-induced lesions. NER involves the concerted action of more than 25 polypeptides in a coordinated fashion. The xeroderma pigmentosum group A protein (XPA) has been suggested to function as a central organizer and damage verifier in NER. How XPA reaches DNA lesions and how the protein is distributed in time and space in living cells are unknown. Here we studied XPA in vivo by using a cell line stably expressing physiological levels of functional XPA fused to green fluorescent protein and by applying quantitative fluorescence microscopy. The majority of XPA moves rapidly through the nucleoplasm with a diffusion rate different from those of other NER factors tested, arguing against a preassembled XPA-containing NER complex. DNA damage induced a transient (∼5-min) immobilization of maximally 30% of XPA. Immobilization depends on XPC, indicating that XPA is not the initial lesion recognition protein in vivo. Moreover, loading of replication protein A on NER lesions was not dependent on XPA. Thus, XPA participates in NER by incorporation of free diffusing molecules in XPC-dependent NER-DNA complexes. This study supports a model for a rapid consecutive assembly of free NER factors, and a relatively slow simultaneous disassembly, after repair. PMID:12897146

  14. SUMOylation of xeroderma pigmentosum group C protein regulates DNA damage recognition during nucleotide excision repair

    PubMed Central

    Akita, Masaki; Tak, Yon-Soo; Shimura, Tsutomu; Matsumoto, Syota; Okuda-Shimizu, Yuki; Shimizu, Yuichiro; Nishi, Ryotaro; Saitoh, Hisato; Iwai, Shigenori; Mori, Toshio; Ikura, Tsuyoshi; Sakai, Wataru; Hanaoka, Fumio; Sugasawa, Kaoru

    2015-01-01

    The xeroderma pigmentosum group C (XPC) protein complex is a key factor that detects DNA damage and initiates nucleotide excision repair (NER) in mammalian cells. Although biochemical and structural studies have elucidated the interaction of XPC with damaged DNA, the mechanism of its regulation in vivo remains to be understood in more details. Here, we show that the XPC protein undergoes modification by small ubiquitin-related modifier (SUMO) proteins and the lack of this modification compromises the repair of UV-induced DNA photolesions. In the absence of SUMOylation, XPC is normally recruited to the sites with photolesions, but then immobilized profoundly by the UV-damaged DNA-binding protein (UV-DDB) complex. Since the absence of UV-DDB alleviates the NER defect caused by impaired SUMOylation of XPC, we propose that this modification is critical for functional interactions of XPC with UV-DDB, which facilitate the efficient damage handover between the two damage recognition factors and subsequent initiation of NER. PMID:26042670

  15. High mobility group protein-mediated transcription requires DNA damage marker γ-H2AX

    PubMed Central

    Singh, Indrabahadur; Ozturk, Nihan; Cordero, Julio; Mehta, Aditi; Hasan, Diya; Cosentino, Claudia; Sebastian, Carlos; Krüger, Marcus; Looso, Mario; Carraro, Gianni; Bellusci, Saverio; Seeger, Werner; Braun, Thomas; Mostoslavsky, Raul; Barreto, Guillermo

    2015-01-01

    The eukaryotic genome is organized into chromatins, the physiological template for DNA-dependent processes including replication, recombination, repair, and transcription. Chromatin-mediated transcription regulation involves DNA methylation, chromatin remodeling, and histone modifications. However, chromatin also contains non-histone chromatin-associated proteins, of which the high-mobility group (HMG) proteins are the most abundant. Although it is known that HMG proteins induce structural changes of chromatin, the processes underlying transcription regulation by HMG proteins are poorly understood. Here we decipher the molecular mechanism of transcription regulation mediated by the HMG AT-hook 2 protein (HMGA2). We combined proteomic, ChIP-seq, and transcriptome data to show that HMGA2-induced transcription requires phosphorylation of the histone variant H2AX at S139 (H2AXS139ph; γ-H2AX) mediated by the protein kinase ataxia telangiectasia mutated (ATM). Furthermore, we demonstrate the biological relevance of this mechanism within the context of TGFβ1 signaling. The interplay between HMGA2, ATM, and H2AX is a novel mechanism of transcription initiation. Our results link H2AXS139ph to transcription, assigning a new function for this DNA damage marker. Controlled chromatin opening during transcription may involve intermediates with DNA breaks that may require mechanisms that ensure the integrity of the genome. PMID:26045162

  16. An unusual case of Streptococcus anginosus group pyomyositis diagnosed using direct 16S ribosomal DNA sequencing

    PubMed Central

    Walkty, Andrew; Embil, John M; Nichol, Kim; Karlowsky, James

    2014-01-01

    Bacteria belonging to the Streptococcus anginosus group (Streptococcus intermedius, Streptococcus constellatus and Streptococcus anginosus) are capable of causing serious pyogenic infections, with a tendency for abscess formation. The present article reports a case of S anginosus group pyomyositis in a 47-year-old man. The pathogen was recovered from one of two blood cultures obtained from the patient, but speciation was initially not performed because the organism was considered to be a contaminant (viridans streptococci group). The diagnosis was ultimately confirmed using 16S ribosomal DNA sequencing of purulent fluid obtained from a muscle abscess aspirate. The present case serves to emphasize that finding even a single positive blood culture of an organism belonging to the S anginosus group should prompt careful evaluation of the patient for a pyogenic focus of infection. It also highlights the potential utility of 16S ribosomal DNA amplification and sequencing in direct pathogen detection from aspirated fluid in cases of pyomyositis in which antimicrobial therapy was initiated before specimen collection. PMID:24634686

  17. Conformational influence of the ribose 2'-hydroxyl group: crystal structures of DNA-RNA chimeric duplexes

    NASA Technical Reports Server (NTRS)

    Egli, M.; Usman, N.; Rich, A.

    1993-01-01

    We have crystallized three double-helical DNA-RNA chimeric duplexes and determined their structures by X-ray crystallography at resolutions between 2 and 2.25 A. The two self-complementary duplexes [r(G)d(CGTATACGC)]2 and [d(GCGT)r(A)d(TACGC)]2, as well as the Okazaki fragment d(GGGTATACGC).r(GCG)d(TATACCC), were found to adopt A-type conformations. The crystal structures are non-isomorphous, and the crystallographic environments for the three chimeras are different. A number of intramolecular interactions of the ribose 2'-hydroxyl groups contribute to the stabilization of the A-conformation. Hydrogen bonds between 2'-hydroxyls and 5'-oxygens or phosphate oxygens, in addition to the previously observed hydrogen bonds to 1'-oxygens of adjacent riboses and deoxyriboses, are observed in the DNA-RNA chimeric duplexes. The crystalline chimeric duplexes do not show a transition between the DNA A- and B-conformations. CD spectra suggest that the Okazaki fragment assumes an A-conformation in solution as well. In this molecule the three RNA residues may therefore lock the complete decamer in the A-conformation. Crystals of an all-DNA strand with the same sequence as the self-complementary chimeras show a morphology which is different from those of the chimera crystals. Moreover, the oligonucleotide does not match any of the sequence characteristics of DNAs usually adopting the A-conformation in the crystalline state (e.g., octamers with short alternating stretches of purines and pyrimidines). In DNA-RNA chimeric duplexes, it is therefore possible that a single RNA residue can drive the conformational equilibrium toward the A-conformation.

  18. Increased levels of chromosomal aberrations and DNA damage in a group of workers exposed to formaldehyde.

    PubMed

    Costa, Solange; Carvalho, Sandra; Costa, Carla; Coelho, Patrícia; Silva, Susana; Santos, Luís S; Gaspar, Jorge F; Porto, Beatriz; Laffon, Blanca; Teixeira, João P

    2015-07-01

    Formaldehyde (FA) is a commonly used chemical in anatomy and pathology laboratories as a tissue preservative and fixative. Because of its sensitising properties, irritating effects and cancer implication, FA accounts probably for the most important chemical-exposure hazard concerning this professional group. Evidence for genotoxic effects and carcinogenic properties in humans is insufficient and conflicting, particularly in regard to the ability of inhaled FA to induce toxicity on other cells besides first contact tissues, such as buccal and nasal cells. To evaluate the effects of exposure to FA in human peripheral blood lymphocytes, a group of 84 anatomy pathology laboratory workers exposed occupationally to FA and 87 control subjects were tested for chromosomal aberrations (CAs) and DNA damage (comet assay). The level of exposure to FA in the workplace air was evaluated. The association between genotoxicity biomarkers and polymorphic genes of xenobiotic-metabolising and DNA repair enzymes were also assessed. The estimated mean level of FA exposure was 0.38±0.03 ppm. All cytogenetic endpoints assessed by CAs test and comet assay % tail DNA (%TDNA) were significantly higher in FA-exposed workers compared with controls. Regarding the effect of susceptibility biomarkers, results suggest that polymorphisms in CYP2E1 and GSTP1 metabolic genes, as well as, XRCC1 and PARP1 polymorphic genes involved in DNA repair pathways are associated with higher genetic damage in FA-exposed subjects. Data obtained in this study show a potential health risk situation of anatomy pathology laboratory workers exposed to FA (0.38 ppm). Implementation of security and hygiene measures may be crucial to decrease risk. The obtained information may also provide new important data to be used by health care programs and by governmental agencies responsible for occupational health and safety. PMID:25711496

  19. Capture-recapture of white-tailed deer using DNA from fecal pellet-groups

    USGS Publications Warehouse

    Goode, Matthew J; Beaver, Jared T; Muller, Lisa I; Clark, Joseph D.; van Manen, Frank T.; Harper, Craig T; Basinger, P Seth

    2014-01-01

    Traditional methods for estimating white-tailed deer population size and density are affected by behavioral biases, poor detection in densely forested areas, and invalid techniques for estimating effective trapping area. We evaluated a noninvasive method of capture—recapture for white-tailed deer (Odocoileus virginianus) density estimation using DNA extracted from fecal pellets as an individual marker and for gender determination, coupled with a spatial detection function to estimate density (spatially explicit capture—recapture, SECR). We collected pellet groups from 11 to 22 January 2010 at randomly selected sites within a 1-km2 area located on Arnold Air Force Base in Coffee and Franklin counties, Tennessee. We searched 703 10-m radius plots and collected 352 pellet-group samples from 197 plots over five two-day sampling intervals. Using only the freshest pellets we recorded 140 captures of 33 different animals (15M:18F). Male and female densities were 1.9 (SE = 0.8) and 3.8 (SE = 1.3) deer km-2, or a total density of 5.8 deer km-2 (14.9 deer mile-2). Population size was 20.8 (SE = 7.6) over a 360-ha area, and sex ratio was 1.0 M: 2.0 F (SE = 0.71). We found DNA sampling from pellet groups improved deer abundance, density and sex ratio estimates in contiguous landscapes which could be used to track responses to harvest or other management actions.

  20. Evaluation of simplified DNA extraction methods for emm typing of group A streptococci.

    PubMed

    Jose, J J M; Brahmadathan, K N

    2006-04-01

    Simplified methods of DNA extraction for amplification and sequencing for emm typing of group A streptococci (GAS) can save valuable time and cost in resource crunch situations. To evaluate this, we compared two methods of DNA extraction directly from colonies with the standard CDC cell lysate method for emm typing of 50 GAS strains isolated from children with pharyngitis and impetigo. For this, GAS colonies were transferred into two sets of PCR tubes. One set was preheated at 94 degrees C for two minutes in the thermal cycler and cooled while the other set was frozen overnight at -20 degrees C and then thawed before adding the PCR mix. For the cell lysate method, cells were treated with mutanolysin and hyaluronidase before heating at 100 degrees C for 10 minutes and cooling immediately as recommended in the CDC method. All 50 strains could be typed by sequencing the hyper variable region of the emm gene after amplification. The quality of sequences and the emm types identified were also identical. Our study shows that the two simplified DNA extraction methods directly from colonies can conveniently be used for typing a large number of GAS strains easily in relatively short time. PMID:16687865

  1. SIV DNA vaccine trial in macaques: post-challenge necropsy in vaccine and control groups.

    PubMed

    Lu, S; Manson, K; Wyand, M; Robinson, H L

    1997-06-01

    In this study we describe the histopathologic findings from nine macaques in a simian immunodeficiency virus (SIV) DNA vaccine trial evaluating the ability of a 5-plasmid DNA vaccine to protect against an uncloned SIVmac251 challenge (Lu et al., J. Virol. 1996, 70, 3978-3991). Three vaccinated and one control macaque developed disease and were euthanized in the first year following challenge. The other four vaccinated and one control macaque remained clinically normal and were euthanized at the end of the trial (60 weeks post-challenge). The necropsy data revealed that both diseased and clinically normal macaques had developed typical SIV-related lymphoid changes, inflammatory disorders and opportunistic infections. All animals had variable degrees of follicular and/or paracortical lymphoid hyperplasia suggesting immune activation. All but one vaccinated macaque and both control macaques had SIV-associated opportunistic infections. Within the small groups of animals, the ability to contain opportunistic infections was superior, and the overall lymphoid changes less severe, in the macaques that had received vaccine DNAs by three routes of inoculation (intravenous, intramuscular and gene gun) than in those that had received control DNAs or vaccine DNAs by gene gun only. In the future it will be important to further test how the route and method of DNA inoculation impact the efficacy of immunodeficiency virus vaccines. PMID:9234548

  2. Genetic Variation Among Vegetative Compatibility Groups of Fusarium oxysporum f. sp. cubense Analyzed by DNA Fingerprinting.

    PubMed

    Bentley, S; Pegg, K G; Moore, N Y; Davis, R D; Buddenhagen, I W

    1998-12-01

    ABSTRACT Genetic variation within a worldwide collection of 208 isolates of Fu-sarium oxysporum f. sp. cubense, representing physiological races 1, 2, 3, and 4 and the 20 reported vegetative compatibility groups (VCGs), was analyzed using modified DNA amplification fingerprinting. Also characterized were 133 isolates that did not belong to any of the reported VCGs of F. oxysporum f. sp. cubense including race 3 isolates from a Heliconia species and isolates from a symptomatic wild banana species growing in the jungle in peninsular Malaysia. The DNA fingerprint patterns were generally VCG specific, irrespective of geographic or host origin. A total of 33 different genotypes were identified within F. oxysporum f. sp. cu-bense; 19 genotypes were distinguished among the isolates that belonged to the 20 reported VCGs, and 14 new genotypes were identified among the isolates that did not belong to any of the existing VCGs. DNA fingerprinting analysis also allowed differentiation of nine clonal lineages within F. oxysporum f. sp. cubense. Five of these lineages each contained numerous closely related VCGs and genotypes, and the remaining four lineages each contained a single genotype. The genetic diversity and geographic distribution of several of these lineages of F. oxysporum f. sp. cubense suggests that they have coevolved with edible bananas and their wild diploid progenitors in Asia. DNA fingerprinting analysis of isolates from the wild pathosystem provides further evidence for the coevolution hypothesis. The genetic isolation and limited geographic distribution of four of the lineages of F. oxysporum f. sp. cubense suggests that the pathogen has also arisen independently, both within and outside of the center of origin of the host. PMID:18944830

  3. Yeast RAD14 and human xeroderma pigmentosum group A DNA-repair genes encode homologous proteins.

    PubMed

    Bankmann, M; Prakash, L; Prakash, S

    1992-02-01

    Xeroderma pigmentosum (XP), a human autosomal recessive disorder, is characterized by extreme sensitivity to sunlight and high incidence of skin cancers. XP cells are defective in the incision step of excision repair of DNA damaged by ultraviolet light. Cell fusion studies have defined seven XP complementation groups, XP-A to XP-G. Similar genetic complexity of excision repair is observed in the yeast Saccharomyces cerevisiae. Mutations in any one of five yeast genes, RAD1, RAD2, RAD3, RAD4, and RAD10, cause a total defect in incision and an extreme sensitivity to ultraviolet light. Here we report the characterization of the yeast RAD14 gene. The available rad14 point mutant is only moderately ultraviolet-sensitive, and it performs a substantial amount of incision of damaged DNA. Our studies with the rad14 deletion (delta) mutation indicate an absolute requirement of RAD14 in incision. RAD14 encodes a highly hydrophilic protein of 247 amino acids containing zinc-finger motifs, and it is similar to the protein encoded by the human XPAC gene that complements XP group A cell lines. PMID:1741034

  4. Conducting polymer based DNA biosensor for the detection of the Bacillus cereus group species

    NASA Astrophysics Data System (ADS)

    Velusamy, Vijayalakshmi; Arshak, Khalil; Korostynska, Olga; Oliwa, Kamila; Adley, Catherine

    2009-05-01

    Biosensor designs are emerging at a significant rate and play an increasingly important role in foodborne pathogen detection. Conducting polymers are excellent tools for the fabrication of biosensors and polypyrrole has been used in the detection of biomolecules due to its unique properties. The prime intention of this paper was to pioneer the design and fabrication of a single-strand (ss) DNA biosensor for the detection of the Bacillus cereus (B.cereus) group species. Growth of B. cereus, results in production of several highly active toxins. Therefore, consumption of food containing >106 bacteria/gm may results in emetic and diarrhoeal syndromes. The most common source of this bacterium is found in liquid food products, milk powder, mixed food products and is of particular concern in the baby formula industry. The electrochemical deposition technique, such as cyclic voltammetry, was used to develop and test a model DNA-based biosensor on a gold electrode electropolymerized with polypyrrole. The electrically conducting polymer, polypyrrole is used as a platform for immobilizing DNA (1μg) on the gold electrode surface, since it can be more easily deposited from neutral pH aqueous solutions of pyrrolemonomers. The average current peak during the electrodeposition event is 288μA. There is a clear change in the current after hybridization of the complementary oligonucleotide (6.35μA) and for the noncomplementary oligonucleotide (5.77μA). The drop in current after each event was clearly noticeable and it proved to be effective.

  5. Transferring the purine 2-amino group from guanines to adenines in DNA changes the sequence-specific binding of antibiotics.

    PubMed Central

    Bailly, C; Waring, M J

    1995-01-01

    The proposition that the 2-amino group of guanine plays a critical role in determining how antibiotics recognise their binding sites in DNA has been tested by relocating it, using tyrT DNA derivative molecules substituted with inosine plus 2,6-diaminopurine (DAP). Irrespective of their mode of interaction with DNA, such GC-specific antibiotics as actinomycin, echinomycin, mithramycin and chromomycin find new binding sites associated with DAP-containing sequences and are excluded from former canonical sites containing I.C base pairs. The converse is found to be the case for a group of normally AT-selective ligands which bind in the minor groove of the helix, such as netropsin: their preferred sites become shifted to IC-rich clusters. Thus the binding sites of all these antibiotics strictly follow the placement of the purine 2-amino group, which accordingly must serve as both a positive and negative effector. The footprinting profile of the 'threading' intercalator nogalamycin is potentiated in DAP plus inosine-substituted DNA but otherwise remains much the same as seen with natural DNA. The interaction of echinomycin with sites containing the TpDAP step in doubly substituted DNA appears much stronger than its interaction with CpG-containing sites in natural DNA. Images PMID:7731800

  6. Chloroplast DNA variation and geographical structure of the Aristolochia kaempferi group (Aristolochiaceae).

    PubMed

    Watanabe, Kana; Kajita, Tadashi; Murata, Jin

    2006-03-01

    The present study documents cpDNA variation in the Aristolochia kaempferi group (Aristolochiaceae), which consists of one Chinese and all Japanese and Taiwanese species of the subgenus Siphisia. In a phylogenetic analysis based on the nucleotide sequences of the matK gene, and the atpB-rbcL and trnS-trnG intergenic spacer regions, 38 haplotypes were recognized in the A. kaempferi group and as many as 24 within A. kaempferi. This is the most haplotypes reported for a single species to date. Although six highly significant major clades were identified in the phylogenetic analysis, they were not congruent with previous classifications. This might be attributed to the specific speciation process, such as convergent evolution, incomplete lineage sorting, and/or reticulate evolution. The six major clades had a clear geographical distribution pattern and were significantly associated with geographical distribution of haplotypes in a nested clade analysis and AMOVA. The results allow us to deduce a scenario in which multiple contractions and expansions of the geographical ranges brought about by Quaternary climatic oscillations affected the patterns of genetic diversity. The present geographic patterns of haplotype distribution within the A. kaempferi group can be explained by the last postglacial range expansion from different refugia, and the boundaries may be suture zones. PMID:21646203

  7. Increased UV resistance in xeroderma pigmentosum group A cells after transformation with a human genomic DNA clone.

    PubMed Central

    Rinaldy, A; Bellew, T; Egli, E; Lloyd, R S

    1990-01-01

    Xeroderma pigmentosum (XP) is an autosomal recessive disease in which the major clinical manifestation is a 2,000-fold enhanced probability of developing sunlight-induced skin tumors, and the molecular basis for the disease is a defective DNA excision repair system. To clone the gene defective in XP complementation group A (XP-A), cDNA clones were isolated by a competition hybridization strategy in which the corresponding mRNAs were more abundant in cells of the obligately heterozygous parents relative to cells of the homozygous proband affected with the disease. In this report, a human genomic DNA clone that contains this cDNA was transformed into two independent homozygous XP-A cell lines, and these transformants displayed partial restoration of resistance to the killing effects of UV irradiation. The abundance of mRNA corresponding to this cDNA appears to correlate well with the observed UV cell survival. The results of unscheduled DNA synthesis after UV exposure indicate that the transformed cells are repair proficient relative to that of the control XP-A cells. However, using this same genomic DNA, transformation of an XP-F cell line did not confer any enhancement of UV survival or promote unscheduled DNA synthesis after UV exposure. Images PMID:2168562

  8. A Human Orthologue of Archaeal DNA Repair Protein Hef is Defective in Fanconi Anemia Complementation Group M

    PubMed Central

    Meetei, Amom Ruhikanta; Medhurst, Annette L.; Ling, Chen; Xue, Yutong; Singh, Thiyam Ramsing; Bier, Patrick; Steltenpool, Jurgen; Stone, Stacie; Dokal, Inderjeet; Mathew, Christopher G.; Hoatlin, Maureen; Joenje, Hans; de Winter, Johan P.; Wang, Weidong

    2005-01-01

    Fanconi anemia (FA) is a genetic disease featuring genomic instability and cancer predisposition1. Nine FA genes have been identified, and their products participate in a DNA damage response network involving BRCA1 and BRCA22,3. We have previously purified a FA core complex containing the FANCL ubiquitin ligase and 6 other FA proteins4–6. Each protein in this complex is essential for monoubiquitination of FANCD2, a key reaction in the FA DNA damage response pathway2,7. Here we show that another component of this complex, FAAP250, is mutated in FA patients of a new complementation group (FA-M). FAAP250, renamed FANCM, has sequence similarity to known DNA repair proteins, including archaeal Hef, yeast Mph1 and human ERCC4/XPF. FANCM can dissociate DNA triplex, possibly due to its ability to translocate on duplex DNA. FANCM is essential for FANCD2 monoubiquitination and becomes hyperphosphorylated in response to DNA damage. Our data suggest an evolutionary link between FA proteins and DNA repair; FANCM may act as an engine that translocates the FA core complex along DNA. PMID:16116422

  9. Acinetobacter variabilis sp. nov. (formerly DNA group 15 sensu Tjernberg & Ursing), isolated from humans and animals.

    PubMed

    Krizova, Lenka; McGinnis, Jana; Maixnerova, Martina; Nemec, Matej; Poirel, Laurent; Mingle, Lisa; Sedo, Ondrej; Wolfgang, William; Nemec, Alexandr

    2015-03-01

    We aimed to define the taxonomic status of 16 strains which were phenetically congruent with Acinetobacter DNA group 15 described by Tjernberg & Ursing in 1989. The strains were isolated from a variety of human and animal specimens in geographically distant places over the last three decades. Taxonomic analysis was based on an Acinetobacter-targeted, genus-wide approach that included the comparative sequence analysis of housekeeping, protein-coding genes, whole-cell profiling based on matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS), an array of in-house physiological and metabolic tests, and whole-genome comparative analysis. Based on analyses of the rpoB and gyrB genes, the 16 strains formed respective, strongly supported clusters clearly separated from the other species of the genus Acinetobacter. The distinctness of the group at the species level was indicated by average nucleotide identity values of ≤82 % between the whole genome sequences of two of the 16 strains (NIPH 2171(T) and NIPH 899) and those of the known species. In addition, the coherence of the group was also supported by MALDI-TOF MS. All 16 strains were non-haemolytic and non-gelatinase-producing, grown at 41 °C and utilized a rather limited number of carbon sources. Virtually every strain displayed a unique combination of metabolic and physiological features. We conclude that the 16 strains represent a distinct species of the genus Acinetobacter, for which the name Acinetobacter variabilis sp. nov. is proposed to reflect its marked phenotypic heterogeneity. The type strain is NIPH 2171(T) ( = CIP 110486(T) = CCUG 26390(T) = CCM 8555(T)). PMID:25510976

  10. Acinetobacter variabilis sp. nov. (formerly DNA group 15 sensu Tjernberg & Ursing), isolated from humans and animals

    PubMed Central

    Krizova, Lenka; McGinnis, Jana; Maixnerova, Martina; Nemec, Matej; Poirel, Laurent; Mingle, Lisa; Sedo, Ondrej; Wolfgang, William

    2015-01-01

    We aimed to define the taxonomic status of 16 strains which were phenetically congruent with Acinetobacter DNA group 15 described by Tjernberg & Ursing in 1989. The strains were isolated from a variety of human and animal specimens in geographically distant places over the last three decades. Taxonomic analysis was based on an Acinetobacter-targeted, genus-wide approach that included the comparative sequence analysis of housekeeping, protein-coding genes, whole-cell profiling based on matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS), an array of in-house physiological and metabolic tests, and whole-genome comparative analysis. Based on analyses of the rpoB and gyrB genes, the 16 strains formed respective, strongly supported clusters clearly separated from the other species of the genus Acinetobacter. The distinctness of the group at the species level was indicated by average nucleotide identity values of ≤82 % between the whole genome sequences of two of the 16 strains (NIPH 2171T and NIPH 899) and those of the known species. In addition, the coherence of the group was also supported by MALDI-TOF MS. All 16 strains were non-haemolytic and non-gelatinase-producing, grown at 41 °C and utilized a rather limited number of carbon sources. Virtually every strain displayed a unique combination of metabolic and physiological features. We conclude that the 16 strains represent a distinct species of the genus Acinetobacter, for which the name Acinetobacter variabilis sp. nov. is proposed to reflect its marked phenotypic heterogeneity. The type strain is NIPH 2171T ( = CIP 110486T = CCUG 26390T = CCM 8555T). PMID:25510976

  11. Yeast high mobility group protein HMO1 stabilizes chromatin and is evicted during repair of DNA double strand breaks

    PubMed Central

    Panday, Arvind; Xiao, LiJuan; Grove, Anne

    2015-01-01

    DNA is packaged into condensed chromatin fibers by association with histones and architectural proteins such as high mobility group (HMGB) proteins. However, this DNA packaging reduces accessibility of enzymes that act on DNA, such as proteins that process DNA after double strand breaks (DSBs). Chromatin remodeling overcomes this barrier. We show here that the Saccharomyces cerevisiae HMGB protein HMO1 stabilizes chromatin as evidenced by faster chromatin remodeling in its absence. HMO1 was evicted along with core histones during repair of DSBs, and chromatin remodeling events such as histone H2A phosphorylation and H3 eviction were faster in absence of HMO1. The facilitated chromatin remodeling in turn correlated with more efficient DNA resection and recruitment of repair proteins; for example, inward translocation of the DNA-end-binding protein Ku was faster in absence of HMO1. This chromatin stabilization requires the lysine-rich C-terminal extension of HMO1 as truncation of the HMO1 C-terminal tail phenocopies hmo1 deletion. Since this is reminiscent of the need for the basic C-terminal domain of mammalian histone H1 in chromatin compaction, we speculate that HMO1 promotes chromatin stability by DNA bending and compaction imposed by its lysine-rich domain and that it must be evicted along with core histones for efficient DSB repair. PMID:25979266

  12. Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum.

    PubMed Central

    Tanaka, K; Satokata, I; Ogita, Z; Uchida, T; Okada, Y

    1989-01-01

    For isolation of the gene responsible for xeroderma pigmentosum (XP) complementation group A, plasmid pSV2gpt and genomic DNA from a mouse embryo were cotransfected into XP2OSSV cells, a group-A XP cell line. Two primary UV-resistant XP transfectants were isolated from about 1.6 X 10(5) pSV2gpt-transformed XP colonies. pSV2gpt and genomic DNA from the primary transfectants were again cotransfected into XP2OSSV cells and a secondary UV-resistant XP transfectant was obtained by screening about 4.8 X 10(5) pSV2gpt-transformed XP colonies. The secondary transfectant retained fewer mouse repetitive sequences. A mouse gene that complements the defect of XP2OSSV cells was cloned into an EMBL3 vector from the genome of a secondary transfectant. Transfections of the cloned DNA also conferred UV resistance on another group-A XP cell line but not on XP cell lines of group C, D, F, or G. Northern blot analysis of poly(A)+ RNA with a subfragment of cloned mouse DNA repair gene as the probe revealed that an approximately 1.0 kilobase mRNA was transcribed in the donor mouse embryo and secondary transfectant, and approximately 1.0- and approximately 1.3-kilobase mRNAs were transcribed in normal human cells, but none of these mRNAs was detected in three strains of group-A XP cells. These results suggest that the cloned DNA repair gene is specific for group-A XP and may be the mouse homologue of the group-A XP human gene. Images PMID:2748601

  13. Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum

    SciTech Connect

    Tanaka, K.; Satokata, I.; Ogita, Z.; Uchida, T.; Okada, Y.

    1989-07-01

    For isolation of the gene responsible for xeroderma pigmentosum (XP) complementation group A, plasmid pSV2gpt and genomic DNA from a mouse embryo were cotransfected into XP2OSSV cells, a group-A XP cell line. Two primary UV-resistant XP transfectants were isolated from about 1.6 X 10(5) pSV2gpt-transformed XP colonies. pSV2gpt and genomic DNA from the primary transfectants were again cotransfected into XP2OSSV cells and a secondary UV-resistant XP transfectant was obtained by screening about 4.8 X 10(5) pSV2gpt-transformed XP colonies. The secondary transfectant retained fewer mouse repetitive sequences. A mouse gene that complements the defect of XP2OSSV cells was cloned into an EMBL3 vector from the genome of a secondary transfectant. Transfections of the cloned DNA also conferred UV resistance on another group-A XP cell line but not on XP cell lines of group C, D, F, or G. Northern blot analysis of poly(A)+ RNA with a subfragment of cloned mouse DNA repair gene as the probe revealed that an approximately 1.0 kilobase mRNA was transcribed in the donor mouse embryo and secondary transfectant, and approximately 1.0- and approximately 1.3-kilobase mRNAs were transcribed in normal human cells, but none of these mRNAs was detected in three strains of group-A XP cells. These results suggest that the cloned DNA repair gene is specific for group-A XP and may be the mouse homologue of the group-A XP human gene.

  14. Threonine phosphorylation prevents promoter DNA binding of the Group B Streptococcus response regulator CovR.

    PubMed

    Lin, Wan-Jung; Walthers, Don; Connelly, James E; Burnside, Kellie; Jewell, Kelsea A; Kenney, Linda J; Rajagopal, Lakshmi

    2009-03-01

    All living organisms communicate with the external environment for their survival and existence. In prokaryotes, communication is achieved by two-component systems (TCS) comprising histidine kinases and response regulators. In eukaryotes, signalling is accomplished by serine/threonine and tyrosine kinases. Although TCS and serine/threonine kinases coexist in prokaryotes, direct cross-talk between these families was first described in Group B Streptococcus (GBS). A serine/threonine kinase (Stk1) and a TCS (CovR/CovS) co-regulate toxin expression in GBS. Typically, promoter binding of regulators like CovR is controlled by phosphorylation of the conserved active site aspartate (D53). In this study, we show that Stk1 phosphorylates CovR at threonine 65. The functional consequence of threonine phosphorylation of CovR in GBS was evaluated using phosphomimetic and silencing substitutions. GBS encoding the phosphomimetic T65E allele are deficient for CovR regulation unlike strains encoding the non-phosphorylated T65A allele. Further, compared with wild-type or T65A CovR, the T65E CovR is unable to bind promoter DNA and is decreased for phosphorylation at D53, similar to Stk1-phosphorylated CovR. Collectively, we provide evidence for a novel mechanism of response regulator control that enables GBS (and possibly other prokaryotes) to fine-tune gene expression for environmental adaptation. PMID:19170889

  15. The high-mobility group protein T160 binds to both linear and cruciform DNA and mediates DNA bending as determined by ring closure.

    PubMed

    Gariglio, M; Ying, G G; Hertel, L; Gaboli, M; Clerc, R G; Landolfo, S

    1997-11-01

    The high-mobility group protein T160 was isolated by screening a phage library from a murine pre-B-cell line L1210. South-Western experiments have previously shown that this protein binds to V-(D)-J recombination signal sequences, suggesting that it may be a sequence-specific DNA-binding protein. However, neither gel-shift nor footprinting analyses have been successfully employed with the T160 protein, despite an extensive effort. In this study, the T160 protein or truncated forms made soluble through denaturing and renaturing cycles in urea were successfully used in gel-shift experiments showing that T160 binds to cruci-form or linear duplex DNA with no apparent sequence specificity. Furthermore, fragments longer than 100 bp efficiently formed covalently closed circular monomers in the presence of T160 and T4 DNA ligase, indicating that the protein is capable of introducing bends into the duplex. Last, tissue distribution by Western blotting analysis showed that the T160 protein is expressed in various murine tissues in addition to those of lymphoid origin. Considering its broad evolutionary conservation (from plants to mammals) also, these results suggest that the functional role of the T160 protein is not limited to V-(D)-J recombination, but might be involved in basic processes such as DNA replication and repairing, where irregular DNA structures are generated and very likely recognized by HMG domain proteins. PMID:9367632

  16. Pyramidal and Chiral Groupings of Gold Nanocrystals Assembled Using DNA Scaffolds

    SciTech Connect

    Mastroianni, Alexander; Claridge, Shelley; Alivisatos, A. Paul

    2009-03-30

    Nanostructures constructed from metal and semiconductor nanocrystals conjugated to, and organized by DNA are an emerging class of material with collective optical properties. We created discrete pyramids of DNA with gold nanocrystals at the tips. By taking small angle X-ray scattering (SAXS) measurments from solutions of these pyramids we confirmed that this pyramidal geometry creates structures which are more rigid in solution than linear DNA. We then took advantage of the tetrahedral symmetry to demonstrate construction of chiral nanostructures.

  17. The Q motif of Fanconi anemia group J protein (FANCJ) DNA helicase regulates its dimerization, DNA binding, and DNA repair function.

    PubMed

    Wu, Yuliang; Sommers, Joshua A; Loiland, Jason A; Kitao, Hiroyuki; Kuper, Jochen; Kisker, Caroline; Brosh, Robert M

    2012-06-22

    The Q motif, conserved in a number of RNA and DNA helicases, is proposed to be important for ATP binding based on structural data, but its precise biochemical functions are less certain. FANCJ encodes a Q motif DEAH box DNA helicase implicated in Fanconi anemia and breast cancer. A Q25A mutation of the invariant glutamine in the Q motif abolished its ability to complement cisplatin or telomestatin sensitivity of a fancj null cell line and exerted a dominant negative effect. Biochemical characterization of the purified recombinant FANCJ-Q25A protein showed that the mutation disabled FANCJ helicase activity and the ability to disrupt protein-DNA interactions. FANCJ-Q25A showed impaired DNA binding and ATPase activity but displayed ATP binding and temperature-induced unfolding transition similar to FANCJ-WT. Size exclusion chromatography and sedimentation velocity analyses revealed that FANCJ-WT existed as molecular weight species corresponding to a monomer and a dimer, and the dimeric form displayed a higher specific activity for ATPase and helicase, as well as greater DNA binding. In contrast, FANCJ-Q25A existed only as a monomer, devoid of helicase activity. Thus, the Q motif is essential for FANCJ enzymatic activity in vitro and DNA repair function in vivo. PMID:22582397

  18. DNA binding and photocleavage specificities of a group of tricationic metalloporphyrins

    NASA Astrophysics Data System (ADS)

    Zhao, Ping; Huang, Jin-Wang; Mei, Wen-Jie; He, Juan; Ji, Liang-Nian

    2010-03-01

    The interactions of 5,10,15-tris(1-methylpyridinium-4-yl)-20-(4-hydroxyphenyl)porphyrinatozinc(II) Zn[TMPyHP] 3+ ( 2) along with Cu[TMPyHP] 3+ ( 3), Co[TMPyHP] 4+ ( 4), Mn[TMPyHP] 4+ ( 5) and the free base porphyrin H 2[TMPyHP] 3+ ( 1) with duplex DNA have been studied by using a combination of absorption, fluorescence titration, surface-enhanced Raman spectroscopy (SERS), induced circular dichroism (ICD) spectroscopy, thermal DNA denaturation, viscosity measurements as well as gel electrophoresis experiment. Their binding modes and intrinsic binding constants ( Kb) to calf DNA (CT DNA) were comparatively studied and were found significantly influenced by different metals coordinated with the porphyrin plane. Except 3, which has four-coordination structure at the metal, all the metal derivatives showed non-intercalative DNA-binding mode and lower Kb than the free base porphyrin 1, most probably due to the steric hindrance results from the axial ligands of the inserted metals which are five or six-coordination structures. Meanwhile, the insertion of metals into cationic porphyrin greatly removed the self-aggregation of the metal-free porphyrins, and thus fully enhanced the singlet oxygen ( 1O 2) productivities in the DNA photocleavage experiments. Therefore, these metalloporphyrins have comparable DNA cleavage ability with the free base porphyrin.

  19. DNA.

    ERIC Educational Resources Information Center

    Felsenfeld, Gary

    1985-01-01

    Structural form, bonding scheme, and chromatin structure of and gene-modification experiments with deoxyribonucleic acid (DNA) are described. Indicates that DNA's double helix is variable and also flexible as it interacts with regulatory and other molecules to transfer hereditary messages. (DH)

  20. Remarkable stabilization of antiparallel DNA triplexes by strong stacking effects of consecutively modified nucleobases containing thiocarbonyl groups.

    PubMed

    Yamada, Kenji; Hattori, Yusaku; Inde, Takeshi; Kanamori, Takashi; Ohkubo, Akihiro; Seio, Kohji; Sekine, Mitsuo

    2013-02-01

    The consecutive arrangement of 2'-deoxy-6-thioguanosines (s(6)Gs) and 4-thiothymidines (s(4)Ts) in antiparallel triplex-forming oligonucleotides (TFOs) considerably stabilized the resulting antiparallel triplexes with high base recognition ability by the strong stacking effects of thiocarbonyl groups. This result was remarkable because chemical modifications of the sugar moieties and nucleobases of antiparallel TFOs generally destabilize triplex structures. Moreover, in comparison with unmodified TFOs, it was found that TFOs containing s(6)Gs and s(4)Ts could selectively bind to the complementary DNA duplex but not to mismatched DNA duplexes or single-stranded RNA. PMID:23287737

  1. Crystal structure of the trithorax group protein ASH2L reveals a forkhead-like DNA binding domain

    SciTech Connect

    Sarvan, Sabina; Avdic, Vanja; Tremblay, Véronique; Chaturvedi, Chandra-Prakash; Zhang, Pamela; Lanouette, Sylvain; Blais, Alexandre; Brunzelle, Joseph S; Brand, Marjorie; Couture, Jean-François

    2012-05-02

    Absent, small or homeotic discs-like 2 (ASH2L) is a trithorax group (TrxG) protein and a regulatory subunit of the SET1 family of lysine methyltransferases. Here we report that ASH2L binds DNA using a forkhead-like helix-wing-helix (HWH) domain. In vivo, the ASH2L HWH domain is required for binding to the {beta}-globin locus control region, histone H3 Lys4 (H3K4) trimethylation and maximal expression of the {beta}-globin gene (Hbb-1), validating the functional importance of the ASH2L DNA binding domain.

  2. Thirteen Virulent and Temperate Bacteriophages of Lactobacillus bulgaricus and Lactobacillus lactis Belong to a Single DNA Homology Group

    PubMed Central

    Mata, Mireille; Trautwetter, Annie; Luthaud, Gisèle; Ritzenthaler, Paul

    1986-01-01

    Thirteen virulent phages and two temperate phages of two closely related bacterial species (Lactobacillus lactis and L. bulgaricus) were compared for their protein composition, their antigenic properties, their restriction endonuclease patterns, and their DNA homology. The immunoblotting studies and the DNA-DNA hybridizations showed that the phages could be differentiated into two groups. One group contained 2 temperate phages of L. bulgaricus and 11 virulent phages of L. lactis. Inside each group, at least two common proteins of identical sizes could be detected for each phage. These proteins were able to cross-react in immunoblotting experiments with an antiserum raised against one phage of the same group. Temperate phage DNAs showed partial homology with DNAs from some virulent phages. These homologies seem to be located on the region coding for the structural proteins since recombinant plasmids coding for one of the major phage proteins of one phage were able to hybridize with the DNAs from phages of the same group. These results suggest that temperate and virulent phages may be related to one another. Images PMID:16347174

  3. Differentiation and grouping of isolates of the Ganoderma lucidum complex by random amplified polymorphic DNA-PCR compared with grouping on the basis of internal transcribed spacer sequences.

    PubMed Central

    Hseu, R S; Wang, H H; Wang, H F; Moncalvo, J M

    1996-01-01

    Laccate polypores of the Ganoderma lucidum species complex are widespread white rot fungi of economic importance, but isolates cannot be identified by traditional taxonomic methods. Parsimony analysis of nucleotide sequences from the internal transcribed spacers (ITS) of the ribosomal gene (rDNA) distinguished six lineages in this species complex. Each ITS lineage may represent one or more putative species. While some isolates have identical ITS sequences, all of them could be clearly differentiated by genetic fingerprinting using random amplified polymorphic DNA (RAPD). To investigate the suitability of RAPD markers for taxonomic identification and grouping of isolates of the G. lucidum complex, RAPD fragments (RAPDs) were used as phenotypic characters in numerical and parsimony analyses. Results show that data from RAPDS do not distinguish the same clades as ITS data do. Groupings based on analysis of RAPD data were very sensitive to the choice of the grouping method used, and no consistent grouping of isolates could be proposed. However, analysis with RAPDs did resolve several robust terminal clades containing putatively conspecific isolates, suggesting that RAPDs might be helpful for systematics at the lower taxonomic levels that are unresolved by ITS sequence data. The limitations of RAPDs for systematics are briefly discussed. The conclusion of this study is that ITS sequences can be used to identify isolates of the G. lucidum complex, whereas RAPDs can be used to differentiate between isolates having identical ITS sequences. The practical implications of these results are briefly illustrated. PMID:8919797

  4. Aeromonas jandaei (formerly genospecies DNA group 9 A. sobria), a new sucrose-negative species isolated from clinical specimens.

    PubMed Central

    Carnahan, A; Fanning, G R; Joseph, S W

    1991-01-01

    A large numerical taxonomy study conducted in 1988 of 165 mostly clinical Aeromonas strains from diverse geographic sources produced a cluster (S = 84%, SSM) of four sucrose-negative strains that included the DNA definition strain for DNA group 9 A. sobria (CDC 0787-80). These four strains, together with five additional strains received in 1989, were subjected to DNA-DNA hybridization (hydroxyapatite, 32P, 60 and 75 degrees C), and all eight strains were closely related to the ninth labeled DNA group 9 definition strain CDC 0787-80 (73 to 86% relatedness at 60 degrees C and 68 to 80% relatedness at 75 degrees C; percent divergence, 2.0 to 3.5). Type strains and DNA definition strains for all other established Aeromonas species were only 35 to 72% related (60 degrees C) to CDC 0787-80. We propose the name Aeromonas jandaei for this highly related group of nine strains, formerly known as DNA group 9 A. sobria. The type strain was designated ATCC 49568 (CDC 0787-80). The nine strains were examined at 36 degrees C and were found to be resistant to 0/129 (vibriostatic agent) and uniformly positive for oxidase, gas production from glucose, indole, lysine decarboxylase, arginine dihydrolase, o-nitrophenyl-beta-D-galactopyranoside, motility (25 degrees C), nitrate reduction, citrate utilization, hemolysis on sheep blood agar, and growth in Trypticase soy broth with no added NaCl. They all fermented D-glucose, D-mannitol, and mannose but did not ferment sucrose, cellobiose, L-arabinose, inositol, salicin, or D-sorbitol. They were uniformly negative for esculin and urea hydrolysis, elastase production, ornithine decarboxylation, and the string test. The antibiogram of A. jandaei resembled that of other aeromonads (resistance to ampicillin and cephalothin), but it differed from most other aeromonads because of resistance to single dilution of colistin and differed from clinical A. veronii biogroup sorbria (formerly A. sobria) by its nearly uniform resistance to cephalothin

  5. Efimov-like phase of a three-stranded DNA and the renormalization-group limit cycle.

    PubMed

    Pal, Tanmoy; Sadhukhan, Poulomi; Bhattacharjee, Somendra M

    2015-04-01

    A three-stranded DNA with short range base pairings only is known to exhibit a classical analog of the quantum Efimov effect, viz., a three-chain bound state at the two-chain melting point where no two are bound. By using a nonperturbative renormalization-group method for a rigid duplex DNA and a flexible third strand, with base pairings and strand exchange, we show that the Efimov-DNA is associated with a limit cycle type behavior of the flow of an effective three-chain interaction. The analysis also shows that thermally generated bubbles play an essential role in producing the effect. A toy model for the flow equations shows the limit cycle in an extended three-dimensional parameter space of the two-chain coupling and a complex three-chain interaction. PMID:25974437

  6. Preferential binding of high mobility group 1 protein to UV-damaged DNA. Role of the COOH-terminal domain.

    PubMed

    Pasheva, E A; Pashev, I G; Favre, A

    1998-09-18

    Binding of chromosomal high mobility group 1 protein (HMG1) to UV-damaged DNA has been studied with oligonucleotides containing a single dipyrimidine site for formation of UV photolesions. Irradiation of an oligonucleotide with unique TT dinucleotide resulted in generation of cyclobutane pyrimidine dimer with no evidence for induction of (6-4) photoproducts, whereas the analysis of irradiated TC-containing oligonucleotide detected (6-4) photoproducts but not cyclobutane pyrimidine dimers. Mobility shift assays have revealed that HMG1 protein binds preferentially to irradiated TT and TC oligonucleotides. Photoreversal of cyclobutane pyrimidine dimers with DNA photolyase and hydrolysis of the (6-4) photoproducts with hot alkali substantially reduced but did not eliminate binding of HMG1. The protein, therefore, appears to bind the two main types of UV damages in DNA, but some other photolesion(s) contributes to the preferential binding of HMG1 to irradiated DNA. By quantifying gel shift assays and considering the efficiencies of lesion formation, we determined dissociation constants of 1.2 +/- 0.5 and 4.0 +/- 1.5 microM for irradiated TT and TC oligonucleotides, respectively, and 70 +/- 20 microM for the control non-irradiated probes. Tryptic removal of the acidic COOH-terminal domain of HMG1 significantly affected binding of the protein to both irradiated and intact oligonucleotides. The potential role of HMG1 in recognition of the UV lesions in DNA is discussed. PMID:9733773

  7. Quaternary ammonium groups exposed at the surface of silica nanoparticles suitable for DNA complexation in the presence of cationic lipids.

    PubMed

    Reinhardt, Nora; Adumeau, Laurent; Lambert, Olivier; Ravaine, Serge; Mornet, Stéphane

    2015-05-28

    The production of silica nanoparticles (NPs) exposing quaternary ammonium groups (NPQ(+)) has been achieved using an optimized chemical surface functionalization protocol. The procedures of surface modification and quaternization of amino groups were validated by diffuse reflectance infrared Fourier transform (DRIFT) and (1)H NMR spectroscopies. Compared to nonquaternized aminated NP, the colloidal stability of NPQ(+) was improved for various pH and salt conditions as assessed by ζ potential and light scattering measurements. In the context of their use for nucleic acid delivery, DNA efficiently bound to NPQ(+) analyzed by cosedimentation assays for a large pH range and various NaCl concentrations and exhibited a better efficacy at basic pH than nonquaternized NP. The study of NPQ(+)/DNA/cationic lipids ternary complexes was carried out with 1,2-dioleoyl-3-trimethylammonium propane (DOTAP) and analyzed by cryo-electron microscopy (cryo-EM). Cryo-EM images showed ternary assemblies where condensed DNA strands are sandwiched between the NPQ(+) surface and the cationic lipid bilayer. Because of an unusual electrostatic colloidal stability of NPQ(+) and a high propensity to bind DNA molecules particularly at high salt concentrations, a novel type of ternary assembly has been formed that might impact the delivery properties of these complexes including their stability in biological environment. PMID:25950202

  8. Immobilization of DNA via oligonucleotides containing an aldehyde or carboxylic acid group at the 5' terminus.

    PubMed Central

    Kremsky, J N; Wooters, J L; Dougherty, J P; Meyers, R E; Collins, M; Brown, E L

    1987-01-01

    A general method for the immobilization of DNA through its 5'-end has been developed. A synthetic oligonucleotide, modified at its 5'-end with an aldehyde or carboxylic acid, was attached to latex microspheres containing hydrazide residues. Using T4 polynucleotide ligase and an oligonucleotide splint, a single stranded 98mer was efficiently joined to the immobilized synthetic fragment. After impregnation of the latex microspheres with the fluorescent dye, Nile Red and attachment of an aldehyde 16mer, 5 X 10(5) bead-DNA conjugates could be detected with a conventional fluorimeter. Images PMID:3562241

  9. Novel Resistance-Nodulation-Cell Division Efflux System AdeDE in Acinetobacter Genomic DNA Group 3

    PubMed Central

    Chau, Sze-Lok; Chu, Yiu-Wai; Houang, Elizabeth T. S.

    2004-01-01

    Resistance-nodulation-cell division type efflux pump AdeDE was identified in acinetobacters belonging to genomic DNA group 3. Inactivation of adeE showed that it may be responsible for reduced susceptibility to amikacin, ceftazidime, chloramphenicol, ciprofloxacin, erythromycin, ethidium bromide, meropenem, rifampin, and tetracycline. However, unlike what was found for other similar efflux systems, the open reading frame for the outer membrane component was not found downstream of the adeDE gene cluster. PMID:15388479

  10. The application of the AMB protective group in the solid-phase synthesis of methylphosphonate DNA analogues.

    PubMed Central

    Kuijpers, W H; Kuyl-Yeheskiely, E; van Boom, J H; van Boeckel, C A

    1993-01-01

    Partially methylphosphonate-modified oligodeoxynucleotides were synthesized on solid-phase by employing the easily removable 2-(acetoxymethyl)benzoyl (AMB) group as base-protecting group. Although a rapid AMB deprotection can be accomplished in methanolic potassium carbonate, the lability of the methylphosphonate linkage towards potassium carbonate/methanol excludes the use of this deprotection reagent. Thus, saturated ammonia solution in methanol was investigated as an alternative reagent for AMB removal. It is demonstrated that the combination of the AMB protective group and ammonia/methanol as deprotection reagent significantly improves the synthesis of methylphosphonate-modified DNA fragments. A mild overnight treatment at room temperature is sufficient for complete removal of the AMB group, whereas deprotection of conventionally protected oligonucleotides requires much longer exposure to basic conditions at elevated temperatures. PMID:8346028

  11. A novel virus genome discovered in an extreme environment suggests recombination between unrelated groups of RNA and DNA viruses

    PubMed Central

    2012-01-01

    Background Viruses are known to be the most abundant organisms on earth, yet little is known about their collective origin and evolutionary history. With exceptionally high rates of genetic mutation and mosaicism, it is not currently possible to resolve deep evolutionary histories of the known major virus groups. Metagenomics offers a potential means of establishing a more comprehensive view of viral evolution as vast amounts of new sequence data becomes available for comparative analysis. Results Bioinformatic analysis of viral metagenomic sequences derived from a hot, acidic lake revealed a circular, putatively single-stranded DNA virus encoding a major capsid protein similar to those found only in single-stranded RNA viruses. The presence and circular configuration of the complete virus genome was confirmed by inverse PCR amplification from native DNA extracted from lake sediment. The virus genome appears to be the result of a RNA-DNA recombination event between two ostensibly unrelated virus groups. Environmental sequence databases were examined for homologous genes arranged in similar configurations and three similar putative virus genomes from marine environments were identified. This result indicates the existence of a widespread but previously undetected group of viruses. Conclusions This unique viral genome carries implications for theories of virus emergence and evolution, as no mechanism for interviral RNA-DNA recombination has yet been identified, and only scant evidence exists that genetic exchange occurs between such distinct virus lineages. Reviewers This article was reviewed by EK, MK (nominated by PF) and AM. For the full reviews, please go to the Reviewers' comments section. PMID:22515485

  12. DNA

    ERIC Educational Resources Information Center

    Stent, Gunther S.

    1970-01-01

    This history for molecular genetics and its explanation of DNA begins with an analysis of the Golden Jubilee essay papers, 1955. The paper ends stating that the higher nervous system is the one major frontier of biological inquiry which still offers some romance of research. (Author/VW)

  13. The Polycomb Group Protein EZH2 Impairs DNA Damage Repair Gene Expression in Human Uterine Fibroids.

    PubMed

    Yang, Qiwei; Nair, Sangeeta; Laknaur, Archana; Ismail, Nahed; Diamond, Michael P; Al-Hendy, Ayman

    2016-03-01

    Uterine fibroids are benign, smooth muscle tumors that occur in approximately 70%-80% of women by age 50 yr. The cellular and molecular mechanism(s) by which uterine fibroids (UFs) develop are not fully understood. Accumulating evidence demonstrates that several genetic abnormalities, including deletions, rearrangements, translocations, as well as mutations, have been found in UFs. These genetic anomalies suggest that low DNA damage repair capacity may be involved in UF formation. The objective of this study was to determine whether expression levels of DNA damage repair-related genes were altered, and how they were regulated in the pathogenesis of UFs. Expression levels of DNA repair-related genes RAD51 and BRCA1 were deregulated in fibroid tissues as compared to adjacent myometrial tissues. Expression levels of chromatin protein enhancer of zeste homolog 2 (EZH2) were higher in a subset of fibroids as compared to adjacent myometrial tissues by both immunohistochemistry and Western blot analysis. Treatment with an inhibitor of EZH2 markedly increased expression levels of RAD51 and BRCA1 in fibroid cells and inhibited cell proliferation paired with cell cycle arrest. Restoring the expression of RAD51 and BRCA1 by treatment with EZH2 inhibitor was dependent on reducing the enrichment of trimethylation of histone 3 lysine 27 epigenetic mark in their promoter regions. This study reveals the important role of EZH2-regulated DNA damage-repair genes via histone methylation in fibroid biology, and may provide novel therapeutic targets for the medical treatment of women with symptomatic UFs. PMID:26888970

  14. Different patterns of evolution for duplicated DNA repair genes in bacteria of the Xanthomonadales group

    PubMed Central

    Martins-Pinheiro, Marinalva; Galhardo, Rodrigo S; Lage, Claudia; Lima-Bessa, Keronninn M; Aires, Karina A; Menck, Carlos FM

    2004-01-01

    Background DNA repair genes encode proteins that protect organisms against genetic damage generated by environmental agents and by-products of cell metabolism. The importance of these genes in life maintenance is supported by their high conservation, and the presence of duplications of such genes may be easily traced, especially in prokaryotic genomes. Results The genome sequences of two Xanthomonas species were used as the basis for phylogenetic analyses of genes related to DNA repair that were found duplicated. Although 16S rRNA phylogenetic analyses confirm their classification at the basis of the gamma proteobacteria subdivision, differences were found in the origin of the various genes investigated. Except for lexA, detected as a recent duplication, most of the genes in more than one copy are represented by two highly divergent orthologs. Basically, one of such duplications is frequently positioned close to other gamma proteobacteria, but the second is often positioned close to unrelated bacteria. These orthologs may have occurred from old duplication events, followed by extensive gene loss, or were originated from lateral gene transfer (LGT), as is the case of the uvrD homolog. Conclusions Duplications of DNA repair related genes may result in redundancy and also improve the organisms' responses to environmental challenges. Most of such duplications, in Xanthomonas, seem to have arisen from old events and possibly enlarge both functional and evolutionary genome potentiality. PMID:15333143

  15. Preparation of carboxyl group-modified palladium nanoparticles in an aqueous solution and their conjugation with DNA

    NASA Astrophysics Data System (ADS)

    Wang, Zhifei; Li, Hongying; Zhen, Shuang; He, Nongyue

    2012-05-01

    The use of nanomaterials in biomolecular labeling and their corresponding detection has been attracting much attention, recently. There are currently very few studies on palladium nanoparticles (Pd NPs) due to their lack of appropriate surface functionalities for conjugation with DNA. In this paper, we thus firstly present an approach to prepare carboxyl group-modified Pd NPs (with an average size of 6 nm) by the use of 11-mercaptoundecanoic acid (MUDA) as a stabilizer in the aqueous solution. The effect of the various reducing reaction conditions on the morphology of the Pd NPs was investigated. The particles were further characterized by TEM, UV-vis, FT-IR and XPS techniques. DNA was finally covalently conjugated to the surface of the Pd NPs through the activation of the carboxyl group, which was confirmed by agarose gel electrophoresis and fluorescence analysis. The resulting Pd NPs-DNA conjugates show high single base pair mismatch discrimination capabilities. This work therefore sets a good foundation for further applications of Pd NPs in bio-analytical research.

  16. Comparison of the genetic diversity of wild and captive groups of Microcebus murinus using the random amplified polymorphic DNA method.

    PubMed

    Neveu, H; Hafen, T; Zimmermann, E; Rumpler, Y

    1998-01-01

    Continued survival of most animal species depends on population management and active protection. It is generally agreed that, in order to avoid extinction of endangered species, ex situ and in situ conservation must be developed in tandem. However, even though many recommendations have been put forward to promote the survival of captive populations, some rapidly become extinct due to loss of genetic diversity (drift effect). Genetic markers, such as random amplified polymorphic DNA (RAPD) markers, can be applied to rapid testing of many individuals. They also permit analysis of very small amounts of DNA, when small species such as mouse lemurs (Microcebus) are to be tested. Using RAPD markers, we compare genetic diversity in four captive groups of Microcebus murinus to that in a sample of 70 wild mouse lemurs. Following the principles of Mendelian inheritance, each amplified fragment of DNA may be considered as a 'locus' (or an amplifying site). The series of bands amplified by a particular primer in any individual is referred to as the individual's 'profile'. We tested 5 primers, or, in the above terms, we studied 98 different 'loci'. Results showed that the captive groups had lost genetic information with respect to the wild sample. Among the four captive groups, the loss of genetic diversity varied according to their number of founders and/or the management of their captive reproduction. Our study of polymorphism permitted us to establish tools for the genetic management of captive breeding, and for the determination of paternity which frequently give better results than behavioural studies; and simulation of introductions or departures of individuals in one very monomorphic group permitted estimation of future increases in its genetic diversity. PMID:9595690

  17. 28s rDNA group-I introns: a powerful tool for identifying strains of Beauveria brongniartii.

    PubMed

    Neuvéglise, C; Brygoo, Y; Riba, G

    1997-04-01

    The nuclear ribosomal DNA of the entomopathogenic fungus Beauveria brongniartii is polymorphic in terms of both restriction site and length. Insertions of 350-450 bp long, identified as group-I introns, were detected in the 28s rDNA. A panel of 47 strains of B. brongniartii, two B. bassiana and one Metarhizium anisopliae of various geographical and biological origins were found to contain 14 variant forms of intron differing in size and restriction pattern, at four different positions. Twelve types of ribosomal large subunit were defined on the basis of variant distribution and compared with strain clustering based on internal transcribed spacers analysis. There was a correlation between the characteristic introns and isolates collected from the sugar cane pest Hoplochelus marginalis. Primers for polymerase chain reaction amplification were chosen from these variants, and used to develop a specific method for detecting strains pathogenic towards Hoplochelus. PMID:9131812

  18. Three Group-I introns in 18S rDNA of Endosymbiotic Algae of Paramecium bursaria from Japan

    NASA Astrophysics Data System (ADS)

    Hoshina, Ryo; Kamako, Shin-ichiro; Imamura, Nobutaka

    2004-08-01

    In the nuclear encoded small subunit ribosomal DNA (18S rDNA) of symbiotic alga of Paramecium bursaria (F36 collected in Japan) possesses three intron-like insertions (Hoshina et al., unpubl. data, 2003). The present study confirmed these exact lengths and insertion sites by reverse transcription-PCR. Two of them were inserted at Escherichia coli 16S rRNA genic position 943 and 1512 that are frequent intron insertion positions, but another insertion position (nearly 1370) was the first finding. Their secondary structures suggested they belong to Group-I intron; one belongs to subgroup IE, others belong to subgroup IC1. Similarity search indicated these introns are ancestral ones.

  19. Fetal blood grouping using cell free DNA - an improved service for RhD negative pregnant women.

    PubMed

    Bills, V L; Soothill, P W

    2014-04-01

    Red cell alloimmunisation involves the transplacental movement of maternally derived red cell antibodies into the fetal circulation, causing red cell haemolysis, fetal anaemia and ultimately fetal death. Current standard UK practice is to prevent sensitisation to the D antigen by administering anti-D at about 28 weeks' gestation to all RhD negative pregnancies. The determination of fetal blood group by non-invasive cell free fetal DNA testing offers an improved and more efficient service to RhD negative pregnant women and avoids the potential iatrogenic harm associated with standard practice. It also has significantly improved the management of women with red cell alloimunisation to D and other antigens. This review summarises the past and future management of red cell alloimmunisation during pregnancy and the impact of ffDNA tests. PMID:24679596

  20. Phylogeny of the Sphaerotilus-Leptothrix group inferred from morphological comparisons, genomic fingerprinting, and 16S ribosomal DNA sequence analyses.

    PubMed

    Siering, P L; Ghiorse, W C

    1996-01-01

    Phase-contrast light microscopy revealed that only one of eight cultivated strains belonging to the Sphaerotilus-Leptothrix group of sheathed bacteria actually produced a sheath in standard growth media. Two Sphaerotilus natans strains produced branched cells, but other morphological characteristics that were used to identify these bacteria were consistent with previously published descriptions. Genomic fingerprints, which were obtained by performing PCR amplification with primers corresponding to enterobacterial repetitive intergenic consensus sequences, were useful for distinguishing between the genera Sphaerotilus and Leptothrix, as well as among individual strains. The complete 16S ribosomal DNA (rDNA) sequences of two strains of "Leptothrix discophora" (strains SP-6 and SS-1) were determined. In addition, partial sequences (approximately 300 nucleotides) of one strain of Leptothrix cholodnii (strain LMG 7171), an unidentified Leptothrix strain (strain NC-1), and four strains of Sphaerotilus natans (strains ATCC 13338T [T = type strain], ATCC 15291, ATCC 29329, and ATCC 29330) were determined. We found that two of the S. natans strains (ATCC 15291 and ATCC 13338T), which differed in morphology and in their genomic fingerprints, had identical sequences in the 300-nucleotide region sequenced. Both parsimony and distance matrix methods were used to infer the evolutionary relationships of the eight strains in a comparison of the 16S rDNA sequences of these organisms with 16S rDNA sequences obtained from ribosomal sequence databases. All of the strains clustered in the Rubrivivax subdivision of the beta subclass of the Proteobacteria, which confirmed previously published conclusions concerning selected individual strains. Additional analyses revealed that all of the S. natans strains clustered in one closely related group, while the Leptothrix strains clustered in two separate lineages that were approximately equidistant from the S. natans cluster. This finding

  1. Complete mtDNA genomes of Filipino ethnolinguistic groups: a melting pot of recent and ancient lineages in the Asia-Pacific region.

    PubMed

    Delfin, Frederick; Min-Shan Ko, Albert; Li, Mingkun; Gunnarsdóttir, Ellen D; Tabbada, Kristina A; Salvador, Jazelyn M; Calacal, Gayvelline C; Sagum, Minerva S; Datar, Francisco A; Padilla, Sabino G; De Ungria, Maria Corazon A; Stoneking, Mark

    2014-02-01

    The Philippines is a strategic point in the Asia-Pacific region for the study of human diversity, history and origins, as it is a cross-road for human migrations and consequently exhibits enormous ethnolinguistic diversity. Following on a previous in-depth study of Y-chromosome variation, here we provide new insights into the maternal genetic history of Filipino ethnolinguistic groups by surveying complete mitochondrial DNA (mtDNA) genomes from a total of 14 groups (11 groups in this study and 3 groups previously published) including previously published mtDNA hypervariable segment (HVS) data from Filipino regional center groups. Comparison of HVS data indicate genetic differences between ethnolinguistic and regional center groups. The complete mtDNA genomes of 14 ethnolinguistic groups reveal genetic aspects consistent with the Y-chromosome, namely: diversity and heterogeneity of groups, no support for a simple dichotomy between Negrito and non-Negrito groups, and different genetic affinities with Asia-Pacific groups that are both ancient and recent. Although some mtDNA haplogroups can be associated with the Austronesian expansion, there are others that associate with South Asia, Near Oceania and Australia that are consistent with a southern migration route for ethnolinguistic group ancestors into the Asia-Pacific, with a timeline that overlaps with the initial colonization of the Asia-Pacific region, the initial colonization of the Philippines and a possible separate post-colonization migration into the Philippine archipelago. PMID:23756438

  2. Complete mtDNA genomes of Filipino ethnolinguistic groups: a melting pot of recent and ancient lineages in the Asia-Pacific region

    PubMed Central

    Delfin, Frederick; Min-Shan Ko, Albert; Li, Mingkun; Gunnarsdóttir, Ellen D; Tabbada, Kristina A; Salvador, Jazelyn M; Calacal, Gayvelline C; Sagum, Minerva S; Datar, Francisco A; Padilla, Sabino G; De Ungria, Maria Corazon A; Stoneking, Mark

    2014-01-01

    The Philippines is a strategic point in the Asia-Pacific region for the study of human diversity, history and origins, as it is a cross-road for human migrations and consequently exhibits enormous ethnolinguistic diversity. Following on a previous in-depth study of Y-chromosome variation, here we provide new insights into the maternal genetic history of Filipino ethnolinguistic groups by surveying complete mitochondrial DNA (mtDNA) genomes from a total of 14 groups (11 groups in this study and 3 groups previously published) including previously published mtDNA hypervariable segment (HVS) data from Filipino regional center groups. Comparison of HVS data indicate genetic differences between ethnolinguistic and regional center groups. The complete mtDNA genomes of 14 ethnolinguistic groups reveal genetic aspects consistent with the Y-chromosome, namely: diversity and heterogeneity of groups, no support for a simple dichotomy between Negrito and non-Negrito groups, and different genetic affinities with Asia-Pacific groups that are both ancient and recent. Although some mtDNA haplogroups can be associated with the Austronesian expansion, there are others that associate with South Asia, Near Oceania and Australia that are consistent with a southern migration route for ethnolinguistic group ancestors into the Asia-Pacific, with a timeline that overlaps with the initial colonization of the Asia-Pacific region, the initial colonization of the Philippines and a possible separate post-colonization migration into the Philippine archipelago. PMID:23756438

  3. Rapid Plant Identification Using Species- and Group-Specific Primers Targeting Chloroplast DNA

    PubMed Central

    Staudacher, Karin; Schallhart, Nikolaus; Mitterrutzner, Evi; Steiner, Eva-Maria; Thalinger, Bettina; Traugott, Michael

    2012-01-01

    Plant identification is challenging when no morphologically assignable parts are available. There is a lack of broadly applicable methods for identifying plants in this situation, for example when roots grow in mixture and for decayed or semi-digested plant material. These difficulties have also impeded the progress made in ecological disciplines such as soil- and trophic ecology. Here, a PCR-based approach is presented which allows identifying a variety of plant taxa commonly occurring in Central European agricultural land. Based on the trnT-F cpDNA region, PCR assays were developed to identify two plant families (Poaceae and Apiaceae), the genera Trifolium and Plantago, and nine plant species: Achillea millefolium, Fagopyrum esculentum, Lolium perenne, Lupinus angustifolius, Phaseolus coccineus, Sinapis alba, Taraxacum officinale, Triticum aestivum, and Zea mays. These assays allowed identification of plants based on size-specific amplicons ranging from 116 bp to 381 bp. Their specificity and sensitivity was consistently high, enabling the detection of small amounts of plant DNA, for example, in decaying plant material and in the intestine or faeces of herbivores. To increase the efficacy of identifying plant species from large number of samples, specific primers were combined in multiplex PCRs, allowing screening for multiple species within a single reaction. The molecular assays outlined here will be applicable manifold, such as for root- and leaf litter identification, botanical trace evidence, and the analysis of herbivory. PMID:22253728

  4. DNA duplexes with reactive dialdehyde groups as novel reagents for cross-linking to restriction- modification enzymes.

    PubMed Central

    Brevnov, M G; Gritsenko, O M; Mikhailov, S N; Efimtseva, E V; Ermolinsky, B S; Van Aerschot, A; Herdewijn, P; Repyk, A V; Gromova, E S

    1997-01-01

    To create new, effective reagents for affinity modification of restriction-modification (R-M) enzymes, a regioselective method for reactive dialdehyde group incorporation into oligonucleotides, based on insertion of a 1-beta-D-galactopyranosylthymine residue, has been developed. We synthesized DNA duplex analogs of the substrates of the Eco RII and Mva I R-M enzymes that contained a galactose or periodate-oxidized galactose residue as single substituents either in the center of the Eco RII (Mva I) recognition site or in the flanking nucleotide sequence. The dependence of binding, cleavage and methylation of these substrate analogs on the modified sugar location in the duplex was determined. Cross-linking of the reagents to the enzymes under different conditions was examined. M. Eco RII covalent attachment to periodate-oxidized substrate analogs proceeded in a specific way and to a large extent depended on the location of the reactive dialdehyde group in the substrate. The yield of covalent attachment to a DNA duplex with a dialdehyde group in the flanking sequence with Eco RII or Mva I methylases was 9-20% and did not exceed 4% for R. Eco RII. PMID:9241245

  5. Identification of group-I introns in the 28s rDNA of the entomopathogenic fungus Beauveria brongniartii.

    PubMed

    Neuvéglise, C; Brygoo, Y

    1994-12-01

    The length of the 28s ribosomal DNA differs significantly between two strains (Bt102 and Bt114) of the entomopathogenic fungus Beauveria brongniartii. RFLP analysis on PCR products revealed the presence of three insertional elements of 350-450 bp in strain Bt114. One of the insertions has been cloned and sequenced and shown to possess all the characteristic sequences and secondary structures of a group-IC intron. Its length is 428 bp and it is devoid of any long open reading frame. The distribution of this intron elsewhere in the genome of Bt114, as well as in the chromosomal ribosomal DNA, was studied. It seems to be present as seven copies in different genes not corresponding to the mitochondrial DNA. The presence of the intron in other strains of B. brongniartii was examined by the hybridization method. Some of them seemed to possess introns with a similar core although others presented no homology with the cloned fragment. PMID:7750145

  6. Function of high-mobility group A proteins in the DNA damage signaling for the induction of apoptosis.

    PubMed

    Fujikane, Ryosuke; Komori, Kayoko; Sekiguchi, Mutsuo; Hidaka, Masumi

    2016-01-01

    O(6)-Methylguanine produced in DNA can pair with thymine during DNA replication, thus leading to a G-to-A transition mutation. To prevent such outcomes, cells harboring O(6)-methylguanine-containing mispair undergo apoptosis that requires the function of mismatch repair (MMR) protein complex. To identify the genes involved in the induction of apoptosis, we performed gene-trap mutagenesis and isolated a clone of mouse cells exhibiting an increased resistance to the killing effect of an alkylating agent, N-methyl-N-nitrosourea (MNU). The mutant carries an insertion in the Hmga2 gene, which belongs to a gene family encoding the high-mobility group A non-histone chromatin proteins. To elucidate the function of HMGA proteins in the apoptosis pathway, we introduced siRNAs for HMGA1 and/or HMGA2 into human HeLa MR cells defective in O(6)-methylguanine-DNA methyltransferase. HMGA1- and HMGA2-single knockdown cells showed an increased resistance to MNU, and HMGA1/HMGA2-double knockdown cells exhibited further increased tolerance compared to the control. The phosphorylation of ATR and CHK1, the appearance of a sub-G1 population, and caspase-9 activation were suppressed in the knockdown cells, although the formation of mismatch recognition complex was unaffected. These results suggest that HMGA family proteins function at the step following the damage recognition in the process of apoptosis triggered by O(6)-methylguanine. PMID:27538817

  7. Function of high-mobility group A proteins in the DNA damage signaling for the induction of apoptosis

    PubMed Central

    Fujikane, Ryosuke; Komori, Kayoko; Sekiguchi, Mutsuo; Hidaka, Masumi

    2016-01-01

    O6-Methylguanine produced in DNA can pair with thymine during DNA replication, thus leading to a G-to-A transition mutation. To prevent such outcomes, cells harboring O6-methylguanine-containing mispair undergo apoptosis that requires the function of mismatch repair (MMR) protein complex. To identify the genes involved in the induction of apoptosis, we performed gene-trap mutagenesis and isolated a clone of mouse cells exhibiting an increased resistance to the killing effect of an alkylating agent, N-methyl-N-nitrosourea (MNU). The mutant carries an insertion in the Hmga2 gene, which belongs to a gene family encoding the high-mobility group A non-histone chromatin proteins. To elucidate the function of HMGA proteins in the apoptosis pathway, we introduced siRNAs for HMGA1 and/or HMGA2 into human HeLa MR cells defective in O6-methylguanine-DNA methyltransferase. HMGA1- and HMGA2-single knockdown cells showed an increased resistance to MNU, and HMGA1/HMGA2-double knockdown cells exhibited further increased tolerance compared to the control. The phosphorylation of ATR and CHK1, the appearance of a sub-G1 population, and caspase-9 activation were suppressed in the knockdown cells, although the formation of mismatch recognition complex was unaffected. These results suggest that HMGA family proteins function at the step following the damage recognition in the process of apoptosis triggered by O6-methylguanine. PMID:27538817

  8. DNA Sequence Variation at the Period Locus Reveals the History of Species and Speciation Events in the Drosophila Virilis Group

    PubMed Central

    Hilton, H.; Hey, J.

    1996-01-01

    The virilis phylad of the Drosophila virilis group consists of five closely related taxa: D. virilis, D. lummei, D. novamexicana, D. americana americana and D. americana texana. DNA sequences from a 2.1-kb pair portion of the period locus were generated in four to eight individuals from each of the five taxa. We found evidence of recombination and high levels of variation within species. We found no evidence of recent natural selection. Surprisingly there was no evidence of divergence between D. a. americana and D. a. texana, and they collectively appear to have had a large historical effective population size. The ranges of these two taxa overlap in a large hybrid zone that has been delineated in the eastern U.S. on the basis of the geographic pattern of a chromosomal fusion. Also surprisingly, D. novamexicana appears to consist of two distinct groups each with low population size and no gene flow between them. PMID:8913746

  9. Testing the validity of Northern European species in the Chrysis ignita species group (Hymenoptera: Chrysididae) with DNA barcoding.

    PubMed

    Soon, Villu; Budrys, Eduardas; Orlovskytė, Svetlana; Paukkunen, Juho; Odegaard, Frode; Ljubomirov, Toshko; Saarma, Urmas

    2014-01-01

    Containing more than a hundred species, the Chrysis ignita species group is the largest and one of the most taxonomically challenging groups in its genus. It has not been possible to resolve the taxonomy of the group using traditional methods due to the lack of robust diagnostic morphological characters. Here we present the results of a molecular analysis designed to delimit species in the Chrysis ignita group for the first time; using mitochondrial sequence data for 364 in-group specimens consisting of all 18 species known to occur in Northern Europe. Two mitochondrial loci were analysed: a COI gene fragment, and a continuous DNA sequence consisting of 16S rRNA, tRNAVal, 12S rRNA and ND4. Two approaches were employed for delimiting species: (1) genetic distance analysis based on the standard COI barcode sequences and; (2) phylogenetic analysis of the COI fragment together with rRNA genes. Both analyses yielded trees with similar topology, but support values for nodes were higher using the second approach. Fifteen species were distinguished in all analyses: Chrysis angustula Schenck, 1856, C. brevitarsis Thomson, 1870, C. clarinicollis Linsenmaier, 1951, C. corusca Valkeila, 1971, C. fulgida Linnaeus, 1761, C. ignita (Linnaeus, 1758), C. impressa Schenck, 1856, C. iris Christ, 1791, C. leptomandibularis Niehuis, 2000, C. longula Abeille de Perrin, 1879, C. ruddii Shuckard, 1837, C. schencki Linsenmaier, 1968, C. subcoriacea Linsenmaier, 1959, C. terminata Dahlbom, 1854 and C. vanlithi Linsenmaier, 1959. The specific status of C. mediata Linsenmaier, 1951 and C. solida Haupt, 1957 was not resolved. Included unidentified specimens grouped in three clusters, two of which are distinctly delimited and apparently represent cryptic species. The specific status of the unidentified samples in the third cluster remained unclear. Moreover, our data suggest the existence of additional cryptic species currently lumped under the names C. pseudobrevitarsis Linsenmaier

  10. A DNA sequence-based study of the Schistosoma indicum (Trematoda: Digenea) group: population phylogeny, taxonomy and historical biogeography.

    PubMed

    Attwood, S W; Fatih, F A; Mondal, M M H; Alim, M A; Fadjar, S; Rajapakse, R P V J; Rollinson, D

    2007-12-01

    Partial (DNA) sequences were collected for 2 mitochondrial loci (Srrna and Lrrna, the rrnS and rrnL rRNA genes respectively) for Schistosoma indicum group species from 4 Southeast Asian countries. The samples included 7 populations, 4 of which were previously unstudied. In 11 cases the combination of locus and population was new. The aim of the study was to provide a phylogeny based on new independent data and multiple populations (earlier studies had mostly used a common set of field samples or laboratory lines) and to examine interrelationships and phylogeography within this species group. Paraphyly of the S. indicum group was confirmed, as was the basal position of Schistosoma incognitum in the Schistosoma phylogeny. Southeast Asian Schistosoma spindale and S. incognitum populations were shown to fall into their respective con-specific cohesive groupings. Estimated divergence times for these taxa were shown to be related to Pleistocene changes in sea level and the radiation of definitive host groups. A revised phylogeographical model is proposed in the light of these findings. PMID:17822572

  11. Genetic polymorphism of Malassezia furfur isolates from Han and Tibetan ethnic groups in China using DNA fingerprinting.

    PubMed

    Zhang, Hao; Zhang, Ruifeng; Ran, Yuping; Dai, Yaling; Lu, Yao; Wang, Peng

    2010-12-01

    Reported isolation rates of Malassezia yeast from human skin show geographic variations. In China, the populations of the Han (1,182.95 million) and Tibetan (5.41 million) ethnic groups are distributed over 9.6 and 3.27 million square kilometers respectively, making biodiversity research feasible and convenient. Malassezia furfur clinical strains (n = 29) isolated from different individuals, with or without associated dermatoses, of these two ethnic groups (15 Han and 12 Tibetan) were identified and analyzed with DNA fingerprinting using single primers specific to minisatellites. Using the Bionumerics software, we found that almost all M. furfur clinical isolates and type strains formed five distinct group clusters according to their associated skin diseases and the ethnic groups of the patients. These findings are the first to focus on the genetic diversity and relatedness of M. furfur in the Tibetan and Han ethnic groups in China and reveal genetic variation associated with related diseases, host ethnicity and geographic origin. PMID:20507265

  12. Species discrimination in the subfamily Ostertagiinae of Northern China: assessment of DNA barcode in a taxonomically challenging group.

    PubMed

    Lv, Jizhou; Zhang, Yongning; Feng, Chunyan; Yuan, Xiangfen; Sun, Degang; Deng, Junhua; Wang, Caixia; Wu, Shaoqiang; Lin, Xiangmei

    2016-03-01

    Gastrointestinal nematodes within the subfamily Ostertagiinae (Teladorsagia, Ostertagia, and Marshallagia et al.) are among the most common infections of domesticated livestock. These parasites are of particular interest, as many of the species within this group are of economic importance worldwide. Traditionally, nematode species designations have been based on morphological criteria. However, this group possesses poorly defined species. There is an urgent need to develop a reliable technique that can distinguish species of Ostertagiinae. DNA barcoding has been proved to be a powerful tool to identify species of birds, mammals, and arthropods, but this technique has not yet been examined for identifying species of Ostertagiinae. In this study, a total of 138 mitochondrial DNA (mtDNA) cytochrome c oxidase subunit I (COI) sequences from individuals representing 11 species of Ostertagiinae were acquired by PCR for the first time. The specimens were collected from pastoral area of northern China. Genetic divergence analyses showed that mean interspecific Kimura two-parameter distances of COI (13.61 %) were about four times higher than the mean value of the intraspecific divergence (3.69 %). Then, the performance of the COI to identify species of Ostertagiinae was evaluated by identification success rates using nearest neighbor (NN) and BLASTn. The results indicated that the rates of correct sequence identification for COI were high (>80 %) when using the NN and BLASTn methods. Besides, the deep lineage divergences are detected in Teladorsagia circumcincta. Meanwhile, the analyses also detected no genetic differentiation between some species such as Ostertagia hahurica and Ostertagia buriatica. These results indicate that the traditional status of species within Ostertagiinae should be closely examined based on the molecular data. PMID:26584827

  13. Two group I ribozymes with different functions in a nuclear rDNA intron.

    PubMed Central

    Decatur, W A; Einvik, C; Johansen, S; Vogt, V M

    1995-01-01

    DiSSU1, a mobile intron in the nuclear rRNA gene of Didymium iridis, was previously reported to contain two independent catalytic RNA elements. We have found that both catalytic elements, renamed GIR1 and GIR2, are group I ribozymes, but with differing functionality. GIR2 carries out the several reactions associated with self-splicing. GIR1 carries out a hydrolysis reaction at an internal processing site (IPS-1). These conclusions are based on the catalytic properties of RNAs transcribed in vitro. Mutation of the P7 pairing segment of GIR2 abrogated self-splicing, while mutation of P7 in GIR1 abrogated hydrolysis at the IPS-1. Much of the P2 stem and all of the associated loop could be deleted without effect on self-splicing. These results are accounted for by a secondary structure model, in which a long P2 pairing segment brings the 5' splice site to the GIR2 catalytic core. GIR1 is the smallest natural group I ribozyme yet reported and is the first example of a group I ribozyme whose presumptive biological function is hydrolysis. We hypothesize that GIR1-mediated cleavage of the excised intron RNA functions in the generation and expression of the mRNA for the intron-encoded endonuclease I-DirI. Images PMID:7556099

  14. Functions of the high mobility group protein, Abf2p, in mitochondrial DNA segregation, recombination and copy number in Saccharomyces cerevisiae.

    PubMed Central

    Zelenaya-Troitskaya, O; Newman, S M; Okamoto, K; Perlman, P S; Butow, R A

    1998-01-01

    Previous studies have established that the mitochondrial high mobility group (HMG) protein, Abf2p, of Saccharomyces cerevisiae influences the stability of wild-type (rho+) mitochondrial DNA (mtDNA) and plays an important role in mtDNA organization. Here we report new functions for Abf2p in mtDNA transactions. We find that in homozygous deltaabf2 crosses, the pattern of sorting of mtDNA and mitochondrial matrix protein is altered, and mtDNA recombination is suppressed relative to homozygous ABF2 crosses. Although Abf2p is known to be required for the maintenance of mtDNA in rho+ cells growing on rich dextrose medium, we find that it is not required for the maintenance of mtDNA in p cells grown on the same medium. The content of both rho+ and rho- mtDNAs is increased in cells by 50-150% by moderate (two- to threefold) increases in the ABF2 copy number, suggesting that Abf2p plays a role in mtDNA copy control. Overproduction of Abf2p by > or = 10-fold from an ABF2 gene placed under control of the GAL1 promoter, however, leads to a rapid loss of rho+ mtDNA and a quantitative conversion of rho+ cells to petites within two to four generations after a shift of the culture from glucose to galactose medium. Overexpression of Abf2p in rho- cells also leads to a loss of mtDNA, but at a slower rate than was observed for rho+ cells. The mtDNA instability phenotype is related to the DNA-binding properties of Abf2p because a mutant Abf2p that contains mutations in residues of both HMG box domains known to affect DNA binding in vitro, and that binds poorly to mtDNA in vivo, complements deltaabf2 cells only weakly and greatly lessens the effect of overproduction on mtDNA instability. In vivo binding was assessed by colocalization to mtDNA of fusions between mutant or wild-type Abf2p and green fluorescent protein.These findings are discussed in the context of a model relating mtDNA copy number control and stability to mtDNA recombination. PMID:9581629

  15. ISC, a Novel Group of Bacterial and Archaeal DNA Transposons That Encode Cas9 Homologs

    PubMed Central

    Kapitonov, Vladimir V.; Makarova, Kira S.

    2015-01-01

    ABSTRACT Bacterial genomes encode numerous homologs of Cas9, the effector protein of the type II CRISPR-Cas systems. The homology region includes the arginine-rich helix and the HNH nuclease domain that is inserted into the RuvC-like nuclease domain. These genes, however, are not linked to cas genes or CRISPR. Here, we show that Cas9 homologs represent a distinct group of nonautonomous transposons, which we denote ISC (insertion sequences Cas9-like). We identify many diverse families of full-length ISC transposons and demonstrate that their terminal sequences (particularly 3′ termini) are similar to those of IS605 superfamily transposons that are mobilized by the Y1 tyrosine transposase encoded by the TnpA gene and often also encode the TnpB protein containing the RuvC-like endonuclease domain. The terminal regions of the ISC and IS605 transposons contain palindromic structures that are likely recognized by the Y1 transposase. The transposons from these two groups are inserted either exactly in the middle or upstream of specific 4-bp target sites, without target site duplication. We also identify autonomous ISC transposons that encode TnpA-like Y1 transposases. Thus, the nonautonomous ISC transposons could be mobilized in trans either by Y1 transposases of other, autonomous ISC transposons or by Y1 transposases of the more abundant IS605 transposons. These findings imply an evolutionary scenario in which the ISC transposons evolved from IS605 family transposons, possibly via insertion of a mobile group II intron encoding the HNH domain, and Cas9 subsequently evolved via immobilization of an ISC transposon. IMPORTANCE Cas9 endonucleases, the effectors of type II CRISPR-Cas systems, represent the new generation of genome-engineering tools. Here, we describe in detail a novel family of transposable elements that encode the likely ancestors of Cas9 and outline the evolutionary scenario connecting different varieties of these transposons and Cas9. PMID:26712934

  16. Japanese Wolves are Genetically Divided into Two Groups Based on an 8-Nucleotide Insertion/Deletion within the mtDNA Control Region.

    PubMed

    Ishiguro, Naotaka; Inoshima, Yasuo; Yanai, Tokuma; Sasaki, Motoki; Matsui, Akira; Kikuchi, Hiroki; Maruyama, Masashi; Hongo, Hitomi; Vostretsov, Yuri E; Gasilin, Viatcheslav; Kosintsev, Pavel A; Quanjia, Chen; Chunxue, Wang

    2016-02-01

    The mitochondrial DNA (mtDNA) control region (198- to 598-bp) of four ancient Canis specimens (two Canis mandibles, a cranium, and a first phalanx) was examined, and each specimen was genetically identified as Japanese wolf. Two unique nucleotide substitutions, the 78-C insertion and the 482-G deletion, both of which are specific for Japanese wolf, were observed in each sample. Based on the mtDNA sequences analyzed, these four specimens and 10 additional Japanese wolf samples could be classified into two groups- Group A (10 samples) and Group B (4 samples)-which contain or lack an 8-bp insertion/deletion (indel), respectively. Interestingly, three dogs (Akita-b, Kishu 25, and S-husky 102) that each contained Japanese wolf-specific features were also classified into Group A or B based on the 8-bp indel. To determine the origin or ancestor of the Japanese wolf, mtDNA control regions of ancient continental Canis specimens were examined; 84 specimens were from Russia, and 29 were from China. However, none of these 113 specimens contained Japanese wolf-specific sequences. Moreover, none of 426 Japanese modern hunting dogs examined contained these Japanese wolf-specific mtDNA sequences. The mtDNA control region sequences of Groups A and B appeared to be unique to grey wolf and dog populations. PMID:26853868

  17. Novel Function of the Fanconi Anemia Group J or RECQ1 Helicase to Disrupt Protein-DNA Complexes in a Replication Protein A-stimulated Manner*

    PubMed Central

    Sommers, Joshua A.; Banerjee, Taraswi; Hinds, Twila; Wan, Bingbing; Wold, Marc S.; Lei, Ming; Brosh, Robert M.

    2014-01-01

    Understanding how cellular machinery deals with chromosomal genome complexity is an important question because protein bound to DNA may affect various cellular processes of nucleic acid metabolism. DNA helicases are at the forefront of such processes, yet there is only limited knowledge how they remodel protein-DNA complexes and how these mechanisms are regulated. We have determined that representative human RecQ and Fe-S cluster DNA helicases are potently blocked by a protein-DNA interaction. The Fanconi anemia group J (FANCJ) helicase partners with the single-stranded DNA-binding protein replication protein A (RPA) to displace BamHI-E111A bound to duplex DNA in a specific manner. Protein displacement was dependent on the ATPase-driven function of the helicase and unique properties of RPA. Further biochemical studies demonstrated that the shelterin proteins TRF1 and TRF2, which preferentially bind the telomeric repeat found at chromosome ends, effectively block FANCJ from unwinding the forked duplex telomeric substrate. RPA, but not the Escherichia coli single-stranded DNA-binding protein or shelterin factor Pot1, stimulated FANCJ ejection of TRF1 from the telomeric DNA substrate. FANCJ was also able to displace TRF2 from the telomeric substrate in an RPA-dependent manner. The stimulation of helicase-catalyzed protein displacement is also observed with the DNA helicase RECQ1, suggesting a conserved functional interaction of RPA-interacting helicases. These findings suggest that partnerships between RPA and interacting human DNA helicases may greatly enhance their ability to dislodge proteins bound to duplex DNA, an activity that is likely to be highly relevant to their biological roles in DNA metabolism. PMID:24895130

  18. DNA polymerase gene sequences indicate western and forest tent caterpillar viruses form a new taxonomic group within baculoviruses.

    PubMed

    Nielsen, Cydney B; Cooper, Dawn; Short, Steven M; Myers, Judith H; Suttle, Curtis A

    2002-11-01

    Baculoviruses infect larval lepidopterans, and thus have potential value as microbial controls of agricultural and forest pests. Understanding their genetic relatedness and host specificity is relevant to the risk assessment of viral insecticides if non-target impacts are to be avoided. DNA polymerase gene sequences have been demonstrated to be useful for inferring genetic relatedness among dsDNA viruses. We have adopted this approach to examine the relatedness among natural isolates of two uncharacterized caterpillar-infecting baculoviruses, Malacosoma californicum pluviale nucleopolyhedrovirus (McplMNPV) and Malacosoma disstria nucleopolyhedrovirus (MadiMNPV), which infect two closely related host species with little to no cross-infectivity. We designed two degenerate primers (BVP1 and BVP2) based on protein motifs conserved among baculoviruses. McplMNPV and MadiMNPV viral DNA was obtained from naturally infected caterpillars collected from geographically distinct sites in the Southern Gulf Islands and Prince George regions of British Columbia, Canada. Sequencing of 0.9 kb PCR amplicons from six McplMNPV and six MadiMNPV isolates obtained from a total of eight sites, revealed very low nucleotide variation among McplMNPV isolates (99.2-100% nucleotide identity) and among MadiMNPV isolates (98.9-100% nucleotide identity). Greater nucleotide variation was observed between viral isolates from the two different caterpillar species (only 84.7-86.1% nucleotide identity). Both maximum parsimony and maximum likelihood phylogenetic analyses support placement of McplMNPV and MadiMNPV in a clade that is distinct from other groups of baculoviruses. PMID:12507483

  19. The effect of leaving groups on binding and reactivity in enzyme-free copying of DNA and RNA.

    PubMed

    Kervio, Eric; Sosson, Marilyne; Richert, Clemens

    2016-07-01

    The template-directed incorporation of nucleotides at the terminus of a growing primer is the basis of the transmission of genetic information. Nature uses polymerases-catalyzed reactions, but enzyme-free versions exist that employ nucleotides with organic leaving groups. The leaving group affects yields, but it was not clear whether inefficient extensions are due to poor binding, low reactivity toward the primer, or rapid hydrolysis. We have measured the binding of a total of 15 different activated nucleotides to DNA or RNA sequences. Further, we determined rate constants for the chemical step of primer extension involving methylimidazolides or oxyazabenzotriazolides of deoxynucleotides or ribonucleotides. Binding constants range from 10 to >500 mM and rate constants from 0.1 to 370 M(-1) h(-1) For aminoterminal primers, a fast covalent step and slow hydrolysis are the main factors leading to high yields. For monomers with weakly pairing bases, the leaving group can improve binding significantly. A detailed mechanistic picture emerges that explains why some enzyme-free primer extensions occur in high yield, while others remain recalcitrant to copying without enzymatic catalysis. A combination of tight binding and rapid extension, coupled with slow hydrolysis induces efficient enzyme-free copying. PMID:27235418

  20. Survey of chimeric IStron elements in bacterial genomes: multiple molecular symbioses between group I intron ribozymes and DNA transposons

    PubMed Central

    Tourasse, Nicolas J.; Stabell, Fredrik B.; Kolstø, Anne-Brit

    2014-01-01

    IStrons are chimeric genetic elements composed of a group I intron associated with an insertion sequence (IS). The group I intron is a catalytic RNA providing the IStron with self-splicing ability, which renders IStron insertions harmless to the host genome. The IS element is a DNA transposon conferring mobility, and thus allowing the IStron to spread in genomes. IStrons are therefore a striking example of a molecular symbiosis between unrelated genetic elements endowed with different functions. In this study, we have conducted the first comprehensive survey of IStrons in sequenced genomes that provides insights into the distribution, diversity, origin and evolution of IStrons. We show that IStrons have a restricted phylogenetic distribution limited to two bacterial phyla, the Firmicutes and the Fusobacteria. Nevertheless, diverse IStrons representing two major groups targeting different insertion site motifs were identified. This taken with the finding that while the intron components of all IStrons belong to the same structural class, they are fused to different IS families, indicates that multiple intron–IS symbioses have occurred during evolution. In addition, introns and IS elements related to those that were at the origin of IStrons were also identified. PMID:25324310

  1. The effect of leaving groups on binding and reactivity in enzyme-free copying of DNA and RNA

    PubMed Central

    Kervio, Eric; Sosson, Marilyne; Richert, Clemens

    2016-01-01

    The template-directed incorporation of nucleotides at the terminus of a growing primer is the basis of the transmission of genetic information. Nature uses polymerases-catalyzed reactions, but enzyme-free versions exist that employ nucleotides with organic leaving groups. The leaving group affects yields, but it was not clear whether inefficient extensions are due to poor binding, low reactivity toward the primer, or rapid hydrolysis. We have measured the binding of a total of 15 different activated nucleotides to DNA or RNA sequences. Further, we determined rate constants for the chemical step of primer extension involving methylimidazolides or oxyazabenzotriazolides of deoxynucleotides or ribonucleotides. Binding constants range from 10 to >500 mM and rate constants from 0.1 to 370 M−1 h−1. For aminoterminal primers, a fast covalent step and slow hydrolysis are the main factors leading to high yields. For monomers with weakly pairing bases, the leaving group can improve binding significantly. A detailed mechanistic picture emerges that explains why some enzyme-free primer extensions occur in high yield, while others remain recalcitrant to copying without enzymatic catalysis. A combination of tight binding and rapid extension, coupled with slow hydrolysis induces efficient enzyme-free copying. PMID:27235418

  2. Detection of bovine group a rotavirus using rapid antigen detection kits, rt-PCR and next-generation DNA sequencing.

    PubMed

    Minami-Fukuda, Fujiko; Nagai, Makoto; Takai, Hikaru; Murakami, Toshiaki; Ozawa, Tadashi; Tsuchiaka, Shinobu; Okazaki, Sachiko; Katayama, Yukie; Oba, Mami; Nishiura, Naomi; Sassa, Yukiko; Omatsu, Tsutomu; Furuya, Tetsuya; Koyama, Satoshi; Shirai, Junsuke; Tsunemitsu, Hiroshi; Fujii, Yoshiki; Katayama, Kazuhiko; Mizutani, Tetsuya

    2013-12-30

    We investigated the sensitivity of human rotavirus rapid antigen detection (RAD) kits, RT-PCR and next-generation DNA sequencing (NGS) for detection of bovine group A rotavirus (RVA). The Dipstick 'Eiken' Rota (Dipstick) showed the highest sensitivity out of the seven RAD kits against all selected strains in limited dilution analyses, which was consistent with the results for equine rotavirus previously reported. RT-PCR had 10⁰-10³-fold higher sensitivity than the Dipstick. NGS using thirteen RT-PCR-negative fecal samples revealed that all samples yielded RVA reads and especially that two of them covered all 11 genome segments. Moreover, mapping reads to reference sequences allowed genotyping. The NGS would be sensitive and useful for analysis of less dependent on specific primers and screening of genotypes. PMID:23912876

  3. Mitochondrial DNA diversity in two ethnic groups in southeastern Kenya: perspectives from the northeastern periphery of the Bantu expansion

    PubMed Central

    Batai, Ken; Babrowski, Kara B.; Arroyo, Juan Pablo; Kusimba, Chapurukha M.; Williams, Sloan R.

    2013-01-01

    The Bantu languages are widely distributed throughout sub-Saharan Africa. Genetic research supports linguists and historians who argue that migration played an important role in the spread of this language family, but the genetic data also indicates a more complex process involving substantial gene flow with resident populations. In order to understand the Bantu expansion process in east Africa, mtDNA hypervariable region I variation in 352 individuals from the Taita and Mijikenda ethnic groups was analyzed, and we evaluated the interactions that took place between the Bantu- and non-Bantu-speaking populations in east Africa. The Taita and Mijikenda are Bantu-speaking agropastoralists from southeastern Kenya, at least some of whose ancestors probably migrated into the area as part of Bantu migrations that began around 3,000 BCE. Our analyses indicate that they show some distinctive differences that reflect their unique cultural histories. The Taita are genetically more diverse than the Mijikenda with larger estimates of genetic diversity. The Taita cluster with other east African groups, having high frequencies of haplogroups from that region, while the Mijikenda have high frequencies of central African haplogroups and cluster more closely with central African Bantu-speaking groups. The non-Bantu speakers who lived in southeastern Kenya before Bantu speaking groups arrived were at least partially incorporated into what are now Bantu-speaking Taita groups. In contrast, gene flow from non-Bantu speakers into the Mijikenda was more limited. These results suggest a more complex demographic history where the nature of Bantu and non-Bantu interactions varied throughout the area. PMID:23382080

  4. Analysis of a human DNA excision repair gene involved in group A xeroderma pigmentosum and containing a zinc-finger domain.

    PubMed

    Tanaka, K; Miura, N; Satokata, I; Miyamoto, I; Yoshida, M C; Satoh, Y; Kondo, S; Yasui, A; Okayama, H; Okada, Y

    1990-11-01

    Xeroderma pigmentosum (XP) is an autosomal recessive disease, characterized by a high incidence of sunlight-induced skin cancer. Cells from people with this condition are hypersensitive to ultraviolet because of a defect in DNA repair. There are nine genetic complementation groups of XP, groups A-H and a variant. We have cloned the mouse DNA repair gene that complements the defect of group A, the XPAC gene. Here we report molecular cloning of human and mouse XPAC complementary DNAs. Expression of XPAC cDNA confers ultraviolet-resistance on several group A cell lines, but not on lines of other XP groups. Almost all group A lines tested showed abnormality or absence of XPAC messenger RNAs. These results indicate that a defective XPAC gene causes group A XP. The human and mouse XPAC genes are located on chromosome 9q34.1 and chromosome 4C2, respectively. Human XPAC cDNA encodes a protein of 273 amino acids with a zinc-finger motif. PMID:2234061

  5. Phylogeny and chromosomal variations in East Asian Carex, Siderostictae group (Cyperaceae), based on DNA sequences and cytological data.

    PubMed

    Yano, Okihito; Ikeda, Hiroshi; Jin, Xiao-Feng; Hoshino, Takuji

    2014-01-01

    Carex (Cyperaceae) is one of the largest genera of the flowering plants, and comprises more than 2,000 species. In Carex, section Siderostictae with broader leaves distributed in East Asia is thought to be an ancestral group. We aimed to clarify the phylogenetic relationships and chromosomal variations within the section Siderostictae, and to examine the relationship of broad-leaved species of the sections Hemiscaposae and Surculosae from East Asia, inferred from DNA sequences and cytological data. Our results indicate that a monophyletic Siderostictae clade, including the sections Hemiscaposae, Siderostictae and Surculosae, as the earliest diverging group in the tribe Cariceae. Low chromosome numbers, 2n = 12 or 24, with large sizes were observed in these three sections. Our results suggest that the genus Carex might have originated or relictly restricted in the East Asia. Geographical distributions of diploid species are restricted in narrower areas, while those of tetraploid species are wider in East Asia. It is concluded that chromosomal variations in Siderostictae clade may have been caused by polyploidization and that tetraploid species may have been able to exploit their habitats by polyploidization. PMID:23857080

  6. A mobile group I intron from Physarum polycephalum can insert itself and induce point mutations in the nuclear ribosomal DNA of saccharomyces cerevisiae.

    PubMed Central

    Muscarella, D E; Vogt, V M

    1993-01-01

    Pp LSU3 is a mobile group I intron in the extrachromosomal nuclear ribosomal DNA (rDNA) of Physarum polycephalum. As found for other mobile introns, Pp LSU3 encodes a site-specific endonuclease, I-Ppo, which mediates "homing" to unoccupied target sites in Physarum rDNA. The recognition sequence for this enzyme is conserved in all eucaryotic nuclear rDNAs. We have introduced this intron into a heterologous species, Saccharomyces cerevisiae, in which nuclear group I introns have not been detected. The expression of Pp LSU3, under control of the inducible GAL10 promoter, was found to be lethal as a consequence of double-strand breaks in the rDNA. However, surviving colonies that are resistant to the lethal effects of I-Ppo because of alterations in the rDNA at the cleavage site were recovered readily. These survivors are of two classes. The first comprises cells that acquired one of three types of point mutations. The second comprises cells in which Pp LSU3 became inserted into the rDNA. In both cases, each resistant survivor appears to carry the same alterations in all approximately 150 rDNA repeats. When it is embedded in yeast rDNA, Pp LSU3 leads to the synthesis of I-Ppo and appears to be mobile in appropriate genetic crosses. The existence of yeast cells carrying a mobile intron should allow dissection of the steps that allow expression of the highly unusual I-Ppo gene. Images PMID:8380887

  7. New pyridinium-based fluorescent dyes: A comparison of symmetry and side-group effects on G-Quadruplex DNA binding selectivity and application in live cell imaging.

    PubMed

    Lu, Yu-Jing; Hu, Dong-Ping; Zhang, Kun; Wong, Wing-Leung; Chow, Cheuk-Fai

    2016-07-15

    A series of C1-, C2-and C3-symmetric pyridinium conjugates with different styrene-like side groups were synthesized and were utilized as G-quadruplex selective fluorescent probes. The new compounds were well-characterized. Their selectivity, sensitivity, and stability towards G-quadruplex were studied by fluorescence titration, native PAGE experiments, FRET and circular dichroism (CD) analyses. These new compounds investigated in the fluorescence assays were preferentially bound with G-quadruplex DNA compared with other type of nucleic acids and it is fascinating to realize the effects of molecular symmetry and associated side groups showing unexpectedly great influence on the fluorescent signal enhancement for the discrimination of G-quadruplexes DNA from other nucleic acids. This may correlate with the pocket symmetry and shape of the G-quadruplex DNA inherently. Among the compounds, a C2-symmetric dye (2,6-bis-((E)-2-(1H-indol-3-yl)-vinyl)-1-methylpyridin-1-ium iodide) with indolyl-groups substituted was screened out from the series giving the best selectivity and sensitivity towards G-quadruplexes DNA, particularly telo21, due to its high equilibrium binding constant (K=2.17×10(5)M(-1)). In addition, the limit of detection (LOD) of the dye to determine telo21 DNA in bioassays was found as low as 33nM. The results of the study give insight and certain crucial factors, such as molecular symmetry and the associated side groups, on developing of effective fluorescent dyes for G-quadruplex DNA applications including G-quadruplex structure stabilization, biosensing and clinical applications. The compound was also demonstrated as a very selective G-quadruplex fluorescent agent for living cell staining and imaging. PMID:26994364

  8. Spectroscopic study on the interaction of ct-DNA with manganese Salen complex containing triphenyl phosphonium groups

    NASA Astrophysics Data System (ADS)

    Dehkordi, Maryam Nejat; Bordbar, Abdol-Khalegh; Lincoln, Per; Mirkhani, Valiollah

    2012-05-01

    The DNA binding properties of a bulky and hydrophobic Schiff base complex of manganese(III) [N,N'-bis(5-(triphenyl phosphonium methyl)salicylidene)-1,2-ethylene diamine chloride Mn(III) acetate] was examined by spectroscopic techniques. UV-vis titration data indicate both hypo and hyperchromic effect with addition of DNA to complex. A competitive binding study showed that the enhanced emission intensity of ethidium bromide (EB) in the presence of DNA was quenched by adding Mn Salen complex. This finding indicates that Mn Salen complex displaces EB from its binding site in DNA. Helix melting studies indicate improvement in the helix stability, and an increase in the melting temperature. The analysis of CD spectra represents the structural changes in DNA due to the binding of Mn Salen complex. The binding constant has been calculated using absorbance and fluorescence data. The results also represent that the binding process proceeds by strong electrostatic and hydrophobic interactions.

  9. Ultraviolet-B-induced DNA damage and ultraviolet-B tolerance mechanisms in species with different functional groups coexisting in subalpine moorlands.

    PubMed

    Wang, Qing-Wei; Kamiyama, Chiho; Hidema, Jun; Hikosaka, Kouki

    2016-08-01

    High doses of ultraviolet-B (UV-B; 280-315 nm) radiation can have detrimental effects on plants, and especially damage their DNA. Plants have DNA repair and protection mechanisms to prevent UV-B damage. However, it remains unclear how DNA damage and tolerance mechanisms vary among field species. We studied DNA damage and tolerance mechanisms in 26 species with different functional groups coexisting in two moorlands at two elevations. We collected current-year leaves in July and August, and determined accumulation of cyclobutane pyrimidine dimer (CPD) as UV-B damage and photorepair activity (PRA) and concentrations of UV-absorbing compounds (UACs) and carotenoids (CARs) as UV-B tolerance mechanisms. DNA damage was greater in dicot than in monocot species, and higher in herbaceous than in woody species. Evergreen species accumulated more CPDs than deciduous species. PRA was higher in Poaceae than in species of other families. UACs were significantly higher in woody than in herbaceous species. The CPD level was not explained by the mechanisms across species, but was significantly related to PRA and UACs when we ignored species with low CPD, PRA and UACs, implying the presence of another effective tolerance mechanism. UACs were correlated negatively with PRA and positively with CARs. Our results revealed that UV-induced DNA damage significantly varies among native species, and this variation is related to functional groups. DNA repair, rather than UV-B protection, dominates in UV-B tolerance in the field. Our findings also suggest that UV-B tolerance mechanisms vary among species under evolutionary trade-off and synergism. PMID:27139425

  10. DNA-damaging activity in ethanol-soluble fractions of feces from New Zealand groups at varying risks of colorectal cancer.

    PubMed

    Ferguson, L R; Alley, P G; Gribben, B M

    1985-01-01

    Using repair-proficient and repair-deficient strains of E. coli, we investigated the application of a liquid incubation assay to measure the DNA-damaging activity of ethanol-soluble fecal extracts. This method appears to be suitable for the study of a wide range of sample types. It was used to measure the DNA-modifying activity of ethanol-soluble fecal extracts from a group of European colorectal cancer patients. Data were compared with those from Europeans of similar age and sex distribution who did not have bowel cancer. We also studied groups of Maoris, Samoans, and European Seventh-Day Adventists who followed an ovo-lacto vegetarian diet. There are significant levels of DNA-modifying materials in the feces of many Europeans on a mixed diet, regardless of whether or not they have cancer. The number of positive samples was less in the Polynesian groups, and there were no samples that could be unequivocally scored as positive in the Seventh-Day Adventist groups. We conclude that diet can significantly reduce the level of ethanol-soluble mutagens, at least in New Zealand Europeans. The data may provide an explanation for the reduced incidence of bowel cancer in Seventh-Day Adventist groups. PMID:3906579

  11. A comparison of different pre-lysis methods and extraction kits for recovery of Streptococcus agalacticae (Lancefield group B Streptococcus) DNA from whole blood.

    PubMed

    Burke, Rachael M; McKenna, James P; Cox, Ciara; Coyle, Peter V; Shields, Michael D; Fairley, Derek J

    2016-10-01

    Sub-optimal recovery of bacterial DNA from whole blood samples can limit the sensitivity of molecular assays to detect pathogenic bacteria. We compared 3 different pre-lysis protocols (none, mechanical pre-lysis and achromopeptidase pre-lysis) and 5 commercially available DNA extraction platforms for direct detection of Group B Streptococcus (GBS) in spiked whole blood samples, without enrichment culture. DNA was extracted using the QIAamp Blood Mini kit (Qiagen), UCP Pathogen Mini kit (Qiagen), QuickGene DNA Whole Blood kit S (Fuji), Speed Xtract Nucleic Acid Kit 200 (Qiagen) and MagNA Pure Compact Nucleic Acid Isolation Kit I (Roche Diagnostics Corp). Mechanical pre-lysis increased yields of bacterial genomic DNA by 51.3 fold (95% confidence interval; 31.6-85.1, p<0.001) and pre-lysis with achromopeptidase by 6.1 fold (95% CI; 4.2-8.9, p<0.001), compared with no pre-lysis. Differences in yield due to pre-lysis were 2-3 fold larger than differences in yield between extraction methods. Including a pre-lysis step can improve the limits of detection of GBS using PCR or other molecular methods without need for culture. PMID:27546716

  12. A distinct group of CpG islands shows differential DNA methylation between replicas of the same cell line in vitro

    PubMed Central

    2013-01-01

    Background CpG dinucleotide-rich genomic DNA regions, known as CpG islands (CGIs), can be methylated at their cytosine residues as an epigenetic mark that is stably inherited during cell mitosis. Differentially methylated regions (DMRs) are genomic regions showing different degrees of DNA methylation in multiple samples. In this study, we focused our attention on CGIs showing different DNA methylation between two culture replicas of the same cell line. Results We used methylation data of 35 cell lines from the Encyclopedia of DNA Elements (ENCODE) consortium to identify CpG islands that were differentially methylated between replicas of the same cell line and denoted them Inter Replicas Differentially Methylated CpG islands (IRDM-CGIs). We identified a group of IRDM-CGIs that was consistently shared by different cell lines, and denoted it common IRDM-CGIs. X chromosome CGIs were overrepresented among common IRDM-CGIs. Autosomal IRDM-CGIs were preferentially located in gene bodies and intergenic regions had a lower G + C content, a smaller mean length, and a reduced CpG percentage. Functional analysis of the genes associated with autosomal IRDM-CGIs showed that many of them are involved in DNA binding and development. Conclusions Our results show that several specific functional and structural features characterize common IRDM-CGIs. They may represent a specific subset of CGIs that are more prone to being differentially methylated for their intrinsic characteristics. PMID:24106769

  13. A feasible approach to evaluate the relative reactivity of NHS-ester activated group with primary amine-derivatized DNA analogue and non-derivatized impurity.

    PubMed

    Dou, Shuping; Virostko, John; Greiner, Dale L; Powers, Alvin C; Liu, Guozheng

    2015-01-01

    Synthetic DNA analogues with improved stability are widely used in life science. The 3'and/or 5' equivalent terminuses are often derivatized by attaching an active group for further modification, but a certain amount of non-derivatized impurity often remains. It is important to know to what extent the impurity would influence further modification. The reaction of an NHS ester with primary amine is one of the most widely used options to modify DNA analogues. In this short communication, a 3'-(NH2-biotin)-derivatized morpholino DNA analogue (MORF) was utilized as the model derivatized DNA analogue. Inclusion of a biotin concomitant with the primary amine at the 3'-terminus allows for the use of streptavidin to discriminate between the products from the derivatized MORF and non-derivatized MORF impurity. To detect the MORF reaction with NHS ester, S-acetyl NHS-MAG3 was conjugated to the DNA analogue for labeling with (99m)Tc, a widely used nuclide in the clinic. It was found that the non-derivatized MORF also reacted with the S-acetyl NHS-MAG3. Radiolabeling of the product yielded an equally high labeling efficiency. Nevertheless, streptavidin binding indicated that under the conditions of this investigation, the non-derivatized MORF was five times less reactive than the amine-derivatized MORF. PMID:25621701

  14. Tuning the DNA Conformational Perturbations Induced by Cytotoxic Platinum-Acridine Bisintercalators: Effect of Metal cis/trans Isomerism and DNA Threading Groups

    PubMed Central

    Choudhury, Jayati Roy; Guddneppanavar, Rajsekhar; Saluta, Gilda; Kucera, Gregory L.; Bierbach, Ulrich

    2009-01-01

    Four highly charged, water soluble platinum-acridine bisintercalating agents have been synthesized. Depending on the cis/trans isomerism of the metal and the nature of the acridine side chains, bisintercalation induces/stabilizes the classical Watson-Crick B-form or a non-B-form. Circular dichroism spectra and chemical footprinting experiments suggest that compound 4, the most active derivative in HL-60 cells, produces a structurally severely perturbed DNA with features of a Hoogsteen base-paired biopolymer. PMID:18457380

  15. Identification of a group of cryptic marine limpet species, Cellana karachiensis (Mollusca: Patellogastropoda) off Veraval coast, India, using mtDNA COI sequencing.

    PubMed

    Joseph, Sneha; Poriya, Paresh; Vakani, Bhavik; Singh, S P; Kundu, Rahul

    2016-01-01

    Present communication reports the phylogenetic relationship between three groups of a marine limpet having different color banding patterns using COI sequencing. Samples were sequenced for mtDNA COI gene using universal primer. Comparative BLAST revealed that all three types were around 99.59% identical with Cellana karachiensis, first record of this species from Indian coasts. Apart from the morphological variations, the mtDNA COI gene analysis revealed around 1% nucleotide variations between these three types. The observed dissimilarity in COI sequences was possibly too little to consider these types as three different species. The derivation of amino acid positions indicated that these types could possibly be a complex of three cryptic species of C. karachiensis. The study proposes that the Oman and Indian populations of C. karachiensis might have derived by allopatric speciation due to geographical isolation. The group of these three cryptic species, sharing same habitat between themselves, possibly showed sympatric speciation. PMID:25109628

  16. Synthesis of S-Adenosyl-L-Methionine Analogs with Extended Transferable Groups for Methyltransferase-Directed Labeling of DNA and RNA.

    PubMed

    Masevičius, Viktoras; Nainytė, Milda; Klimašauskas, Saulius

    2016-01-01

    S-Adenosyl-L-methionine (AdoMet) is a ubiquitous methyl donor for a variety of biological methylation reactions catalyzed by methyltransferases (MTases). AdoMet analogs with extended propargylic chains replacing the sulfonium-bound methyl group can serve as surrogate cofactors for many DNA and RNA MTases enabling covalent deposition of these linear chains to their cognate targets sites in DNA or RNA. Here we describe synthetic procedures for the preparation of two representative examples of AdoMet analogs with a transferable hex-2-ynyl group carrying a terminal azide or amine functionality. Our approach is based on direct chemoselective alkylation of S-adenosyl-L-homocysteine at sulfur with corresponding nosylates under acidic conditions. We also describe synthetic routes to 6-substituted hex-2-yn-1-ols and their conversion to the corresponding nosylates. Using these protocols, synthetic AdoMet analogs can be prepared within 1 to 2 weeks. PMID:26967468

  17. Mutational analysis of Mycobacterium UvrD1 identifies functional groups required for ATP hydrolysis, DNA unwinding, and chemomechanical coupling.

    PubMed

    Sinha, Krishna Murari; Glickman, Michael S; Shuman, Stewart

    2009-05-19

    Mycobacterial UvrD1 is a DNA-dependent ATPase and a Ku-dependent 3' to 5' DNA helicase. The UvrD1 motor domain resembles that of the prototypal superfamily I helicases UvrD and PcrA. Here we performed a mutational analysis of UvrD1 guided by the crystal structure of a DNA-bound Escherichia coli UvrD-ADP-MgF(3) transition state mimetic. Alanine scanning and conservative substitutions identified amino acids essential for both ATP hydrolysis and duplex unwinding, including those implicated in phosphohydrolase chemistry via transition state stabilization (Arg308, Arg648, Gln275), divalent cation coordination (Glu236), or activation of the nucleophilic water (Glu236, Gln275). Other residues important for ATPase/helicase activity include Phe280 and Phe72, which interact with the DNA 3' single strand tail. ATP hydrolysis was uncoupled from duplex unwinding by mutations at Glu609 (in helicase motif V), which contacts the ATP ribose sugar. Introducing alanine in lieu of the adenine-binding "Q motif" glutamine (Gln24) relaxed the substrate specificity in NTP hydrolysis, e.g., eliciting a gain of function as a UTPase/TTPase, although the Q24A mutant still relied on ATP/dATP for duplex unwinding. Our studies highlight the role of the Q motif as a substrate filter and the contributions of adenosine-binding residues as couplers of NTP hydrolysis to motor activity. The Ku-binding function of UvrD1 lies within its C-terminal 270 amino acid segment. Here we found that deleting the 90 amino acid C-terminal domain, which is structurally uncharacterized, diminished DNA unwinding, without affecting ATP hydrolysis or binding to the DNA helicase substrate, apparently by affecting the strength of the UvrD1-Ku interaction. PMID:19317511

  18. Complete structure of nuclear rDNA of the obligate plant parasite Plasmodiophora brassicae: intraspecific polymorphisms in the exon and group I intron of the large subunit rDNA.

    PubMed

    Niwa, Rieko; Kawahara, Ai; Murakami, Hiroharu; Tanaka, Shuhei; Ezawa, Tatsuhiro

    2011-07-01

    Plasmodiophora brassicae is a soil-borne obligate intracellular parasite in the phylum Cercozoa of the Rhizaria that causes clubroot disease of crucifer crops. To control the disease, understanding the distribution and infection routes of the pathogen is essential, and thus development of reliable molecular markers to discriminate geographic populations is required. In this study, the nuclear ribosomal RNA gene (rDNA) repeat unit of P. brassicae was determined, with particular emphasis on the structure of large subunit (LSU) rDNA, in which polymorphic regions were expected to be present. The complete rDNA complex was 9513bp long, which included the small subunit, 5.8S and LSU rDNAs as well as the internal transcribed spacer and intergenic spacer regions. Among eight field populations collected from throughout Honshu Island, Japan, a 1.1 kbp region of the LSU rDNA, including the divergent 8 domain, exhibited intraspecific polymorphisms that reflected geographic isolation of the populations. Two new group I introns were found in this region in six out of the eight populations, and the sequences also reflected their geographic isolation. The polymorphic region found in this study may have potential for the development of molecular markers for discrimination of field populations/isolates of this organism. PMID:21497131

  19. mtDNA G10398A variation provides risk to type 2 diabetes in population group from the Jammu region of India

    PubMed Central

    Sharma, Varun; Sharma, Indu; Singh, Vishav Pratap; Verma, Sonali; Pandita, Anil; Singh, Vinod; Rai, Ekta; Sharma, Swarkar

    2014-01-01

    Mitochondrion plays an integral role in glucose metabolism and insulin secretion. Mitochondrial electron-transport chain (ETC) is involved in adenosine triphosphate (ATP) generation and ATP mediated insulin secretion in pancreatic β-cells. β-cell dysfunction is a critical component in the pathogenesis of type 2 diabetes (T2D). The mtDNA G10398A variation (amino acid change: Alanine → Threonine) within the NADH dehydrogenase (ND3) subunit of complex I of mtDNA ETC, has emerged as a variation of clinical significance in various disorders including T2D. This variation is supposed to result in altered complex I function, leading to an increased rate of electron leakage and reactive oxygen species (ROS) production, which might cause β-cell damage and impaired insulin secretion. The aim of the study was to explore the association of mtDNA G10398A variation with T2D in a total of 439 samples (196 T2D cases and 243 healthy controls) belonging to the Jammu region of Jammu and Kashmir (J&K). The candidate gene association analyses showed significant association of mtDNA G10398A variant with T2D and the estimated odds ratio (OR) was 2.83 (1.64–4.90 at 95% CI) in the studied population group. The extent of genetic heterogeneity in T2D and diversity of the Indian population groups, make such replication studies pertinent to understand the etiology of T2D in these population groups. PMID:25606409

  20. Comparative restriction endonuclease maps of proviral DNA of the primate type C simian sarcoma-associated virus and gibbon ape leukemia virus group.

    PubMed Central

    Trainor, C D; Wong-Staal, F; Reitz, M S

    1982-01-01

    Extrachromosomal DNA was purified from canine thymus cells acutely infected with different strains of infectious primate type C viruses of the woolly monkey (simian) sarcoma helper virus and gibbon ape leukemia virus group. All DNA preparations contained linear proviral molecules of 9.1 to 9.2 kilobases, at least some of which represent complete infectious proviral DNA. Cells infected with a replication-defective fibroblast-transforming sarcoma virus and its helper, a replication-competent nontransforming helper virus, also contained a 6.6- to 6.7-kilobase DNA. These proviral DNA molecules were digested with different restriction endonucleases, and the resultant fragments were oriented to the viral RNA by a combination of partial digestions, codigestion with more than one endonuclease, digestion of integrated proviral DNA, and hybridization with 3'- and 5'-specific viral probes. The 3'- and 5'-specific probes each hybridized to fragments from both ends of proviral DNA, indicating that, in common with those of other retroviruses, these proviruses contain a large terminal redundancy at both ends, each of which consists of sequences derived from both the 3' and 5' regions of the viral RNA. The proviral sequences are organized 3',5'-unique-3',5'. Four restriction enzymes (KpnI, SmaI, PstI, and SstI) recognized sites within the large terminal redundancies, and these sites were conserved within all the isolates tested. This suggests that both the 3' and 5' ends of the genomic RNA of these viruses are extremely closely related. In contrast, the restriction sites within the unique portion of the provirus were not strongly conserved within this group of viruses, even though they were related along most of their genomes. Whereas the 5' 60 to 70% of the RNA of these viruses was more closely related by liquid hybridization experiments than was the 3' 30 to 40%, restriction sites within this region were not preferentially conserved, suggesting that small sequence differences or

  1. European Mitochondrial DNA Haplogroups and Metabolic Changes during Antiretroviral Therapy in AIDS Clinical Trials Group Study A5142

    PubMed Central

    Hulgan, Todd; Haubrich, Richard; Riddler, Sharon A.; Tebas, Pablo; Ritchie, Marylyn D.; McComsey, Grace A.; Haas, David W.; Canter, Jeffrey A.

    2010-01-01

    Background Mitochondrial DNA (mtDNA) influences metabolic diseases and perhaps antiretroviral therapy (ART) complications. We explored associations between European mtDNA haplogroups and metabolic changes among A5142 participants. Methods 757 ART-naïve subjects were randomized to one of three class-sparing ART regimens including efavirenz and/or lopinavir/ritonavir with or without nucleoside reverse transcriptase inhibitors (NRTIs). Non-randomized NRTIs included stavudine, tenofovir, or zidovudine, each with lamivudine. Fasting lipid profiles and whole-body dual-energy X-ray absorptiometry (DEXA) were performed. Nine European mtDNA haplogroups were determined for 231 self-identified non-Hispanic white subjects. Metabolic changes from baseline to 96 weeks were analyzed by haplogroup. Results Median age was 39 years, 9% were female, and 37%, 32%, and 30% were randomized to NRTI-containing regimens with either efavirenz or lopinavir/ritonavir, and an NRTI-sparing regimen respectively. Among NRTI-containing regimens, 51% included zidovudine, 28% tenofovir, and 21% stavudine. Compared with other haplogroups, mtDNA haplogroup I (N=10) had higher baseline non-HDL cholesterol (160 mg/dL [interquartile range 137–171] vs. 120 mg/dL [104–136]; p=0.005), a decrease in non-HDL cholesterol over 96 weeks (−14% [−20-+6] vs. +25% [+8-+51]; p<0.001), tended to have more baseline extremity fat, and had more extremity fat loss by DEXA (−13% [−31-+12] vs. +9% [−13-+26]; p=0.08) and lipoatrophy (50% vs. 20%; p=0.04). Haplogroup W (N=5; all randomized to NRTI-sparing regimens) had the greatest increase in extremity fat (+35.5% [+26.8 - +54.9]; P=0.02). Conclusions Lipids and extremity fat were associated with European mtDNA haplogroups in this HIV-infected population. These preliminary results suggest that mitochondrial genomics may influence metabolic parameters before and during ART. PMID:20871389

  2. A group of chromosomal proteins is specifically released by spermine and loses DNA-binding activity upon phosphorylation.

    PubMed Central

    Van den Broeck, D; Van der Straeten, D; Van Montagu, M; Caplan, A

    1994-01-01

    Biologically relevant concentrations as low as 500 microM spermine led to the specific release of chromatin-associated proteins from nuclei of rice (Oryza sativa) seedlings. Using a southwestern technique, it was shown that several of these proteins bind DNA. This affinity was lost upon in organello phosphorylation by an endogenous kinase. The effect of spermine was very specific. Spermidine was far less effective and putrescine was essentially ineffective in releasing these proteins. The most abundant spermine-released protein was shown to be homologous to the maize HMG1 protein. Our results suggest that spermine induces the release of spermine-released proteins by changing DNA conformation. Binding of these proteins might be sensitive to long-range changes in chromosome structure caused by torsional stress. PMID:7991684

  3. Both the folate cycle and betaine-homocysteine methyltransferase contribute methyl groups for DNA methylation in mouse blastocysts.

    PubMed

    Zhang, Baohua; Denomme, Michelle M; White, Carlee R; Leung, Kit-Yi; Lee, Martin B; Greene, Nicholas D E; Mann, Mellissa R W; Trasler, Jacquetta M; Baltz, Jay M

    2015-03-01

    The embryonic pattern of global DNA methylation is first established in the inner cell mass (ICM) of the mouse blastocyst. The methyl donor S-adenosylmethionine (SAM) is produced in most cells through the folate cycle, but only a few cell types generate SAM from betaine (N,N,N-trimethylglycine) via betaine-homocysteine methyltransferase (BHMT), which is expressed in the mouse ICM. Here, mean ICM cell numbers decreased from 18-19 in controls to 11-13 when the folate cycle was inhibited by the antifolate methotrexate and to 12-14 when BHMT expression was knocked down by antisense morpholinos. Inhibiting both pathways, however, much more severely affected ICM development (7-8 cells). Total SAM levels in mouse blastocysts decreased significantly only when both pathways were inhibited (from 3.1 to 1.6 pmol/100 blastocysts). DNA methylation, detected as 5-methylcytosine (5-MeC) immunofluorescence in isolated ICMs, was minimally affected by inhibition of either pathway alone but decreased by at least 45-55% when both BHMT and the folate cycle were inhibited simultaneously. Effects on cell numbers and 5-MeC levels in the ICM were completely rescued by methionine (immediate SAM precursor) or SAM. Both the folate cycle and betaine/BHMT appear to contribute to a methyl pool required for normal ICM development and establishing initial embryonic DNA methylation. PMID:25466894

  4. DNA Barcoding of Rhodiola (Crassulaceae): A Case Study on a Group of Recently Diversified Medicinal Plants from the Qinghai-Tibetan Plateau

    PubMed Central

    Zhang, Jian-Qiang; Meng, Shi-Yong; Wen, Jun; Rao, Guang-Yuan

    2015-01-01

    DNA barcoding, the identification of species using one or a few short standardized DNA sequences, is an important complement to traditional taxonomy. However, there are particular challenges for barcoding plants, especially for species with complex evolutionary histories. We herein evaluated the utility of five candidate sequences — rbcL, matK, trnH-psbA, trnL-F and the internal transcribed spacer (ITS) — for barcoding Rhodiola species, a group of high-altitude plants frequently used as adaptogens, hemostatics and tonics in traditional Tibetan medicine. Rhodiola was suggested to have diversified rapidly recently. The genus is thus a good model for testing DNA barcoding strategies for recently diversified medicinal plants. This study analyzed 189 accessions, representing 47 of the 55 recognized Rhodiola species in the Flora of China treatment. Based on intraspecific and interspecific divergence and degree of monophyly statistics, ITS was the best single-locus barcode, resolving 66% of the Rhodiola species. The core combination rbcL+matK resolved only 40.4% of them. Unsurprisingly, the combined use of all five loci provided the highest discrimination power, resolving 80.9% of the species. However, this is weaker than the discrimination power generally reported in barcoding studies of other plant taxa. The observed complications may be due to the recent diversification, incomplete lineage sorting and reticulate evolution of the genus. These processes are common features of numerous plant groups in the high-altitude regions of the Qinghai-Tibetan Plateau. PMID:25774915

  5. Thermostable group II intron reverse transcriptase fusion proteins and their use in cDNA synthesis and next-generation RNA sequencing

    PubMed Central

    Mohr, Sabine; Ghanem, Eman; Smith, Whitney; Sheeter, Dennis; Qin, Yidan; King, Olga; Polioudakis, Damon; Iyer, Vishwanath R.; Hunicke-Smith, Scott; Swamy, Sajani; Kuersten, Scott; Lambowitz, Alan M.

    2013-01-01

    Mobile group II introns encode reverse transcriptases (RTs) that function in intron mobility (“retrohoming”) by a process that requires reverse transcription of a highly structured, 2–2.5-kb intron RNA with high processivity and fidelity. Although the latter properties are potentially useful for applications in cDNA synthesis and next-generation RNA sequencing (RNA-seq), group II intron RTs have been difficult to purify free of the intron RNA, and their utility as research tools has not been investigated systematically. Here, we developed general methods for the high-level expression and purification of group II intron-encoded RTs as fusion proteins with a rigidly linked, noncleavable solubility tag, and we applied them to group II intron RTs from bacterial thermophiles. We thus obtained thermostable group II intron RT fusion proteins that have higher processivity, fidelity, and thermostability than retroviral RTs, synthesize cDNAs at temperatures up to 81°C, and have significant advantages for qRT-PCR, capillary electrophoresis for RNA-structure mapping, and next-generation RNA sequencing. Further, we find that group II intron RTs differ from the retroviral enzymes in template switching with minimal base-pairing to the 3′ ends of new RNA templates, making it possible to efficiently and seamlessly link adaptors containing PCR-primer binding sites to cDNA ends without an RNA ligase step. This novel template-switching activity enables facile and less biased cloning of nonpolyadenylated RNAs, such as miRNAs or protein-bound RNA fragments. Our findings demonstrate novel biochemical activities and inherent advantages of group II intron RTs for research, biotechnological, and diagnostic methods, with potentially wide applications. PMID:23697550

  6. Thermostable group II intron reverse transcriptase fusion proteins and their use in cDNA synthesis and next-generation RNA sequencing.

    PubMed

    Mohr, Sabine; Ghanem, Eman; Smith, Whitney; Sheeter, Dennis; Qin, Yidan; King, Olga; Polioudakis, Damon; Iyer, Vishwanath R; Hunicke-Smith, Scott; Swamy, Sajani; Kuersten, Scott; Lambowitz, Alan M

    2013-07-01

    Mobile group II introns encode reverse transcriptases (RTs) that function in intron mobility ("retrohoming") by a process that requires reverse transcription of a highly structured, 2-2.5-kb intron RNA with high processivity and fidelity. Although the latter properties are potentially useful for applications in cDNA synthesis and next-generation RNA sequencing (RNA-seq), group II intron RTs have been difficult to purify free of the intron RNA, and their utility as research tools has not been investigated systematically. Here, we developed general methods for the high-level expression and purification of group II intron-encoded RTs as fusion proteins with a rigidly linked, noncleavable solubility tag, and we applied them to group II intron RTs from bacterial thermophiles. We thus obtained thermostable group II intron RT fusion proteins that have higher processivity, fidelity, and thermostability than retroviral RTs, synthesize cDNAs at temperatures up to 81°C, and have significant advantages for qRT-PCR, capillary electrophoresis for RNA-structure mapping, and next-generation RNA sequencing. Further, we find that group II intron RTs differ from the retroviral enzymes in template switching with minimal base-pairing to the 3' ends of new RNA templates, making it possible to efficiently and seamlessly link adaptors containing PCR-primer binding sites to cDNA ends without an RNA ligase step. This novel template-switching activity enables facile and less biased cloning of nonpolyadenylated RNAs, such as miRNAs or protein-bound RNA fragments. Our findings demonstrate novel biochemical activities and inherent advantages of group II intron RTs for research, biotechnological, and diagnostic methods, with potentially wide applications. PMID:23697550

  7. Towards zirconium phosphonate-based microarrays for probing DNA-protein interactions: critical influence of the location of the probe anchoring groups.

    PubMed

    Monot, Julien; Petit, Marc; Lane, Sarah M; Guisle, Isabelle; Léger, Jean; Tellier, Charles; Talham, Daniel R; Bujoli, Bruno

    2008-05-14

    Terminal phosphate groups on double-stranded DNA probes bind strongly to glass substrates coated with a zirconium phosphonate monolayer, and probes immobilized in this way as microarrays can be used to detect protein targets. The sensitivity of the microarray was shown to be enhanced by the use of a polyguanine segment ((G)n , n > or = 5) as a spacer between the phosphate linker and the protein interaction domain. More importantly, the presence of phosphate linkers on both ends of the dsDNA probes leads to significant enhancement of target capture. The relevant characteristics of the different probes when bound to the surface were determined, by the original use of a combination of surface characterization techniques (XPS, AFM, and Sarfus). In this context, the location of the phosphate linkers in the duplex probes was found to result in different probe surface coverage and presentation on the surface, which affect subsequent interactions with the target protein. PMID:18407629

  8. Prognostic implications of DNA aneuploidy in 156 untreated multiple myeloma patients. Castelano-Leonés (Spain) Cooperative Group for the Study of Monoclonal Gammopathies.

    PubMed

    García-Sanz, R; Orfão, A; González, M; Moro, M J; Hernández, J M; Ortega, F; Borrego, D; Carnero, M; Casanova, F; Jiménez, R

    1995-05-01

    In this study the incidence of DNA aneuploidy in a large series of untreated multiple myeloma (MM) patients was assessed in order to determine its clinical and prognostic significance. A total of 156 MM patients were included in the study. DNA measurements were performed in all cases at diagnosis using two different flow cytometry methods: (1) propidium iodide (PI) staining on isolated nuclei, and (2) CD38/PI double staining on whole cells. The DNA ploidy status was correlated with the most relevant clinical and haematological disease characteristics. From the 156 cases analysed, 91 (58%) were aneuploid (56% hyperdiploid and 2% hypodiploid). The correlation between the two techniques on the detection of DNA aneuploidy was excellent, although CD38/PI double staining would be preferable in cases with < 5% of DNA aneuploid plasma cells (PC). Upon comparing the clinical and haematological disease characteristics of hyperdiploid versus diploid cases, the former group was characterized by a lower age, reduced incidence of anaemia, lower beta 2M levels, higher proliferative activity within the residual normal haemopoietic cells, increased expression of CD56 antigen in PC, and higher proportion of PB CD4+ T cells. In contrast, diploid cases had a higher expression of the CD10, CD20 and CD15 antigens and greater numbers of PB CD56+CD3- NK cells (P < 0.05). Circulating PC were identified in six cases, all being diploid. Overall survival was significantly longer in hyperdiploid compared to diploid MM (P = 0.02). These results show that over 50% of MM patients are aneuploid, almost all of them being hyperdiploid. This characteristic is associated with better prognosis. PMID:7786771

  9. A Computational Study of the Hydrolysis of dGTP Analogues with Halomethylene Modified Leaving Groups in Solution: Implications for the Mechanism of DNA Polymerases

    PubMed Central

    Kamerlin, Shina C. L.; McKenna, Charles E.; Goodman, Myron F.; Warshel, A.

    2009-01-01

    DNA polymerases are a family of enzymes responsible for regulating DNA replication and repair, which in turn maintains the integrity of the genome. However, despite intensive kinetic, crystallographic and computational studies, elucidation of the detailed enzymatic mechanism still presents a significant challenge. We recently developed an alternative strategy for exploring the fidelity and mechanism of DNA polymerases, by probing leaving group effects on nucleotidyl transfer using a series of dGTP bisphosphonate analogues in which the β, γ-bridging oxygen was replaced by a series of substituted methylene groups (X=CYZ, where Y,Z=H, halogen or other substituent). Pre-steady state kinetic measurements of DNA polymerase-catalyzed incorporation of correctly base paired (R) and mispaired (W) analogues demonstrated a strong linear free energy relationship (LFER) between the polymerase rate constant (kpol) and the highest pKa of the free bisphosphonic acid corresponding to the leaving group. However, unexpectedly, the data segregated into two distinctly different linear correlations depending on the nature of the substituent. The discrepancy between the two lines was considerably greater when the dGTP analogue formed an incorrect (G•T) rather than a correct (G•C) base pair, although the reason for this phenomenon remains unexplained. Here, we have evaluated the complete free energy surfaces for bisphosphonate hydrolysis in aqueous solution and evaluated the corresponding LFER. Our study, which employs several alternative solvation models, finds a split of the calculated LFER for the mono- and dihalogen compounds into two parallel lines, reflecting their behavior in the polymerase-catalyzed condensation reaction. We suggest that the division into two linear subsets may be a generalized solvation phenomenon involving the overall electrostatic interaction between the substrates and enzyme and would be observed in solution in the absence of the enzyme. In contrast, the

  10. Characterization of I-Ppo, an intron-encoded endonuclease that mediates homing of a group I intron in the ribosomal DNA of Physarum polycephalum.

    PubMed Central

    Muscarella, D E; Ellison, E L; Ruoff, B M; Vogt, V M

    1990-01-01

    A novel and only recently recognized class of enzymes is composed of the site-specific endonucleases encoded by some group I introns. We have characterized several aspects of I-Ppo, the endonuclease that mediates the mobility of intron 3 in the ribosomal DNA of Physarum polycephalum. This intron is unique among mobile group I introns in that it is located in nuclear DNA. We found that I-Ppo is encoded by an open reading frame in the 5' half of intron 3, upstream of the sequences required for self-splicing of group I introns. Either of two AUG initiation codons could start this reading frame, one near the beginning of the intron and the other in the upstream exon, leading to predicted polypeptides of 138 and 160 amino acid residues. The longer polypeptide was the major form translated in vitro in a reticulocyte extract. From nuclease assays of proteins synthesized in vitro with partially deleted DNAs, we conclude that both polypeptides possess endonuclease activity. We also have expressed I-Ppo in Escherichia coli, using a bacteriophage T7 RNA polymerase expression system. The longer polypeptide also was the predominant form made in this system. It showed enzymatic activity in bacteria in vivo, as demonstrated by the cleavage of a plasmid carrying the target site. Like several other intron-encoded endonucleases, I-Ppo makes a four-base staggered cut in its ribosomal DNA target sequence, very near the site where intron 3 becomes integrated in crosses of intron 3-containing and intron 3-lacking Physarum strains. Images PMID:2355911

  11. Ampelomyces mycoparasites from apple powdery mildew identified as a distinct group based on single-stranded conformation polymorphism analysis of the rDNA ITS region.

    PubMed

    Szentiványi, Orsolya; Kiss, Levente; Russell, John C; Kovács, Gábor M; Varga, Krisztina; Jankovics, Tünde; Lesemann, Silke; Xu, Xiang-Ming; Jeffries, Peter

    2005-04-01

    Pycnidial fungi belonging to the genus Ampelomyces are the most common natural antagonists of powdery mildews worldwide. During a study of the interactions between apple powdery mildew (Podosphaera leucotricha) and Ampelomyces mycoparasites, 52 new Ampelomyces isolates were obtained from P. leucotricha and, in addition, 13 new isolates from other species of the Erysiphaceae in four European countries. Their genetic diversity was screened using single-stranded conformation polymorphism (SSCP) analysis of the internal transcribed spacer (ITS) region of the ribosomal DNA (rDNA). For comparison, 24 isolates obtained from genetic resource collections or other sources were included in this study. Based on the ITS-SSCP patterns, the isolates were placed in eight groups. The isolates belonged to two types based on their growth in culture. The faster-growing and the slower-growing isolates were included in different SSCP groups. A phylogenetic analysis of the ITS sequences of representatives of these groups confirmed the results obtained with the SSCP method, and showed that the faster-growing isolates do not belong to Ampelomyces as suggested by earlier studies. All the isolates from P. leucotricha fell into a distinct SSCP group of genetically homogeneous isolates. This suggests that Ampelomyces mycoparasites which occur in apple powdery mildew are slightly different from the other Ampelomyces groups which contain mycoparasites from various powdery mildew species. This may be because the main growth period of Ampelomyces mycoparasites in apple powdery mildew is isolated in time from that of Ampelomyces isolates that occur in other species of the Erysiphaceae. P. leucotricha starts its life-cycle early in the season, usually in March-April, while most powdery mildews are active in the same environments only late in the year. PMID:15912930

  12. Synthesis and Structure of a New Copper(II) Coordination Polymer Alternately Bridged by Oxamido and Carboxylate Groups: Evaluation of DNA/BSA Binding and Cytotoxic Activities.

    PubMed

    Jin, Xiao-Ting; Zheng, Kang; Li, Yan-Tuan; Wu, Zhi-Yong; Yan, Cui-Wei

    2015-08-01

    A new one-dimensional (1D) copper(II) coordination polymer {[Cu2 (dmaepox)(dabt)](NO3) · 0.5 H2 O}n , where H3 dmaepox and dabt denote N-benzoato-N'-(3-methylaminopropyl)oxamide and 2,2'-diamino-4,4'-bithiazole, respectively, was synthesized and characterized by single-crystal X-ray diffraction and other methods. The crystal structure analysis revealed that the two copper(II) ions are bridged alternately by cis-oxamido and carboxylato groups to form a 1-D coordination polymer with the corresponding Cu · · · Cu separations of 5.1946(19) and 5.038(2) Å. There is a three-dimensional supramolecular structure constructed by hydrogen bonding and π-π stacking interactions in the crystal. The reactivity towards herring sperm DNA (HS-DNA) and bovine serum albumin (BSA) indicated that the copper(II) polymer can interact with the DNA in the mode of intercalation, and bind to BSA responsible for quenching of tryptophan fluorescence by the static quenching mechanism. The in vitro cytotoxicity suggested that the copper(II) polymer exhibits cytotoxic effects against the selected tumor cell lines. PMID:25940657

  13. Adjusting the DNA Interaction and Anticancer Activity of Pt(II) N-Heterocyclic Carbene Complexes by Steric Shielding of the Trans Leaving Group.

    PubMed

    Muenzner, Julienne K; Rehm, Tobias; Biersack, Bernhard; Casini, Angela; de Graaf, Inge A M; Worawutputtapong, Pawida; Noor, Awal; Kempe, Rhett; Brabec, Viktor; Kasparkova, Jana; Schobert, Rainer

    2015-08-13

    Five platinum(II) complexes bearing a (1,3-dibenzyl)imidazol-2-ylidene ligand but different leaving groups trans to it were examined for cytotoxicity, DNA and cell cycle interference, vascular disrupting properties, and nephrotoxicity. The cytotoxicity of complexes 3a-c increased with the steric shielding of their leaving chloride ligand, and complex 3c, featuring two triphenylphosphanes, was the most efficacious, with submicromolar IC50 concentrations. Complexes 3a-c interacted with DNA in electrophoretic mobility shift and ethidium bromide binding assays. The cationic complex 3c did not bind coordinatively to DNA but led to its aggregation, damage that is not amenable to the usual repair mechanisms. Accordingly, it arrested the cell cycle of melanoma cells in G1 phase, whereas cis-dichlorido[(1,3-dibenzyl)imidazol-2-ylidene](dimethyl sulfoxide) platinum(II) 3a induced G2/M phase arrest. Complex 3c also disrupted the blood vessels in the chorioallantoic membrane of fertilized chicken eggs. Ex vivo studies using precision-cut tissue slices suggested the nephrotoxicities of 3a-c to be clinically manageable. PMID:26182125

  14. HPV E6/E7 mRNA versus HPV DNA biomarker in cervical cancer screening of a group of Macedonian women.

    PubMed

    Duvlis, Sotirija; Popovska-Jankovic, Katerina; Arsova, Zorica Sarafinovska; Memeti, Shaban; Popeska, Zaneta; Plaseska-Karanfilska, Dijana

    2015-09-01

    High risk types of human papillomaviruses E6/E7 oncogenes and their association with tumor suppressor genes products are the key factors of cervical carcinogenesis. This study proposed them as specific markers for cervical dysplasia screening. The aim of the study is to compare the clinical and prognostic significance of HPV E6/E7 mRNA as an early biomarker versus HPV DNA detection and cytology in triage of woman for cervical cancer. The study group consists of 413 women: 258 NILM, 26 ASC-US, 81 LSIL, 41 HSIL, and 7 unsatisfactory cytology. HPV4AACE screening, real-time multiplex PCR and MY09/11 consensus PCR primers methods were used for the HPV DNA detection. The real-time multiplex nucleic acid sequence-based assay (NucliSENS EasyQ HPV assay) was used for HPV E6/E7 mRNA detection of the five most common high risk HPV types in cervical cancer (16, 18, 31, 33, and 45). The results show that HPV E6/E7 mRNA testing had a higher specificity 50% (95% CI 32-67) and positive predictive value (PPV) 62% (95% CI 46-76) for CIN2+ compared to HPV DNA testing that had specificity of 18% (95% CI 7-37) and PPV 52% (95% CI 39-76) respectively. The higher specificity and PPV of HPV E6/E7 mRNA testing are valuable in predicting insignificant HPV DNA infection among cases with borderline cytological finding. It can help in avoiding aggressive procedures (biopsies and over-referral of transient HPV infections) as well as lowering patient's anxiety and follow up period. PMID:25880030

  15. Analysis of the distribution and evolution of the ATP-dependent DNA ligases of bacteria delineates a distinct phylogenetic group 'Lig E'.

    PubMed

    Williamson, Adele; Hjerde, Erik; Kahlke, Tim

    2016-01-01

    Prior to the discovery of a minimal ATP-dependent DNA ligase in Haemophilus influenzae, bacteria were thought to only possess a NAD-dependent ligase, which was involved in sealing of Okazaki fragments. We now know that a diverse range of bacterial species possess up to six of these accessory bacterial ATP-dependent DNA ligases (b-ADLs), which vary in size and enzymatic domain associations. Here we compare the domain structure of different types of b-ADLs and investigate their distribution among the bacterial domain to describe possible evolutionary trajectories that gave rise to the sequence and structural diversity of these enzymes. Previous biochemical and genetic analyses have delineated three main classes of these enzymes: Lig B, Lig C and Lig D, which appear to have descended from a common ancestor within the bacterial domain. In the present study, we delineate a fourth group of b-ADLs, Lig E, which possesses a number of unique features at the primary and tertiary structural levels. The biochemical characteristics, domain structure and inferred extracellular location sets this group apart from the other b-ADLs. The results presented here indicate that the Lig E type ligases were horizontally transferred into bacteria in a separate event from other b-ADLs possibly from a bacteriophage. PMID:26412580

  16. MtDNA Haplogroup A10 Lineages in Bronze Age Samples Suggest That Ancient Autochthonous Human Groups Contributed to the Specificity of the Indigenous West Siberian Population

    PubMed Central

    Pilipenko, Aleksandr S.; Trapezov, Rostislav O.; Zhuravlev, Anton A.; Molodin, Vyacheslav I.; Romaschenko, Aida G.

    2015-01-01

    Background The craniometric specificity of the indigenous West Siberian human populations cannot be completely explained by the genetic interactions of the western and eastern Eurasian groups recorded in the archaeology of the area from the beginning of the 2nd millennium BC. Anthropologists have proposed another probable explanation: contribution to the genetic structure of West Siberian indigenous populations by ancient human groups, which separated from western and eastern Eurasian populations before the final formation of their phenotypic and genetic features and evolved independently in the region over a long period of time. This hypothesis remains untested. From the genetic point of view, it could be confirmed by the presence in the gene pool of indigenous populations of autochthonous components that evolved in the region over long time periods. The detection of such components, particularly in the mtDNA gene pool, is crucial for further clarification of early regional genetic history. Results and Conclusion We present the results of analysis of mtDNA samples (n = 10) belonging to the A10 haplogroup, from Bronze Age populations of West Siberian forest-steppe (V—I millennium BC), that were identified in a screening study of a large diachronic sample (n = 96). A10 lineages, which are very rare in modern Eurasian populations, were found in all the Bronze Age groups under study. Data on the A10 lineages’ phylogeny and phylogeography in ancient West Siberian and modern Eurasian populations suggest that A10 haplogroup underwent a long-term evolution in West Siberia or arose there autochthonously; thus, the presence of A10 lineages indicates the possible contribution of early autochthonous human groups to the genetic specificity of modern populations, in addition to contributions of later interactions of western and eastern Eurasian populations. PMID:25950581

  17. DNA fingerprinting and anastomosis grouping reveal similar genetic diversity in Rhizoctonia species infecting turfgrasses in the transition zone of USA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rhizoctonia blight (sensu lato) is a common and serious disease of many turfgrass species. The most widespread causal agent, R. solani, consists of several genetically different subpopulations. Though hyphal anastomosis reactions have been used to group Rhizoctonia species, they are time consuming a...

  18. Phylogenetic analysis of LSU and SSU rDNA group I introns of lichen photobionts associated with the genera Xanthoria and Xanthomendoza (Teloschistaceae, lichenized Ascomycetes)

    PubMed Central

    Nyati, Shyam; Bhattacharya, Debashish; Werth, Silke; Honegger, Rosmarie

    2013-01-01

    We studied group I introns in sterile cultures of selected groups of lichen photobionts, focusing on Trebouxia species associated with Xanthoria s. lat. (including Xanthomendoza spp.; lichen-forming ascomycetes). Group I introns were found inserted after position 798 (Escherichia coli numbering) in the large subunit (LSU) rRNA in representatives of the green algal genera Trebouxia and Asterochloris. The 798 intron was found in about 25% of Xanthoria photobionts including several reference strains obtained from algal culture collections. An alignment of LSU-encoded rDNA intron sequences revealed high similarity of these sequences allowing their phylogenetic analysis. The 798 group I intron phylogeny was largely congruent with a phylogeny of the Internal Transcribed Spacer Region (ITS), indicating that the insertion of the intron most likely occurred in the common ancestor of the genera Trebouxia and Asterochloris. The intron was vertically inherited in some taxa, but lost in others. The high sequence similarity of this intron to one found in Chlorella angustoellipsoidea suggests that the 798 intron was either present in the common ancestor of Trebouxiophyceae, or that its present distribution results from more recent horizontal transfers, followed by vertical inheritance and loss. Analysis of another group I intron shared by these photobionts at small subunit (SSU) position 1512 supports the hypothesis of repeated lateral transfers of this intron among some taxa, but loss among others. Our data confirm that the history of group I introns is characterized by repeated horizontal transfers, and suggests that some of these introns have ancient origins within Chlorophyta. PMID:24415800

  19. PolA1, a Putative DNA Polymerase I, Is Coexpressed with PerR and Contributes to Peroxide Stress Defenses of Group A Streptococcus

    PubMed Central

    Toukoki, Chadia

    2013-01-01

    The peroxide stress response regulator PerR coordinates the oxidative-stress defenses of group A Streptococcus (GAS). We now show that PerR is expressed from an operon encoding a putative DNA polymerase I (PolA1), among other GAS products. A polA1 deletion mutant exhibited wild-type growth but showed reduced capacity to repair DNA damage caused by UV light or ciprofloxacin. Mutant bacteria were hypersensitive to H2O2, compared with the wild type or a complemented mutant strain, and remained severely attenuated even after adaptation at sublethal H2O2 levels, whereas wild-type bacteria could adapt to withstand peroxide challenge under identical conditions. The hypersensitivity of the mutant was reversed when bacteria were grown in iron-depleted medium and challenged in the presence of a hydroxyl radical scavenger, results that indicated sensitivity to hydroxyl radicals generated by Fenton chemistry. The peroxide resistance of a perR polA1 double mutant following adaptation at sublethal H2O2 levels was decreased 9-fold relative to a perR single mutant, thus implicating PolA1 in PerR-mediated defenses against peroxide stress. Cultures of the polA1 mutant grown with or without prior H2O2 exposure yielded considerably lower numbers of rifampin-resistant mutants than cultures of the wild type or the complemented mutant strain, a finding consistent with PolA1 lacking proofreading activity. We conclude that PolA1 promotes genome sequence diversity while playing an essential role in oxidative DNA damage repair mechanisms of GAS, dual functions predicted to confer optimal adaptive capacity and fitness in the host. Together, our studies reveal a unique genetic and functional relationship between PerR and PolA1 in streptococci. PMID:23204468

  20. Synthesis and structure of a new tetracopper(II) complex bridged both by oxamido and phenolato groups: Cytotoxic activity, and reactivity towards DNA and BSA

    NASA Astrophysics Data System (ADS)

    Xu, Xiao-Wen; Li, Xue-Jie; Zhan, Shu-Hui; Li, Yan-Tuan; Wu, Zhi-Yong; Yan, Cui-Wei

    2013-05-01

    A new tetracopper(II) complex bridged both by oxamido and phenolato groups, namely [Cu4(chmpoxd)2(dabt)2](ClO4)2, where H3chmpoxd and dabt stand for N-(5-chloro-2-hydroxyl-phenyl)-N'-[3-(methylamino)propyl]oxamide and 2,2'-diamino-4,4'-bithiazole, respectively, has been synthesized and characterized by elemental analyses, molar conductance measurements, IR and electronic spectra studies, and single-crystal X-ray diffraction. The crystal structure reveals a centrosymmetric circular tetranuclear cation [Cu4(chmpoxd)2(dabt)2]2+ assembled by a pair of cis-oxamido-bridged bicopper(II) units via μ2-phenolato bridges, in which one copper(II) atom is located in a slightly distorted square-planar environment, while the other is in a square-pyramidal geometry. The Cu⋯Cu separations through the oxamido and the phenolato bridges are 5.2217(12) and 3.7042(11) Å, respectively. In vitro cytotoxicity experiment shows that the tetracopper(II) complex exhibits cytotoxic activity against the SMMC7721 and A549 cell lines. The reactivities towards HS-DNA and protein BSA revealed that the tetracopper(II) complex can interact with HS-DNA in the mode of intercalation, and the complex binds to BSA responsible for quenching of tryptophan fluorescence by static quenching mechanism.

  1. Characterization of polybacterial clinical samples using a set of group-specific broad-range primers targeting the 16S rRNA gene followed by DNA sequencing and RipSeq analysis

    PubMed Central

    Lekang, Katrine; Langeland, Nina; Wiker, Harald G.

    2011-01-01

    The standard use of a single universal broad-range PCR in direct 16S rDNA sequencing from polybacterial samples leaves the minor constituents at risk of remaining undetected because all bacterial DNA will be competing for the same reagents. In this article we introduce a set of three broad-range group-specific 16S rDNA PCRs that together cover the clinically relevant bacteria and apply them in the investigation of 25 polybacterial clinical samples. Mixed DNA chromatograms from samples containing more than one species per primer group were analysed using RipSeq Mixed (iSentio, Norway), a web-based application for the interpretation of chromatograms containing up to three different species. The group-specific PCRs reduced complexity in the resulting DNA chromatograms and made the assay more sensitive in situations with unequal species concentrations. Together this allowed for identification of a significantly higher number of bacterial species than did standard direct sequencing with a single universal primer pair and RipSeq analysis (95 vs 51). The method could improve microbiological diagnostics for important groups of patients and can be established in any laboratory with experience in direct 16S rDNA sequencing. PMID:21436365

  2. A study of genetic polymorphisms in mitochondrial DNA hypervariable regions I and II of the five major ethnic groups and Vedda population in Sri Lanka.

    PubMed

    Ranasinghe, Ruwandi; Tennekoon, Kamani H; Karunanayake, Eric H; Lembring, Maria; Allen, Marie

    2015-11-01

    Diversity of the hypervariable regions (HV) I and II of the mitochondrial genome was studied in maternally unrelated Sri Lankans (N=202) from six ethnic groups (i.e.: Sinhalese, Sri Lankan Tamil, Muslim, Malay, Indian Tamil and Vedda). DNA was extracted from blood and buccal swabs and HVI and HVII regions were PCR amplified and sequenced. Resulting sequences were aligned and edited between 16024-16365 and 73-340 regions and compared with revised Cambridge reference sequences (rCRS). One hundred and thirty-five unique haplotypes and 22 shared haplotypes were observed. A total of 145 polymorphic sites and 158 polymorphisms were observed. Hypervariable region I showed a higher polymorphic variation than hypervariable region II. Nucleotide diversities were quite low and similar for all ethnicities apart from a slightly higher value for Indian Tamils and a much lower value for the Vedda population compared to the other groups. When the total population was considered South Asian (Indian) haplogroups were predominant, but there were differences in the distribution of phylo-geographical haplogroups between ethnic groups. Sinhalese, Sri Lankan Tamil and Vedda populations had a considerable presence of West Eurasian haplogroups. About 2/3rd of the Vedda population comprised of macro-haplogroup N or its subclades R and U, whereas macro-haplogroup M was predominant in all other populations. The Vedda population clustered separately from other groups and Sri Lankan Tamils showed a closer genetic affiliation to Sinhalese than to Indian Tamils. Thus this study provides useful information for forensic analysis and anthropological studies of Sri Lankans. PMID:26065620

  3. SOXE transcription factors form selective dimers on non-compact DNA motifs through multifaceted interactions between dimerization and high-mobility group domains

    PubMed Central

    Huang, Yong-Heng; Jankowski, Aleksander; Cheah, Kathryn S. E.; Prabhakar, Shyam; Jauch, Ralf

    2015-01-01

    The SOXE transcription factors SOX8, SOX9 and SOX10 are master regulators of mammalian development directing sex determination, gliogenesis, pancreas specification and neural crest development. We identified a set of palindromic SOX binding sites specifically enriched in regulatory regions of melanoma cells. SOXE proteins homodimerize on these sequences with high cooperativity. In contrast to other transcription factor dimers, which are typically rigidly spaced, SOXE group proteins can bind cooperatively at a wide range of dimer spacings. Using truncated forms of SOXE proteins, we show that a single dimerization (DIM) domain, that precedes the DNA binding high mobility group (HMG) domain, is sufficient for dimer formation, suggesting that DIM : HMG rather than DIM:DIM interactions mediate the dimerization. All SOXE members can also heterodimerize in this fashion, whereas SOXE heterodimers with SOX2, SOX4, SOX6 and SOX18 are not supported. We propose a structural model where SOXE-specific intramolecular DIM:HMG interactions are allosterically communicated to the HMG of juxtaposed molecules. Collectively, SOXE factors evolved a unique mode to combinatorially regulate their target genes that relies on a multifaceted interplay between the HMG and DIM domains. This property potentially extends further the diversity of target genes and cell-specific functions that are regulated by SOXE proteins. PMID:26013289

  4. Genetic variation in DNA-repair pathways and response to radiochemotherapy in esophageal adenocarcinoma: a retrospective cohort study of the Eastern Cooperative Oncology Group

    PubMed Central

    2011-01-01

    Background Recent data in esophageal cancer suggests the variant allele of a single-nucleotide polymorphism (SNP) in XRCC1 may be associated with resistance to radiochemotherapy. However, this SNP has not been assessed in a histologically homogeneous clinical trial cohort that has been treated with a uniform approach. In addition, whether germline DNA may serve as a surrogate for tumor genotype at this locus is unknown in this disease. Our objective was to assess this SNP in relation to the pathologic complete response (pCR) rate in subjects with esophageal adenocarcinoma who received cisplatin-based preoperative radiochemotherapy in a multicenter clinical trial (Eastern Cooperative Oncology Group 1201). As a secondary aim, we investigated the rate of allelic imbalance between germline and tumor DNA. Methods Eighty-one eligible treatment-naïve subjects with newly diagnosed resectable esophageal adenocarcinoma received radiotherapy (45 Gy) concurrent with cisplatin-based chemotherapy, with planned subsequent surgical resection. The primary endpoint was pCR, defined as complete absence of tumor in the surgical specimen after radiochemotherapy. Using germline DNA from 60 subjects, we examined the base-excision repair SNP, XRCC1 Arg399Gln, and 4 other SNPs in nucleotide excision (XPD Lys751Gln and Asp312Asn, ERCC1 3' flank) and double-stranded break (XRCC2 5' flank) repair pathways, and correlated genotype with pCR rate. Paired tumor tissue was used to estimate the frequency of allelic imbalance at the XRCC1 SNP. Results The variant allele of the XRCC1 SNP (399Gln) was detected in 52% of subjects. Only 6% of subjects with the variant allele experienced a pCR, compared to 28% of subjects without the variant allele (odds ratio 5.37 for failing to achieve pCR, p = 0.062). Allelic imbalance at this locus was found in only 10% of informative subjects, suggesting that germline genotype may reflect tumor genotype at this locus. No significant association with pCR was noted

  5. Molecular cloning, sequence, and expression of a human GDP-L-fucose:. beta. -D-galactoside 2-. alpha. -L-fucosyltransferase cDNA that can form the H blood group antigen

    SciTech Connect

    Larsen, R.D.; Ernst, L.K.; Nair, R.P.; Lowe, J.B. )

    1990-09-01

    The authors have previously used a gene-transfer scheme to isolate a human genomic DNA fragment that determines expression of a GDP-L-fucose:{beta}D-galactoside 2-{alpha}-L-fucosyltransferase. Although this fragment determined expression of an {alpha}(1,2)FT whose kinetic properties mirror those of the human H blood group {alpha}(1,2)FT, their precise nature remained undefined. They describe here the molecular cloning, sequence, and expression of a human of cDNA corresponding to these human genomic sequences. When expressed in COS-1 cells, the cDNA directs expression of cell surface H structures and a cognate {alpha}(1,2)FT activity with properties analogous to the human H blood group {alpha}(1,2)FT. The cDNA sequence predicts a 365-amino acid polypeptide characteristic of a type II transmembrane glycoprotein with a domain structure analogous to that of other glycosyltransferases but without significant primary sequence similarity to these or other known proteins. To directly demonstrate that the cDNA encodes an {alpha}(1,2)FT, the COOH-terminal domain predicted to be Golgi-resident was expressed in COS-1 cells as a catalytically active, secreted, and soluble protein A fusion peptide. Southern blot analysis showed that this cDNA identified DNA sequences syntenic to the human H locus on chromosome 19. These results strongly suggest that this cloned {alpha}(1,2)FT cDNA represents the product of the human H blood group locus.

  6. Virion-associated cofactor high-mobility group DNA-binding protein-1 facilitates transposition from the herpes simplex virus/Sleeping Beauty amplicon vector platform.

    PubMed

    de Silva, Suresh; Lotta, Louis T; Burris, Clark A; Bowers, William J

    2010-11-01

    The development of the integration-competent, herpes simplex virus/Sleeping Beauty (HSV/SB) amplicon vector platform has created a means to efficiently and stably deliver therapeutic transcription units (termed "transgenons") to neurons within the mammalian brain. Furthermore, an investigation into the transposition capacity of the HSV/SB vector system revealed that the amplicon genome provides an optimal substrate for the transposition of transgenons at least 12 kb in length [de Silva, S., Mastrangelo, M.A., Lotta, L.T., Jr., Burris, C.A., Federoff, H.J., and Bowers, W.J. ( 2010 ). Gene Ther. 17, 424-431]. These results prompted an investigation into the factors that may contribute toward efficient transposition from the HSV/SB amplicon. One of the cellular cofactors known to play a key role during SB-mediated transposition is the high-mobility group DNA-binding protein-1 (HMGB1). Our present investigation into the role of HMGB1 during amplicon-based transposition revealed that transposition is not strictly dependent on the presence of cellular HMGB1, contrary to what had been previously demonstrated with plasmid-based SB transposition. We have shown for the first time that during amplicon preparation, biologically active HMGB1 derived from the packaging cell line is copackaged into amplicon vector particles. As a result, HSV/SB amplicon virions arrive prearmed with HMGB1 protein at levels sufficient for facilitating SB-mediated transposition in the transduced mammalian cell. PMID:20568967

  7. Mucosal immunization with high-mobility group box 1 in chitosan enhances DNA vaccine-induced protection against coxsackievirus B3-induced myocarditis.

    PubMed

    Wang, Maowei; Yue, Yan; Dong, Chunsheng; Li, Xiaoyun; Xu, Wei; Xiong, Sidong

    2013-11-01

    Coxsackievirus B3 (CVB3), a small single-stranded RNA virus, belongs to the Picornaviridae family. Its infection is the most common cause of myocarditis, with no vaccine available. Gastrointestinal mucosa is the major entry port for CVB3; therefore, the induction of local immunity in mucosal tissues may help control initial viral infections and alleviate subsequent myocardial injury. Here we evaluated the ability of high-mobility group box 1 (HMGB1) encapsulated in chitosan particles to enhance the mucosal immune responses induced by the CVB3-specific mucosal DNA vaccine chitosan-pVP1. Mice were intranasally coimmunized with 4 doses of chitosan-pHMGB1 and chitosan-pVP1 plasmids, at 2-week intervals, and were challenged with CVB3 4 weeks after the last immunization. Compared with chitosan-pVP1 immunization alone, coimmunization with chitosan-pHMGB1 significantly (P < 0.05) enhanced CVB3-specific fecal secretory IgA levels and promoted mucosal T cell immune responses. In accordance, reduced severity of myocarditis was observed in coimmunized mice, as evidenced by significantly (P < 0.05) reduced viral loads, decreased myocardial injury, and increased survival rates. Flow cytometric analysis indicated that HMGB1 enhanced dendritic cell (DC) recruitment to mesenteric lymph nodes and promoted DC maturation, which might partly account for its mucosal adjuvant effect. This strategy may represent a promising approach to candidate vaccines against CVB3-induced myocarditis. PMID:24027262

  8. Variations of SSU rDNA group I introns in different isolates of Cordyceps militaris and the loss of an intron during cross-mating.

    PubMed

    Lian, Tiantian; Yang, Tao; Sun, Junde; Guo, Suping; Yang, Huaijun; Dong, Caihong

    2014-08-01

    Cordyceps militaris, the type species of genus Cordyceps, is one of the most popular mushrooms and a nutraceutical in eastern Asia. It is considered a model organism for the study of Cordyceps species because it can complete its life cycle when cultured in vitro. In the present study, the occurrence and sequence variation of SSU rDNA group I introns, Cmi.S943 and Cmi.S1199, among different isolates of C. militaris were analyzed. Based on the secondary structure predictions, the Cmi.S943 intron has been placed in subgroup IC1, and the Cmi.S1199 intron has been placed in subgroup IE. No significant similarity between Cmi.S943 and Cmi.S1199 suggested different origins. Three genotypes, based on the frequency and distribution of introns, were described to discriminate the 57 surveyed C. militaris strains. It was found that the genotype was related to the stroma characteristics. The stromata of all of the genotype II strains, which possessed only Cmi.S943, could produce perithecium. In contrast, the stromata of all genotype III strains, which had both Cmi.S943 and Cmi.S1199, could not produce perithecium. Cmi.S1199 showed the lowest level of intra-specific variation among the tested strains. Group I introns can be lost during strain cross-mating. Therefore, we presumed that during cross-mating and recombination, intron loss could be driven by positive Darwinian selection due to the energetic cost of transcribing long introns. PMID:24996897

  9. Sperm DNA oxidative damage and DNA adducts.

    PubMed

    Jeng, Hueiwang Anna; Pan, Chih-Hong; Chao, Mu-Rong; Lin, Wen-Yi

    2015-12-01

    The objective of this study was to investigate DNA damage and adducts in sperm from coke oven workers who have been exposed to polycyclic aromatic hydrocarbons. A longitudinal study was conducted with repeated measurements during spermatogenesis. Coke-oven workers (n=112) from a coke-oven plant served the PAH-exposed group, while administrators and security personnel (n=67) served the control. Routine semen parameters (concentration, motility, vitality, and morphology) were analyzed simultaneously; the assessment of sperm DNA integrity endpoints included DNA fragmentation, bulky DNA adducts, and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dGuo). The degree of sperm DNA fragmentation was measured using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay and sperm chromatin structure assay (SCSA). The PAH-exposed group had a significant increase in bulky DNA adducts and 8-oxo-dGuo compared to the control subjects (Ps=0.002 and 0.045, respectively). Coke oven workers' percentages of DNA fragmentation and denaturation from the PAH-exposed group were not significantly different from those of the control subjects (Ps=0.232 and 0.245, respectively). Routine semen parameters and DNA integrity endpoints were not correlated. Concentrations of 8-oxo-dGuo were positively correlated with percentages of DNA fragmentation measured by both TUNEL and SCSA (Ps=0.045 and 0.034, respectively). However, the concentrations of 8-oxo-dGuo and percentages of DNA fragmentation did not correlate with concentrations of bulky DNA adducts. In summary, coke oven workers with chronic exposure to PAHs experienced decreased sperm DNA integrity. Oxidative stress could contribute to the degree of DNA fragmentation. Bulky DNA adducts may be independent of the formation of DNA fragmentation and oxidative adducts in sperm. Monitoring sperm DNA integrity is recommended as a part of the process of assessing the impact of occupational and environmental toxins on sperm

  10. Phosphorus-nitrogen compounds: Part 28. Syntheses, structural characterizations, antimicrobial and cytotoxic activities, and DNA interactions of new phosphazenes bearing vanillinato and pendant ferrocenyl groups

    NASA Astrophysics Data System (ADS)

    Tümer, Yasemin; Asmafiliz, Nuran; Kılıç, Zeynel; Hökelek, Tuncer; Yasemin Koç, L.; Açık, Leyla; Yola, Mehmet Lütfi; Solak, Ali Osman; Öner, Yağmur; Dündar, Devrim; Yavuz, Makbule

    2013-10-01

    The gradually Cl replacement reactions of spirocyclic mono (1 and 2) and bisferrocenyl cyclotriphosphazenes (3-5) with the potassium salt of 4-hydroxy-3-methoxybenzaldehyde (potassium vanillinate) gave mono (1a-5a), geminal (gem-1b-5b), non-geminal (cis-1b, cis-5b and trans-2b-5b), tri (1c-5c) and tetra-substituted phosphazenes (1d-5d). Some phosphazenes have stereogenic P-center(s). The chirality of 4c was verified using chiral HPLC column. Electrochemical behaviors were influenced only by the number of ferrocene groups, but not the length of the amine chains and the substituent(s). The structures of the new phosphazenes were determined by FTIR, MS, 1H, 13C and 31P NMR, HSQC and HMBC spectral data. The solid-state structures of cis-1b and 4d were examined by single crystal X-ray diffraction techniques. The twelve phosphazene derivatives were screened for antimicrobial activity and the compounds 5a, cis-1b and 2c exhibited the highest antibacterial activity against G(+) and G(-) bacteria. In addition, it was found that overall gem-1b inhibited the growth of Mycobacterium tuberculosis. The compounds 1d, 2d and 4d were tested in HeLa cancer cell lines. Among these compounds, 2d had cytotoxic effect on HeLa cell in the first 48 h. Moreover, interactions between compounds 2a, gem-1b, gem-2b, cis-1b, 2c, 3c, 4c, 5c, 1d, 2d and 4d, and pBR322 plasmid DNA were investigated.

  11. A 127 kDa component of a UV-damaged DNA-binding complex, which is defective in some xeroderma pigmentosum group E patients, is homologous to a slime mold protein.

    PubMed Central

    Takao, M; Abramic, M; Moos, M; Otrin, V R; Wootton, J C; McLenigan, M; Levine, A S; Protic, M

    1993-01-01

    A cDNA which encodes a approximately 127 kDa UV-damaged DNA-binding (UV-DDB) protein with high affinity for (6-4)pyrimidine dimers [Abramic', M., Levine, A.S. & Protic', M., J. Biol. Chem. 266: 22493-22500, 1991] has been isolated from a monkey cell cDNA library. The presence of this protein in complexes bound to UV-damaged DNA was confirmed by immunoblotting. The human cognate of the UV-DDB gene was localized to chromosome 11. UV-DDB mRNA was expressed in all human tissues examined, including cells from two patients with xeroderma pigmentosum (group E) that are deficient in UV-DDB activity, which suggests that the binding defect in these cells may reside in a dysfunctional UV-DDB protein. Database searches have revealed significant homology of the UV-DDB protein sequence with partial sequences of yet uncharacterized proteins from Dictyostelium discoideum (44% identity over 529 amino acids) and Oryza sativa (54% identity over 74 residues). According to our results, the UV-DDB polypeptide belongs to a highly conserved, structurally novel family of proteins that may be involved in the early steps of the UV response, e.g., DNA damage recognition. Images PMID:8371985

  12. Evolution of eukaryotic single-stranded DNA viruses of the Bidnaviridae family from genes of four other groups of widely different viruses

    NASA Astrophysics Data System (ADS)

    Krupovic, Mart; Koonin, Eugene V.

    2014-06-01

    Single-stranded (ss)DNA viruses are extremely widespread, infect diverse hosts from all three domains of life and include important pathogens. Most ssDNA viruses possess small genomes that replicate by the rolling-circle-like mechanism initiated by a distinct virus-encoded endonuclease. However, viruses of the family Bidnaviridae, instead of the endonuclease, encode a protein-primed type B DNA polymerase (PolB) and hence break this pattern. We investigated the provenance of all bidnavirus genes and uncover an unexpected turbulent evolutionary history of these unique viruses. Our analysis strongly suggests that bidnaviruses evolved from a parvovirus ancestor from which they inherit a jelly-roll capsid protein and a superfamily 3 helicase. The radiation of bidnaviruses from parvoviruses was probably triggered by integration of the ancestral parvovirus genome into a large virus-derived DNA transposon of the Polinton (polintovirus) family resulting in the acquisition of the polintovirus PolB gene along with terminal inverted repeats. Bidnavirus genes for a receptor-binding protein and a potential novel antiviral defense modulator are derived from dsRNA viruses (Reoviridae) and dsDNA viruses (Baculoviridae), respectively. The unusual evolutionary history of bidnaviruses emphasizes the key role of horizontal gene transfer, sometimes between viruses with completely different genomes but occupying the same niche, in the emergence of new viral types.

  13. Evolution of eukaryotic single-stranded DNA viruses of the Bidnaviridae family from genes of four other groups of widely different viruses

    PubMed Central

    Krupovic, Mart; Koonin, Eugene V.

    2014-01-01

    Single-stranded (ss)DNA viruses are extremely widespread, infect diverse hosts from all three domains of life and include important pathogens. Most ssDNA viruses possess small genomes that replicate by the rolling-circle-like mechanism initiated by a distinct virus-encoded endonuclease. However, viruses of the family Bidnaviridae, instead of the endonuclease, encode a protein-primed type B DNA polymerase (PolB) and hence break this pattern. We investigated the provenance of all bidnavirus genes and uncover an unexpected turbulent evolutionary history of these unique viruses. Our analysis strongly suggests that bidnaviruses evolved from a parvovirus ancestor from which they inherit a jelly-roll capsid protein and a superfamily 3 helicase. The radiation of bidnaviruses from parvoviruses was probably triggered by integration of the ancestral parvovirus genome into a large virus-derived DNA transposon of the Polinton (polintovirus) family resulting in the acquisition of the polintovirus PolB gene along with terminal inverted repeats. Bidnavirus genes for a receptor-binding protein and a potential novel antiviral defense modulator are derived from dsRNA viruses (Reoviridae) and dsDNA viruses (Baculoviridae), respectively. The unusual evolutionary history of bidnaviruses emphasizes the key role of horizontal gene transfer, sometimes between viruses with completely different genomes but occupying the same niche, in the emergence of new viral types. PMID:24939392

  14. Changes in Fat Mitochondrial DNA and Function in Subjects Randomized to Abacavir-Lamivudine or Tenofovir DF–Emtricitabine With Atazanavir-Ritonavir or Efavirenz: AIDS Clinical Trials Group Study A5224s, Substudy of A5202

    PubMed Central

    McComsey, Grace A.; Daar, Eric S.; O'Riordan, MaryAnn; Collier, Ann C.; Kosmiski, Lisa; Santana, Jorge L.; Fichtenbaum, Carl J.; Fink, Heidi; Sax, Paul E.; Libutti, Daniel E.; Gerschenson, Mariana

    2013-01-01

    Background. The effect of nonthymidine nucleoside reverse-transcriptase inhibitors (NRTIs) on fat mitochondrial DNA (mtDNA) content and function is unclear. Methods. A5202 randomized antiretroviral therapy–naive human immunodeficiency virus–infected subjects to abacavir-lamivudine (ABC/3TC) versus tenofovir DF–emtricitabine (TDF/FTC) with efavirenz (EFV) or atazanavir-ritonavir (ATV/r). A5224s, substudy of A5202, enrolled 269 subjects with fat measurements by dual-energy x-ray absorptiometry and computed tomography. A subset of subjects underwent fat biopsies at baseline and week 96 for mtDNA content (real-time polymerase chain reaction) and oxidative phosphorylation nicotinamide adenine dinucleotide (reduced) dehydrogenase (complex I) and cytochrome c oxidase (complex IV) activity levels (immunoassays). Intent-to-treat analyses were performed using analysis of variance and paired t tests. Results. Fifty-six subjects (87% male; median age, 39 years) were included; their median body mass index, CD4 cell count, and fat mtDNA level were 26 kg/m2, 227 cells/μL, and 1197 copies/cell, respectively. Fat mtDNA content decreased within the ABC/3TC and TDF/FTC groups (combining EFV and ATV/r arms; median change, −341 [interquartile range, −848 to 190; P = .03] and −400 [−661 to −221; P < .001] copies/cell, respectively), but these changes did not differ significantly between the 2 groups (P = .57). Complex I and IV activity decreased significantly in the TDF/FTC group (median change, −12.45 [interquartile range, −24.70 to 2.90; P = .003] and −8.25 [−13.90 to −1.30; P < .001], optical density × 103/µg, respectively) but not the ABC/3TC group. Differences between the ABC/3TC and TDF/FTC groups were significant for complex I (P = .03). Conclusions. ABC/3TC and TDF/FTC significantly and similarly decreased fat mtDNA content, but only TDF/FTC decreased complex I and complex IV activity levels. Clinical Trials Registration. NCT00118898. PMID

  15. Reactions of 5-methylcytosine cation radicals in DNA and model systems: thermal deprotonation from the 5-methyl group vs. excited state deprotonation from sugar

    PubMed Central

    Adhikary, Amitava; Kumar, Anil; Palmer, Brian J.; Todd, Andrew D.; Heizer, Alicia N.; Sevilla, Michael D.

    2014-01-01

    Purpose To study the formation and subsequent reactions of the 5-methyl-2′-deoxycytidine cation radical (5-Me-2′-dC•+) in nucleosides and DNA-oligomers and compare to one electron oxidized thymidine. Materials and methods Employing electron spin resonance (ESR), cation radical formation and its reactions were investigated in 5-Me-2′-dC, thymidine (Thd) and their derivatives, in fully double stranded (ds) d[GC*GC*GC*GC*]2 and in the 5-Me-C/A mismatched, d[GGAC*AAGC:CCTAATCG], where C* = 5-Me-C. Results We report 5-Me-2′-dC•+ production by one-electron oxidation of 5-Me-2′-dC by Cl2•− via annealing in the dark at 155 K. Progressive annealing of 5-Me-2′-dC•+ at 155 K produces the allylic radical (C-CH2•). However, photoexcitation of 5-Me-2′-dC•+ by 405 nm laser or by photoflood lamp leads to only C3′• formation. Photoexcitation of N3-deprotonated thyminyl radical in Thd and its 5′-nucleotides leads to C3′• formation but not in 3′-TMP which resulted in the allylic radical (U-CH2•) and C5′• production. For excited 5-Me-2′,3′-ddC•+, absence of the 3′-OH group does not prevent C3′• formation. For d[GC*GC*GC*GC*]2 and d[GGAC*AAGC:CCTAATCG], intra-base paired proton transferred form of G cation radical (G(N1-H)•:C(+H+)) is found with no observable 5-Me-2′-dC•+ formation. Photoexcitation of (G(N1-H)•:C(+H+)) in d[GC*GC*GC*GC*]2 produced only C1′• and not the expected photoproducts from 5-Me-2′-dC•+. However, photoexcitation of (G(N1-H)•:C(+H+)) in d[GGAC*AAGC:CCTAATCG] led to C5′• and C1′• formation. Conclusions C-CH2• formation from 5-Me-2′-dC•+ occurs via ground state deprotonation from C5-methyl group on the base. In the excited 5-Me-2′-dC•+ and 5-Me-2′,3′-ddC•+, spin and charge localization at C3′ followed by deprotonation leads to C3′• formation. Thus, deprotonation from C3′ in the excited cation radical is kinetically controlled and sugar C-H bond energies are

  16. Functional regulation of the DNA damage-recognition factor DDB2 by ubiquitination and interaction with xeroderma pigmentosum group C protein

    PubMed Central

    Matsumoto, Syota; Fischer, Eric S.; Yasuda, Takeshi; Dohmae, Naoshi; Iwai, Shigenori; Mori, Toshio; Nishi, Ryotaro; Yoshino, Ken-ichi; Sakai, Wataru; Hanaoka, Fumio; Thomä, Nicolas H.; Sugasawa, Kaoru

    2015-01-01

    In mammalian nucleotide excision repair, the DDB1–DDB2 complex recognizes UV-induced DNA photolesions and facilitates recruitment of the XPC complex. Upon binding to damaged DNA, the Cullin 4 ubiquitin ligase associated with DDB1–DDB2 is activated and ubiquitinates DDB2 and XPC. The structurally disordered N-terminal tail of DDB2 contains seven lysines identified as major sites for ubiquitination that target the protein for proteasomal degradation; however, the precise biological functions of these modifications remained unknown. By exogenous expression of mutant DDB2 proteins in normal human fibroblasts, here we show that the N-terminal tail of DDB2 is involved in regulation of cellular responses to UV. By striking contrast with behaviors of exogenous DDB2, the endogenous DDB2 protein was stabilized even after UV irradiation as a function of the XPC expression level. Furthermore, XPC competitively suppressed ubiquitination of DDB2 in vitro, and this effect was significantly promoted by centrin-2, which augments the DNA damage-recognition activity of XPC. Based on these findings, we propose that in cells exposed to UV, DDB2 is protected by XPC from ubiquitination and degradation in a stochastic manner; thus XPC allows DDB2 to initiate multiple rounds of repair events, thereby contributing to the persistence of cellular DNA repair capacity. PMID:25628365

  17. The effects of linear assembly of two carbazole groups on acid-base and DNA-binding properties of a ruthenium(II) complex

    NASA Astrophysics Data System (ADS)

    Chen, Xi; Xue, Long-Xin; Ju, Chun-Chuan; Wang, Ke-Zhi

    2013-07-01

    A novel Ru(II) complex of [Ru(bpy)2(Hbcpip)](ClO4)2 {where bpy = 2,2-bipyridine, Hbcpip = 2-(4-(9H-3,9'-bicarbazol-9-yl)phenyl)-1H-imidazo[4,5-f][1,10]phenanthroline} is synthesized and characterized. Calf-thymus DNA-binding properties of the complex were studied by UV-vis absorption and luminescence titrations, steady-state emission quenching by [Fe(CN)6]4-, DNA competitive binding with ethidium bromide, thermal denaturation and DNA viscosity measurements. The results indicate that the complex partially intercalated into the DNA with a binding constant of (5.5 ± 1.4) × 105 M-1 in buffered 50 mM NaCl. The acid-base properties of the complex were also studied by UV-visible and luminescence spectrophotometric pH titrations, and ground- and excited-state acidity ionization constant values were derived.

  18. A biosensing of Toxoplasma gondii DNA with CdTe/Fe3O4 dual functional quantum dot as reporter group

    NASA Astrophysics Data System (ADS)

    Liang, Chu; Xu, Shichao; Yang, Juan; Zhang, Jimei; Dai, Zhao; Sun, Bo; Sun, Shuqing; Feng, Tielin; Zi, Yan; Liu, Jingwei; Luo, Hao

    2009-07-01

    Toxoplasma gondii is an intestinal coccidium that parasitizes members of the cat family as definitive hosts and has a wide range of intermediate hosts. Infection is common in many warm-blooded animals, including humans, the early detection of Toxoplasma gondii was concerned in recent years. In the current research, we presented a fast, specific, and sensitive sensing probe to detect Toxoplasma gondii DNA based on mechanism of fluorescence energy transfer (FRET), and a magnetic-fluorescent CdTe/Fe3O4 core-shell quantum dots (mQDs) was utilized as energy donor, and a commercial quencher (BHQ-2) was used as energy acceptor, respectively. The CdTe/Fe3O4 mQDs were prepared by layer-by-layer (LBL) process at ambient temperature. The sensing probe was fabricated through labeling a stem-loop Toxoplasma gondii DNA oligonucleotide with mQDs at the 5' end and BHQ-2 at 3' end, respectively, and the resulting sensing probe can be simply isolated and purified from the reactant with a common magnet. Properties of mQDs and sensing probe were determined by transmission electron microscopy (TEM) and fluorescence spectrum (FS). The TEM data demonstrated that the size of mQDs was ~20nm. the FS data indicated fluorescence intensity (FI) was doubled after the complete complimentary target Toxoplasma gondii DNA was introduced comparing with the FI before addition of target Toxoplasma gondii DNA. Moreover, only weak FI change was observed when the target DNA with one-mismatch base pair was added, this result revealed the sensing probe has high sensitivity and specificity. The current sensing probe will has great potential applications in the life science and related research.

  19. DNA Nanotechnology-- Architectures Designed with DNA

    NASA Astrophysics Data System (ADS)

    Han, Dongran

    As the genetic information storage vehicle, deoxyribonucleic acid (DNA) molecules are essential to all known living organisms and many viruses. It is amazing that such a large amount of information about how life develops can be stored in these tiny molecules. Countless scientists, especially some biologists, are trying to decipher the genetic information stored in these captivating molecules. Meanwhile, another group of researchers, nanotechnologists in particular, have discovered that the unique and concise structural features of DNA together with its information coding ability can be utilized for nano-construction efforts. This idea culminated in the birth of the field of DNA nanotechnology which is the main topic of this dissertation. The ability of rationally designed DNA strands to self-assemble into arbitrary nanostructures without external direction is the basis of this field. A series of novel design principles for DNA nanotechnology are presented here, from topological DNA nanostructures to complex and curved DNA nanostructures, from pure DNA nanostructures to hybrid RNA/DNA nanostructures. As one of the most important and pioneering fields in controlling the assembly of materials (both DNA and other materials) at the nanoscale, DNA nanotechnology is developing at a dramatic speed and as more and more construction approaches are invented, exciting advances will emerge in ways that we may or may not predict.

  20. Mitochondrial DNA variation within and among regional populations of longtail macaques (Macaca fascicularis) in relation to other species of the fascicularis group of macaques.

    PubMed

    Smith, David Glenn; McDonough, John W; George, Debra A

    2007-02-01

    An 835 base pair (bp) fragment of mitochondrial DNA (mtDNA) was sequenced to characterize genetic variation within and among 1,053 samples comprising five regional populations each of longtail macaques (Macaca fascicularis) and rhesus macaques (Macaca mulatta), and one sample each of Japanese (M. fuscata) and Taiwanese (M. cyclopis) macaques. The mtDNA haplotypes of longtail macaques clustered in two large highly structured clades (Fas1 and Fas2) of a neighbor-joining tree that were reciprocally monophyletic with respect to those representing rhesus macaques, Japanese macaques, and Taiwanese macaques. Both clades exhibited haplotypes of Indonesian and Malaysian longtail macaques widely dispersed throughout them; however, longtail macaques from Indochina, Philippines, and Mauritius each clustered in a separate well-defined clade together with one or a few Malaysian and/or Indonesian longtail macaques, suggesting origins on the Sunda shelf. Longtail macaques from Malaysia and Indonesia were far more genetically diverse, and those from Mauritius were far less diverse than any other population studied. Nucleotide diversity between mtDNA sequences of longtail macaques from different geographic regions is, in some cases, greater than that between Indian and Chinese rhesus macaques. Approximately equal amounts of genetic diversity are due to differences among animals in the same regional population, different regional populations, and different species. A greater proportion of genetic variance was explained by interspecies differences when Japanese and Taiwanese macaques were regarded as regional populations of rhesus macaques than when they were treated as separate species. Rhesus macaques from China were more closely related to both Taiwanese and Japanese macaques than to their own conspecifics from India. PMID:17177314

  1. DNA codes

    SciTech Connect

    Torney, D. C.

    2001-01-01

    We have begun to characterize a variety of codes, motivated by potential implementation as (quaternary) DNA n-sequences, with letters denoted A, C The first codes we studied are the most reminiscent of conventional group codes. For these codes, Hamming similarity was generalized so that the score for matched letters takes more than one value, depending upon which letters are matched [2]. These codes consist of n-sequences satisfying an upper bound on the similarities, summed over the letter positions, of distinct codewords. We chose similarity 2 for matches of letters A and T and 3 for matches of the letters C and G, providing a rough approximation to double-strand bond energies in DNA. An inherent novelty of DNA codes is 'reverse complementation'. The latter may be defined, as follows, not only for alphabets of size four, but, more generally, for any even-size alphabet. All that is required is a matching of the letters of the alphabet: a partition into pairs. Then, the reverse complement of a codeword is obtained by reversing the order of its letters and replacing each letter by its match. For DNA, the matching is AT/CG because these are the Watson-Crick bonding pairs. Reversal arises because two DNA sequences form a double strand with opposite relative orientations. Thus, as will be described in detail, because in vitro decoding involves the formation of double-stranded DNA from two codewords, it is reasonable to assume - for universal applicability - that the reverse complement of any codeword is also a codeword. In particular, self-reverse complementary codewords are expressly forbidden in reverse-complement codes. Thus, an appropriate distance between all pairs of codewords must, when large, effectively prohibit binding between the respective codewords: to form a double strand. Only reverse-complement pairs of codewords should be able to bind. For most applications, a DNA code is to be bi-partitioned, such that the reverse-complementary pairs are separated

  2. The katablepharids are a distant sister group of the Cryptophyta: A proposal for Katablepharidophyta divisio nova/ Kathablepharida phylum novum based on SSU rDNA and beta-tubulin phylogeny.

    PubMed

    Okamoto, Noriko; Inouye, Isao

    2005-08-01

    The katablepharids are a morphologically well-defined group of heterotrophic flagellates. Since their original description in 1939, they have been classified in the Cryptophyceae (Cryptophyta) based on their similar cell shape, flagellar orientation, and the presence of ejectisomes visible by light microscopy. However, electron microscopy suggests that the katablepharids are distinct from cryptomonads. A possible affinity with the Alveolata has been proposed which is mainly based on the resemblance of their feeding apparatus to the apical complex of the Apicomplexa or to the tentacles of the Ciliophora. In this study, we provide the first SSU rDNA and beta-tubulin molecular sequence data for two katablepharids: Katablepharis japonica sp. nov. and Leucocryptos marina. We reveal that the katablepharids are not closely related to the Alveolata; rather, phylogenetic reconstruction analyses of SSU rDNA and beta-tubulin suggest that the katablepharids are a distant sister group of the Cryptophyta. We therefore conclude that the katablepharids should be a group equivalent to the Cryptophyta and propose Katablepharidophyta divisio nova (ICBN)/Kathablepharida phylum novum (ICZN). PMID:16171184

  3. Cleaving DNA with DNA

    NASA Astrophysics Data System (ADS)

    Carmi, Nir; Balkhi, Shameelah R.; Breaker, Ronald R.

    1998-03-01

    A DNA structure is described that can cleave single-stranded DNA oligonucleotides in the presence of ionic copper. This ``deoxyribozyme'' can self-cleave or can operate as a bimolecular complex that simultaneously makes use of duplex and triplex interactions to bind and cleave separate DNA substrates. Bimolecular deoxyribozyme-mediated strand scission proceeds with a kobs of 0.2 min-1, whereas the corresponding uncatalyzed reaction could not be detected. The duplex and triplex recognition domains can be altered, making possible the targeted cleavage of single-stranded DNAs with different nucleotide sequences. Several small synthetic DNAs were made to function as simple ``restriction enzymes'' for the site-specific cleavage of single-stranded DNA.

  4. DNA-cell conjugates

    DOEpatents

    Hsiao, Shih-Chia; Francis, Matthew B.; Bertozzi, Carolyn; Mathies, Richard; Chandra, Ravi; Douglas, Erik; Twite, Amy; Toriello, Nicholas; Onoe, Hiroaki

    2016-05-03

    The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

  5. Immobilization of DNA in polyacrylamide gel for the manufacture of DNA and DNA-oligonucleotide microchips.

    SciTech Connect

    Proudnikov, D.; Timofeev, E.; Mirzabekov, A.; Center for Mechanistic Biology and Biotechnology; Engelhardt Inst. of Molecular Biology

    1998-05-15

    Activated DNA was immobilized in aldehyde-containing polyacrylamide gel for use in manufacturing the MAGIChip (microarrays of gel-immobilized compounds on a chip). First, abasic sites were generated in DNA by partial acidic depurination. Amino groups were then introduced into the abasic sites by reaction with ethylenediamine and reduction of the aldimine bonds formed. It was found that DNA could be fragmented at the site of amino group incorporation or preserved mostly unfragmented. In similar reactions, both amino-DNA and amino-oligonucleotides were attached through their amines to polyacrylamide gel derivatized with aldehyde groups. Single- and double-stranded DNA of 40 to 972 nucleotides or base pairs were immobilized on the gel pads to manufacture a DNA microchip. The microchip was hybridized with fluorescently labeled DNA-specific oligonucleotide probes. This procedure for immobilization of amino compounds was used to manufacture MAGIChips containing both DNA and oligonucleotides.

  6. DNA repair

    SciTech Connect

    Friedberg, E.C.; Hanawalt, P.C. )

    1988-01-01

    Topics covered in this book included: Eukaryote model systems for DNA repair study; Sensitive detection of DNA lesions and their repair; and Defined DNA sequence probes for analysis of mutagenesis and repair.

  7. Partial mitochondrial DNA sequences suggest the existence of a cryptic species within the Leucosphyrus group of the genus Anopheles (Diptera: Culicidae), forest malaria vectors, in northern Vietnam

    PubMed Central

    2010-01-01

    Background During the last decade, Southeast Asian countries have been very successful in reducing the burden of malaria. However, malaria remains endemic in these countries, especially in remote and forested areas. The Leucosphyrus group of the genus Anopheles harbors the most important malaria vectors in forested areas of Southeast Asia. In Vietnam, previous molecular studies have resulted in the identification of only Anopheles dirus sensu stricto (previously known as An. dirus species A) among the Leucosphyrus group members. However, Vietnamese entomologists have recognized that mosquitoes belonging to the Leucosphyrus group in northern Vietnam exhibit morphological characteristics similar to those of Anopheles takasagoensis, which has been reported only from Taiwan. Here, we aimed to confirm the genetic and morphological identities of the members of the Leucosphyrus group in Vietnam. Results In the molecular phylogenetic trees reconstructed using partial COI and ND6 mitochondrial gene sequences, samples collected from southern and central Vietnam clustered together with GenBank sequences of An. dirus that were obtained from Thailand. However, samples from northern Vietnam formed a distinct clade separated from both An. dirus and An. takasagoensis by other valid species. Conclusions The results suggest the existence of a cryptic species in northern Vietnam that is morphologically similar to, but phylogenetically distant from both An. dirus and An. takasagoensis. We have tentatively designated this possible cryptic species as Anopheles aff. takasagoensis for convenience, until a valid name is assigned. However, it is difficult to distinguish the species solely on the basis of morphological characteristics. Further studies on such as karyotypes and polytene chromosome banding patterns are necessary to confirm whether An. aff. takasagoensis is a valid species. Moreover, studies on (1) the geographic distribution, which is potentially spreading along the Vietnam

  8. PEX12, the Pathogenic Gene of Group III Zellweger Syndrome: cDNA Cloning by Functional Complementation on a CHO Cell Mutant, Patient Analysis, and Characterization of Pex12p

    PubMed Central

    Okumoto, Kanji; Shimozawa, Nobuyuki; Kawai, Atsusi; Tamura, Shigehiko; Tsukamoto, Toshiro; Osumi, Takashi; Moser, Hugo; Wanders, Ronald J. A.; Suzuki, Yasuyuki; Kondo, Naomi; Fujiki, Yukio

    1998-01-01

    Rat PEX12 cDNA was isolated by functional complementation of peroxisome deficiency of a mutant CHO cell line, ZP109 (K. Okumoto, A. Bogaki, K. Tateishi, T. Tsukamoto, T. Osumi, N. Shimozawa, Y. Suzuki, T. Orii, and Y. Fujiki, Exp. Cell Res. 233:11–20, 1997), using a transient transfection assay and an ectopic, readily visible marker, green fluorescent protein. This cDNA encodes a 359-amino-acid membrane protein of peroxisomes with two transmembrane segments and a cysteine-rich zinc finger, the RING motif. A stable transformant of ZP109 with the PEX12 was morphologically and biochemically restored for peroxisome biogenesis. Pex12p was shown by expression of bona fide as well as epitope-tagged Pex12p to expose both N- and C-terminal regions to the cytosol. Fibroblasts derived from patients with the peroxisome deficiency Zellweger syndrome of complementation group III (CG-III) were also complemented for peroxisome biogenesis with PEX12. Two unrelated patients of this group manifesting peroxisome deficiency disorders possessed homozygous, inactivating PEX12 mutations: in one, Arg180Thr by one point mutation, and in the other, deletion of two nucleotides in codons for 291Asn and 292Ser, creating an apparently unchanged codon for Asn and a codon 292 for termination. These results indicate that the gene encoding peroxisome assembly factor Pex12p is a pathogenic gene of CG-III peroxisome deficiency. Moreover, truncation and site mutation studies, including patient PEX12 analysis, demonstrated that the cytoplasmically oriented N- and C-terminal parts of Pex12p are essential for biological function. PMID:9632816

  9. Wrinkled DNA.

    PubMed Central

    Arnott, S; Chandrasekaran, R; Puigjaner, L C; Walker, J K; Hall, I H; Birdsall, D L; Ratliff, R L

    1983-01-01

    The B form of poly d(GC):poly d(GC) in orthorhombic microcrystallites in oriented fibers has a secondary structure in which a dinucleotide is the repeated motif rather than a mononucleotide as in standard, smooth B DNA. One set of nucleotides (probably GpC) has the same conformations as the smooth form but the alternate (CpG) nucleotides have a different conformation at C3'-O3'. This leads to a distinctive change in the orientation of the phosphate groups. Similar perturbations can be detected in other poly d(PuPy):poly d(PuPy) DNAs such as poly d(IC):poly d(IC) and poly d(AT):poly d(AT) in their D forms which have tetragonal crystal environments. This suggests that such perturbations are intrinsic to all stretches of duplex DNA where purines and pyrimidines alternate and may play a role in the detection and exploitation of such sequences by regulatory proteins. Images PMID:6572358

  10. Nanoparticle bridge DNA biosensor

    NASA Astrophysics Data System (ADS)

    Huang, Hong-Wen

    A new DNA sensing method is demonstrated in which DNA hybridization events lead to the formation of nanoparticle satellites that bridge two electrodes and are detected electrically. The hybridization events are exclusively carried out only on specific locations, the surfaces of C-ssDNA modified 50 nm GNPs. The uniqueness of this work is that only a small number of T-ccDNA molecules (<10) is required to form the nanoparticle satellites, allowing ultra-sensitive DNA sensing. The principle of this new DNA sensing technique has been demonstrated using target DNA and three-base-pair-mismatched DNA in 20nM concentrations. Three single-stranded DNA (ssDNA) system is used in our experiment which includes Capture-ssDNA (C-ssDNA), Target-ssDNA (T-ssDNA) and Probe-ssDNA (P-ssDNA). Both C-ssDNA and P-ssDNA are modified by a thiol group and can hybridize with different portions of T-ssDNA. T-ssDNA requires no modification in three ssDNA system, which is beneficial in many applications. C-ssDNA modified 50nm gold nanoparticle (C-50au) and P-ssDNA modified 30nm gold nanoparticle (P-30au) are prepared through the reaction of thiol-gold chemical bonding between thiolated ssDNA and gold nanoparticle (GNP) (C-ssDNA with 50nm GNP, P-ssDNA with 30nm GNP). We controllably place the C-50au only on the SiO2 band surface (˜ 90nm width) between two gold electrodes (source and drain electrodes) by forming positively- and negatively-charged self-assembled monolayers (SAMs) on SiO2 and gold surface, respectively. DNA modified GNP is negatively charged due to ionization of phosphate group on DNA back bone. C-50au therefore is negatively charged and can only be attracted toward SiO2 area (repelled by negatively charged gold electrode surface). The amine group of positively-charged SAMs on SiO2 surface is then passivated by converting to non-polar methyl functional group after C-50au placement. P-30au is first hybridized with T-ssDNA in the solution phase (T-P- 30au formed) and is introduced

  11. Mitochondrial DNA.

    ERIC Educational Resources Information Center

    Wright, Russell G.; Bottino, Paul J.

    1986-01-01

    Provides background information for teachers on mitochondrial DNA, pointing out that it may have once been a free-living organism. Includes a ready-to-duplicate exercise titled "Using Microchondrial DNA to Measure Evolutionary Distance." (JN)

  12. A high-mobility group box 1 that binds to DNA, enhances pro-inflammatory activity, and acts as an anti-infection molecule in black rockfish, Sebastes schlegelii.

    PubMed

    Xin-Peng, Zhao; Yong-Hua, Hu; Yong, Liu; Jing-Jing, Wang; Guang-Hua, Wang; Ren-Jie, Wang; Min, Zhang

    2016-09-01

    High-mobility group box (HMGB) 1 is a chromosomal protein that plays critical roles in DNA transcription, replication and repair. In addition, HMGB1 functions as a pro-inflammatory molecule in many vertebrates and invertebrates. In teleosts, very limited studies of HMGB1 have been reported. In this study, we identified a HMGB1 homologue (SsHMGB1) from black rockfish (Sebastes schlegelii) and analyzed its structure, expression and biological function. The open reading frame of SsHMGB1 is 621 bp, with a 5'-untranslated region (UTR) of 62 bp and a 3'-UTR of 645 bp. SsHMGB1 contains two typical HMG boxes and an acidic C-terminal tail. The deduced amino acid sequence of SsHMGB1 shares the highest overall identity (89.4%) with the HMGB1 of Anoplopoma fimbria. The expression of SsHMGB1 occurred in multiple tissues and was highest in the brain. Moreover, the mRNA level of SsHMGB1 in head kidney (HK) macrophages could be induced by Listonella anguillarum in a time-dependent manner. Recombinant SsHMGB1 purified from Escherichia coli (i) bound DNA fragments in a dose-dependent manner; and (ii) induced the expression of cytokines in HK macrophages, including a significant increase in TNF-α activity and enhanced mRNA level of TNF13B and IL-1 β, which are known to be involved in antibacterial defense; moreover, (iii) significantly improved the macrophage bactericidal activity together with reduced pathogen dissemination and replication of bacteria in fish kidney. These results indicated that SsHMGB1 is a novel HMGB1 that possesses apparent immunoregulatory properties and is likely to be involved in fighting bacterial infection. PMID:27492120

  13. DNA Banking

    SciTech Connect

    Reilly, P.R. )

    1992-11-01

    The author is involved in the ethical, legal, and social issues of banking of DNA and data from DNA analysis. In his attempt to determine the extent of DNA banking in the U.S., the author surveyed some commercial companies performing DNA banking services. This article summarizes the results of that survey, with special emphasis on the procedures the companies use to protect the privacy of individuals. 4 refs.

  14. DNA sequences, recombinant DNA molecules and processes producing human phospholipase inhibitor polypeptides

    SciTech Connect

    Wallner, B.P.; Pepinsky, R.B.; Garwin, J.L.

    1989-11-07

    This patent describes a recombinant DNA molecule. In comprises a DNA sequence coding for a phospholopase inhibitor polypeptide and being selected from the group consisting of: the cDNA insert of ALC, DNA sequences which code on expression for a phospholopase inhibitor, and DNA sequences which are degenerate as a result of the genetic code to either of the foregoing DNA sequences and which code on expression for a phospholipase inhibitor.

  15. Dna Sequencing

    DOEpatents

    Tabor, Stanley; Richardson, Charles C.

    1995-04-25

    A method for sequencing a strand of DNA, including the steps off: providing the strand of DNA; annealing the strand with a primer able to hybridize to the strand to give an annealed mixture; incubating the mixture with four deoxyribonucleoside triphosphates, a DNA polymerase, and at least three deoxyribonucleoside triphosphates in different amounts, under conditions in favoring primer extension to form nucleic acid fragments complementory to the DNA to be sequenced; labelling the nucleic and fragments; separating them and determining the position of the deoxyribonucleoside triphosphates by differences in the intensity of the labels, thereby to determine the DNA sequence.

  16. Use of DNA arrays to identify a mutation in the negative regulator, csrR, responsible for the high virulence of a naturally occurring type M3 group A streptococcus clinical isolate.

    PubMed

    Miyoshi-Akiyama, Tohru; Ikebe, Tadayoshi; Watanabe, Haruo; Uchiyama, Takehiko; Kirikae, Teruo; Kawamura, Yoshiaki

    2006-06-15

    We previously reported that type M3 group A streptococcus (GAS) showed a wide range of 50% lethal dose values in mice. Analysis using DNA arrays indicated that the most virulent strain, M3-f, expressed significantly higher levels of the products of several virulence genes than did the other M3 isolates. Sequencing of the csrS, csrR, luxS, and rgg genes in the isolates showed that the M-3f csrR gene contained a specific point mutation. Disruption of wild-type (wt) csrR in an M3 strain increased its virulence and the expression of hyaluronic acid, whereas complementation with wt but not type M3-f csrR attenuated these changes. Expression experiments showed that type M3-f CsrR counteracted the effects of wt CsrR. Although wt CsrR bound to the hasA promoter region, type M3-f CsrR did not. Thus, the high virulence of the type M3-f strain is associated with the decreased binding of type M3-f CsrR to its target sequences. PMID:16703511

  17. Cinnamate-based DNA photolithography

    PubMed Central

    Romulus, Joy; Li, Minfeng; Sha, Ruojie; Royer, John; Wu, Kun-Ta; Xu, Qin

    2013-01-01

    As demonstrated by means of DNA nanoconstructs[1], as well as DNA functionalization of nanoparticles[2-4] and micrometre-scale colloids[5-8], complex self-assembly processes require components to associate with particular partners in a programmable fashion. In many cases the reversibility of the interactions between complementary DNA sequences is an advantage[9]. However, permanently bonding some or all of the complementary pairs may allow for flexibility in design and construction[10]. Here, we show that the substitution of a pair of complementary bases by a cinnamate group provides an efficient, addressable, UV light-based method to covalently bond complementary DNA. To show the potential of this approach, we wrote micrometre-scale patterns on a surface via UV light and demonstrate the reversible attachment of conjugated DNA and DNA-coated colloids. Our strategy enables both functional DNA photolithography and multi-step, specific binding in self-assembly processes. PMID:23685865

  18. DNA Immunization

    PubMed Central

    Wang, Shixia; Lu, Shan

    2013-01-01

    DNA immunization was discovered in early 1990s and its use has been expanded from vaccine studies to a broader range of biomedical research, such as the generation of high quality polyclonal and monoclonal antibodies as research reagents. In this unit, three common DNA immunization methods are described: needle injection, electroporation and gene gun. In addition, several common considerations related to DNA immunization are discussed. PMID:24510291

  19. Patterning nanocrystals using DNA

    SciTech Connect

    Williams, Shara Carol

    2003-09-01

    One of the goals of nanotechnology is to enable programmed self-assembly of patterns made of various materials with nanometer-sized control. This dissertation describes the results of experiments templating arrangements of gold and semiconductor nanocrystals using 2'-deoxyribonucleic acid (DNA). Previously, simple DNA-templated linear arrangements of two and three nanocrystals structures have been made.[1] Here, we have sought to assemble larger and more complex nanostructures. Gold-DNA conjugates with 50 to 100 bases self-assembled into planned arrangements using strands of DNA containing complementary base sequences. We used two methods to increase the complexity of the arrangements: using branched synthetic doublers within the DNA covalent backbone to create discrete nanocrystal groupings, and incorporating the nanocrystals into a previously developed DNA lattice structure [2][3] that self-assembles from tiles made of DNA double-crossover molecules to create ordered nanoparticle arrays. In the first project, the introduction of a covalently-branched synthetic doubler reagent into the backbone of DNA strands created a branched DNA ''trimer.'' This DNA trimer templated various structures that contained groupings of three and four gold nanoparticles, giving promising, but inconclusive transmission electron microscopy (TEM) results. Due to the presence of a variety of possible structures in the reaction mixtures, and due to the difficulty of isolating the desired structures, the TEM and gel electrophoresis results for larger structures having four particles, and for structures containing both 5 and 10 nm gold nanoparticles were inconclusive. Better results may come from using optical detection methods, or from improved sample preparation. In the second project, we worked toward making two-dimensional ordered arrays of nanocrystals. We replicated and improved upon previous results for making DNA lattices, increasing the size of the lattices to a length greater than

  20. DNA ALTERATIONS

    EPA Science Inventory

    The exposure of an organism to genotoxic chemicals may induce a cascade of genetic events. nitially, structural alterations to DNA are formed. ext, the DNA damage is processed and subsequently expressed in mutant gene products. inally, diseases result from the genetic damage. he ...

  1. Recombinant DNA means and method

    SciTech Connect

    Alford, B.L.; Mao, J.I.; Moir, D.T.; Taunton-Rigby, A.; Vovis, G.F.

    1987-05-19

    This patent describes a transformed living cell selected from the group consisting of fungi, yeast and bacteria, and containing genetic material derived from recombinant DNA material and coding for bovine rennin.

  2. Chilean Pitavia more closely related to Oceania and Old World Rutaceae than to Neotropical groups: evidence from two cpDNA non-coding regions, with a new subfamilial classification of the family

    PubMed Central

    Groppo, Milton; Kallunki, Jacquelyn A.; Pirani, José Rubens; Antonelli, Alexandre

    2012-01-01

    Abstract The position of the plant genus Pitavia within an infrafamilial phylogeny of Rutaceae (rue, or orange family) was investigated with the use of two non-coding regions from cpDNA, the trnL-trnF region and the rps16 intron. The only species of the genus, Pitavia punctata Molina, is restricted to the temperate forests of the Coastal Cordillera of Central-Southern Chile and threatened by loss of habitat. The genus traditionally has been treated as part of tribe Zanthoxyleae (subfamily Rutoideae) where it constitutes the monogeneric tribe Pitaviinae. This tribe and genus are characterized by fruits of 1 to 4 fleshy drupelets, unlike the dehiscent fruits typical of the subfamily. Fifty-five taxa of Rutaceae, representing 53 genera (nearly one-third of those in the family) and all subfamilies, tribes, and almost all subtribes of the family were included. Parsimony and Bayesian inference were used to infer the phylogeny; six taxa of Meliaceae, Sapindaceae, and Simaroubaceae, all members of Sapindales, were also used as out-groups. Results from both analyses were congruent and showed Pitavia as sister to Flindersia and Lunasia, both genera with species scattered through Australia, Philippines, Moluccas, New Guinea and the Malayan region, and phylogenetically far from other Neotropical Rutaceae, such as the Galipeinae (Galipeeae, Rutoideae) and Pteleinae (Toddalieae, former Toddalioideae). Additionally, a new circumscription of the subfamilies of Rutaceae is presented and discussed. Only two subfamilies (both monophyletic) are recognized: Cneoroideae (including Dictyolomatoideae, Spathelioideae, Cneoraceae, and Ptaeroxylaceae) and Rutoideae (including not only traditional Rutoideae but also Aurantioideae, Flindersioideae, and Toddalioideae). As a consequence, Aurantioideae (Citrus and allies) is reduced to tribal rank as Aurantieae. PMID:23717188

  3. [DNA computing].

    PubMed

    Błasiak, Janusz; Krasiński, Tadeusz; Popławski, Tomasz; Sakowski, Sebastian

    2011-01-01

    Biocomputers can be an alternative for traditional "silicon-based" computers, which continuous development may be limited due to further miniaturization (imposed by the Heisenberg Uncertainty Principle) and increasing the amount of information between the central processing unit and the main memory (von Neuman bottleneck). The idea of DNA computing came true for the first time in 1994, when Adleman solved the Hamiltonian Path Problem using short DNA oligomers and DNA ligase. In the early 2000s a series of biocomputer models was presented with a seminal work of Shapiro and his colleguas who presented molecular 2 state finite automaton, in which the restriction enzyme, FokI, constituted hardware and short DNA oligomers were software as well as input/output signals. DNA molecules provided also energy for this machine. DNA computing can be exploited in many applications, from study on the gene expression pattern to diagnosis and therapy of cancer. The idea of DNA computing is still in progress in research both in vitro and in vivo and at least promising results of these research allow to have a hope for a breakthrough in the computer science. PMID:21735816

  4. Interaction of chlorpromazine with DNA.

    PubMed

    Motohashi, N; Kamata, K; Meyer, R

    1990-01-01

    The mechanism of the potential anticancer agent chlorpromazine hydrochloride (CPZ) with DNA was investigated by the techniques of high-performance liquid chromatography (HPLC), viscosity and Raman spectroscopy. It has been suggested from HPLC work that DNA nucleotides (except nucleosides) from either a CPZ-DNA system or a CPZ-nucleotide system. Furthermore, the shear stress of the viscosity of the CPZ-DNA system and the CPZ-nucleotide systems ware shown to be apparently the higher increasing than that of DNA and nucleotide alone. These systems had non-Newtonian properties for the formation of the CPZ-DNA and the CPZ-nucleotide systems under experimental conditions. The Raman spectra showed a dramatic difference at 982 cm-1 due to the symmetric P-O stretching vibration of the PO4(2-) group between dGMP and the CPZ-dGMP system. PMID:2285233

  5. Group Counseling

    ERIC Educational Resources Information Center

    Mahler, Clarence A.

    1971-01-01

    This article reviews the major concerns of group counseling and differentiates among group guidance, group counseling, and group therapy. It also evaluates the research status of group counseling and presents implications for the future of this approach. Comment by Carl E. Thoresen follows. (Author)

  6. Can Epstein-Barr virus DNA load in nasopharyngeal brushings or whole blood predict recurrent nasopharyngeal carcinoma in a non-endemic region? A prospective nationwide study of the Dutch Head and Neck Oncology Cooperative Group.

    PubMed

    Stoker, Sharon D; Wildeman, Maarten A; Novalic, Zlata; Fles, Renske; van der Noort, Vincent; de Bree, Remco; Braunius, Weibel W; van den Broek, Guido B; Kreike, Bas; Kross, Kenneth W; Juwana, Hedy; Ramayanti, Octavia; Verkuijlen, Sandra A W M; de Boer, Jan Paul; Greijer, Astrid E; Middeldorp, Jaap M; Tan, I Bing

    2016-06-01

    This study estimated the value of quantitative measurements of EBV markers in the clinical management of nasopharyngeal carcinoma in a non-endemic area. The aim was to predict prognosis and detect recurrent and residual disease. In 72 patients, EBV DNA load in blood and nasopharyngeal brushes, and IgA VCA-p18 and EBNA1 in plasma were measured at different time points. At diagnosis and post-treatment, a cut-off value was used for detecting disease [positive (PPV) and negative (NPV) predictive value]. The markers were correlated as a continuous variable with tumor stage, disease-free survival (DFS) and overall survival (OS). The Cox hazard ratio model assessed hazard ratios. At diagnosis, the markers were above the COV in 45, 92, 85 and 83 % of the patients, respectively. Post-treatment, DNA load test in blood and brush had the best discriminating power (blood DNA load test: PPV 39 % and NPV 97 %, brush for local disease: PPV 75 % and NPV 99 %). Post-treatment, DNA load in blood was the best predictor for OS and DFS [hazard ratio 3.2 (95 % CI 1.51-3.5) and 2.3 (95 % CI 1.72-5.8)]. Assessing the EBV DNA load in blood has significant prognostic value, although the clinical value is for discussion. The EBV DNA load in the brush might improve early detection of local failures post-treatment. PMID:25929413

  7. Novel applications of DNA materials

    NASA Astrophysics Data System (ADS)

    Ogata, Naoya; Yamaoka, Kanji; Yoshida, Junichi

    2009-08-01

    This paper describes preparations of innovative photonic devices based on high purity DNA molecules which are obtained from Salmon roe. DNA molecules have characteristic features of double helical chain structures where aromatic compounds can intercalate into the stacked layers so that various optically active aromatic dyes indicate strong enhancement effects of photonic activities. Thus, various DNA photonic devices have been developed in the world in terms of optical switches, electro-luminescence (EL), lasers and so on. However, these DNA photonic devices adsorb moisture in the air because of hydrophilic character of DNA molecules, leading to decrease photonic activities. Nevertheless, it was found by my group that a novel hybridization method of the dye-intercalated DNA molecules by means of so-called so-gel process increased stabilities and durability of DNA photonic devices under environmental changes. Also, hybridization of dye-intercalated DNA devices with synthetic polymers including fluorinated poly(methylmethacrylate ) or polycarbonates was successfully carried out by solution blending method, followed by casting the solution to obtain these films which showed stability and durability increases of these DNA photonic devices. DNA-lipid complexes showed a very strong fluorescence amplification by chelating with rare earth metals such as Europium or Telbiumu compounds. This paper also describes the chelate effects of rare earth metal compounds for light amplifications.

  8. Dancing DNA.

    ERIC Educational Resources Information Center

    Pennisi, Elizabeth

    1991-01-01

    An imaging technique that uses fluorescent dyes and allows scientists to track DNA as it moves through gels or in solution is described. The importance, opportunities, and implications of this technique are discussed. (KR)

  9. Unravelling DNA

    NASA Astrophysics Data System (ADS)

    Conroy, Rs; Danilowicz, C.

    2004-04-01

    The forces involved in the biology of life are carefully balanced between stopping thermal fluctuations ripping our DNA apart and having bonds weak enough to allow enzymes to function. The application of recently developed techniques for measuring piconewton forces and imaging at the nanometre scale on a molecule-by-molecule basis has dramatically increased the impact of single-molecule biophysics. This article describes the most commonly used techniques for imaging and manipulating single biomolecules. Using these techniques, the mechanical properties of DNA can be investigated, for example through measurements of the forces required to stretch and unzip the DNA double helix. These properties determine the ease with which DNA can be folded into the cell nucleus and the size and complexity of the accompanying cellular machinery. Part of this cellular machinery is enzymes, which manipulate, repair and transcribe the DNA helix. Enzymatic function is increasingly being investigated at the single molecule level to give better understanding of the forces and processes involved in the genetic cycle. One of the challenges is to transfer this understanding of single molecules into living systems. Already there have been some notable successes, such as the development of techniques for gene expression through the application of mechanical forces to cells, and the imaging and control of viral infection of a cell. This understanding and control of DNA has also been used to design molecules, which can self-assemble into a range of structures.

  10. Tunnelling microscopy of DNA

    NASA Astrophysics Data System (ADS)

    Selci, Stefano; Cricenti, Antonio

    1991-01-01

    Uncoated DNA molecules marked with an activated tris (1-aziridinyl) phosphine oxide (TAPO) solution were deposited on gold substrates and imaged in air with a high resolution Scanning Tunnelling Microscope (STM). The STM operated simultaneously in the constant-current and gap-modulated mode. Highly reproducible STM images have been obtained and interpreted in terms of expected DNA structure. The main periodicity, regularly presented in molecules several hundred Ångstrom long, ranges from 25 Å to 35 Å with an average diameter of 22 Å. Higher resolution images of the minor groove have revealed the phosphate groups along the DNA backbones. Constant-current images of TAPO deposited on gold show a crystalline structure of rows of molecules with a side-by-side spacing of 3 Å.

  11. Comparison of Immunogenicity in Rhesus Macaques of Transmitted-Founder, HIV-1 Group M Consensus, and Trivalent Mosaic Envelope Vaccines Formulated as a DNA Prime, NYVAC, and Envelope Protein Boost

    PubMed Central

    Hulot, Sandrine L.; Korber, Bette; Giorgi, Elena E.; Vandergrift, Nathan; Saunders, Kevin O.; Balachandran, Harikrishnan; Mach, Linh V.; Lifton, Michelle A.; Pantaleo, Giuseppe; Tartaglia, Jim; Phogat, Sanjay; Jacobs, Bertram; Kibler, Karen; Perdiguero, Beatriz; Gomez, Carmen E.; Esteban, Mariano; Rosati, Margherita; Felber, Barbara K.; Pavlakis, George N.; Parks, Robert; Lloyd, Krissey; Sutherland, Laura; Scearce, Richard; Letvin, Norman L.; Seaman, Michael S.; Alam, S. Munir; Montefiori, David; Liao, Hua-Xin; Haynes, Barton F.

    2015-01-01

    ABSTRACT An effective human immunodeficiency virus type 1 (HIV-1) vaccine must induce protective antibody responses, as well as CD4+ and CD8+ T cell responses, that can be effective despite extraordinary diversity of HIV-1. The consensus and mosaic immunogens are complete but artificial proteins, computationally designed to elicit immune responses with improved cross-reactive breadth, to attempt to overcome the challenge of global HIV diversity. In this study, we have compared the immunogenicity of a transmitted-founder (T/F) B clade Env (B.1059), a global group M consensus Env (Con-S), and a global trivalent mosaic Env protein in rhesus macaques. These antigens were delivered using a DNA prime-recombinant NYVAC (rNYVAC) vector and Env protein boost vaccination strategy. While Con-S Env was a single sequence, mosaic immunogens were a set of three Envs optimized to include the most common forms of potential T cell epitopes. Both Con-S and mosaic sequences retained common amino acids encompassed by both antibody and T cell epitopes and were central to globally circulating strains. Mosaics and Con-S Envs expressed as full-length proteins bound well to a number of neutralizing antibodies with discontinuous epitopes. Also, both consensus and mosaic immunogens induced significantly higher gamma interferon (IFN-γ) enzyme-linked immunosorbent spot assay (ELISpot) responses than B.1059 immunogen. Immunization with these proteins, particularly Con-S, also induced significantly higher neutralizing antibodies to viruses than B.1059 Env, primarily to tier 1 viruses. Both Con-S and mosaics stimulated more potent CD8-T cell responses against heterologous Envs than did B.1059. Both antibody and cellular data from this study strengthen the concept of using in silico-designed centralized immunogens for global HIV-1 vaccine development strategies. IMPORTANCE There is an increasing appreciation for the importance of vaccine-induced anti-Env antibody responses for preventing HIV-1

  12. What Is Mitochondrial DNA?

    MedlinePlus

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  13. Ancient DNA

    PubMed Central

    Willerslev, Eske; Cooper, Alan

    2004-01-01

    In the past two decades, ancient DNA research has progressed from the retrieval of small fragments of mitochondrial DNA from a few late Holocene specimens, to large-scale studies of ancient populations, phenotypically important nuclear loci, and even whole mitochondrial genome sequences of extinct species. However, the field is still regularly marred by erroneous reports, which underestimate the extent of contamination within laboratories and samples themselves. An improved understanding of these processes and the effects of damage on ancient DNA templates has started to provide a more robust basis for research. Recent methodological advances have included the characterization of Pleistocene mammal populations and discoveries of DNA preserved in ancient sediments. Increasingly, ancient genetic information is providing a unique means to test assumptions used in evolutionary and population genetics studies to reconstruct the past. Initial results have revealed surprisingly complex population histories, and indicate that modern phylogeographic studies may give misleading impressions about even the recent evolutionary past. With the advent and uptake of appropriate methodologies, ancient DNA is now positioned to become a powerful tool in biological research and is also evolving new and unexpected uses, such as in the search for extinct or extant life in the deep biosphere and on other planets. PMID:15875564

  14. DNA vaccines

    NASA Astrophysics Data System (ADS)

    Gregersen, Jens-Peter

    2001-12-01

    Immunization by genes encoding immunogens, rather than with the immunogen itself, has opened up new possibilities for vaccine research and development and offers chances for new applications and indications for future vaccines. The underlying mechanisms of antigen processing, immune presentation and regulation of immune responses raise high expectations for new and more effective prophylactic or therapeutic vaccines, particularly for vaccines against chronic or persistent infectious diseases and tumors. Our current knowledge and experience of DNA vaccination is summarized and critically reviewed with particular attention to basic immunological mechanisms, the construction of plasmids, screening for protective immunogens to be encoded by these plasmids, modes of application, pharmacokinetics, safety and immunotoxicological aspects. DNA vaccines have the potential to accelerate the research phase of new vaccines and to improve the chances of success, since finding new immunogens with the desired properties is at least technically less demanding than for conventional vaccines. However, on the way to innovative vaccine products, several hurdles have to be overcome. The efficacy of DNA vaccines in humans appears to be much less than indicated by early studies in mice. Open questions remain concerning the persistence and distribution of inoculated plasmid DNA in vivo, its potential to express antigens inappropriately, or the potentially deleterious ability to insert genes into the host cell's genome. Furthermore, the possibility of inducing immunotolerance or autoimmune diseases also needs to be investigated more thoroughly, in order to arrive at a well-founded consensus, which justifies the widespread application of DNA vaccines in a healthy population.

  15. Linking two DNA duplexes with a rigid linker for DNA nanotechnology.

    PubMed

    Tashiro, Ryu; Iwamoto, Masahiro; Morinaga, Hironobu; Emura, Tomoko; Hidaka, Kumi; Endo, Masayuki; Sugiyama, Hiroshi

    2015-08-18

    DNA has recently emerged as a promising material for the construction of nanosized architectures. Chemically modified DNA has been suggested to be an important component of such architectural building blocks. We have designed and synthesized a novel H-shaped DNA oligonucleotide dimer that is cross-linked with a structurally rigid linker composed of phenylene and ethynylene groups. A rotatable DNA unit was constructed through the self-assembly of this H-shaped DNA component and two complementary DNA oligonucleotides. In addition to the rotatable unit, a locked DNA unit containing two H-shaped DNA components was also constructed. As an example of an extended locked structure, a hexagonal DNA origami dimer and oligomer were constructed by using H-shaped DNA as linkers. PMID:26130712

  16. Linking two DNA duplexes with a rigid linker for DNA nanotechnology

    PubMed Central

    Tashiro, Ryu; Iwamoto, Masahiro; Morinaga, Hironobu; Emura, Tomoko; Hidaka, Kumi; Endo, Masayuki; Sugiyama, Hiroshi

    2015-01-01

    DNA has recently emerged as a promising material for the construction of nanosized architectures. Chemically modified DNA has been suggested to be an important component of such architectural building blocks. We have designed and synthesized a novel H-shaped DNA oligonucleotide dimer that is cross-linked with a structurally rigid linker composed of phenylene and ethynylene groups. A rotatable DNA unit was constructed through the self-assembly of this H-shaped DNA component and two complementary DNA oligonucleotides. In addition to the rotatable unit, a locked DNA unit containing two H-shaped DNA components was also constructed. As an example of an extended locked structure, a hexagonal DNA origami dimer and oligomer were constructed by using H-shaped DNA as linkers. PMID:26130712

  17. Group X

    SciTech Connect

    Fields, Susannah

    2007-08-16

    This project is currently under contract for research through the Department of Homeland Security until 2011. The group I was responsible for studying has to remain confidential so as not to affect the current project. All dates, reference links and authors, and other distinguishing characteristics of the original group have been removed from this report. All references to the name of this group or the individual splinter groups has been changed to 'Group X'. I have been collecting texts from a variety of sources intended for the use of recruiting and radicalizing members for Group X splinter groups for the purpose of researching the motivation and intent of leaders of those groups and their influence over the likelihood of group radicalization. This work included visiting many Group X websites to find information on splinter group leaders and finding their statements to new and old members. This proved difficult because the splinter groups of Group X are united in beliefs, but differ in public opinion. They are eager to tear each other down, prove their superiority, and yet remain anonymous. After a few weeks of intense searching, a list of eight recruiting texts and eight radicalizing texts from a variety of Group X leaders were compiled.

  18. Group Flow and Group Genius

    ERIC Educational Resources Information Center

    Sawyer, Keith

    2015-01-01

    Keith Sawyer views the spontaneous collaboration of group creativity and improvisation actions as "group flow," which organizations can use to function at optimum levels. Sawyer establishes ideal conditions for group flow: group goals, close listening, complete concentration, being in control, blending egos, equal participation, knowing…

  19. Isopermutation group

    SciTech Connect

    Muktibodh, A. S.

    2015-03-10

    The concept of ‘Isotopy’ as formulated by Ruggero Maria Santilli [1, 2, 3] plays a vital role in the development of Iso mathematics. Santilli defined iso-fields of characteristic zero. In this paper we extend this definition to define Iso-Galois fields [4] which are essentially of non-zero characteristic. Isotopically isomorphic realizations of a group define isopermutation group which gives a clear cut distinction between automorphic groups and isotopic groups.

  20. Separation Group.

    ERIC Educational Resources Information Center

    Addington, Jean

    1992-01-01

    Describes eight-week short-term group designed to help separated or divorced men and women move through related adjustment phase in focused group setting. Discusses constructs that form the foundations of this short-term psychoeducational and support group and presents brief overview of psychological difficulties that occur as result of marital…

  1. DNA computing.

    PubMed

    Gibbons, A; Amos, M; Hodgson, D

    1997-02-01

    DNA computation is a novel and exciting recent development at the interface of computer science and molecular biology. We describe the current activity in this field following the seminal work of Adleman, who recently showed how techniques of molecular biology may be applied to the solution of a computationally intractable problem. PMID:9013647

  2. DNA Music.

    ERIC Educational Resources Information Center

    Miner, Carol; della Villa, Paula

    1997-01-01

    Describes an activity in which students reverse-translate proteins from their amino acid sequences back to their DNA sequences then assign musical notes to represent the adenine, guanine, cytosine, and thymine bases. Data is obtained from the National Institutes of Health (NIH) on the Internet. (DDR)

  3. DNA Investigations.

    ERIC Educational Resources Information Center

    Mayo, Ellen S.; Bertino, Anthony J.

    1991-01-01

    Presents a simulation activity that allow students to work through the exercise of DNA profiling and to grapple with some analytical and ethical questions involving a couple arranging with a surrogate mother to have a baby. Can be used to teach the principles of restriction enzyme digestion, gel electrophoresis, and probe hybridization. (MDH)

  4. DNA Methylation

    PubMed Central

    Marinus, M.G.; Løbner-Olesen, A.

    2014-01-01

    The DNA of E. coli contains 19,120 6-methyladenines and 12,045 5-methylcytosines in addition to the four regular bases and these are formed by the postreplicative action of three DNA methyltransferases. The majority of the methylated bases are formed by the Dam and Dcm methyltransferases encoded by the dam (DNA adenine methyltransferase) and dcm (DNA cytosine methyltransferase) genes. Although not essential, Dam methylation is important for strand discrimination during repair of replication errors, controlling the frequency of initiation of chromosome replication at oriC, and regulation of transcription initiation at promoters containing GATC sequences. In contrast, there is no known function for Dcm methylation although Dcm recognition sites constitute sequence motifs for Very Short Patch repair of T/G base mismatches. In certain bacteria (e.g., Vibrio cholerae, Caulobacter crescentus) adenine methylation is essential and in C. crescentus, it is important for temporal gene expression which, in turn, is required for coordinating chromosome initiation, replication and division. In practical terms, Dam and Dcm methylation can inhibit restriction enzyme cleavage; decrease transformation frequency in certain bacteria; decrease the stability of short direct repeats; are necessary for site-directed mutagenesis; and to probe eukaryotic structure and function. PMID:26442938

  5. Galaxy groups

    SciTech Connect

    Brent Tully, R.

    2015-02-01

    Galaxy groups can be characterized by the radius of decoupling from cosmic expansion, the radius of the caustic of second turnaround, and the velocity dispersion of galaxies within this latter radius. These parameters can be a challenge to measure, especially for small groups with few members. In this study, results are gathered pertaining to particularly well-studied groups over four decades in group mass. Scaling relations anticipated from theory are demonstrated and coefficients of the relationships are specified. There is an update of the relationship between light and mass for groups, confirming that groups with mass of a few times 10{sup 12}M{sub ⊙} are the most lit up while groups with more and less mass are darker. It is demonstrated that there is an interesting one-to-one correlation between the number of dwarf satellites in a group and the group mass. There is the suggestion that small variations in the slope of the luminosity function in groups are caused by the degree of depletion of intermediate luminosity systems rather than variations in the number per unit mass of dwarfs. Finally, returning to the characteristic radii of groups, the ratio of first to second turnaround depends on the dark matter and dark energy content of the universe and a crude estimate can be made from the current observations of Ω{sub matter}∼0.15 in a flat topology, with a 68% probability of being less than 0.44.

  6. Galaxy Groups

    NASA Astrophysics Data System (ADS)

    Tully, R. Brent

    2015-02-01

    Galaxy groups can be characterized by the radius of decoupling from cosmic expansion, the radius of the caustic of second turnaround, and the velocity dispersion of galaxies within this latter radius. These parameters can be a challenge to measure, especially for small groups with few members. In this study, results are gathered pertaining to particularly well-studied groups over four decades in group mass. Scaling relations anticipated from theory are demonstrated and coefficients of the relationships are specified. There is an update of the relationship between light and mass for groups, confirming that groups with mass of a few times {{10}12}{{M}⊙ } are the most lit up while groups with more and less mass are darker. It is demonstrated that there is an interesting one-to-one correlation between the number of dwarf satellites in a group and the group mass. There is the suggestion that small variations in the slope of the luminosity function in groups are caused by the degree of depletion of intermediate luminosity systems rather than variations in the number per unit mass of dwarfs. Finally, returning to the characteristic radii of groups, the ratio of first to second turnaround depends on the dark matter and dark energy content of the universe and a crude estimate can be made from the current observations of {{Ω}matter}˜ 0.15 in a flat topology, with a 68% probability of being less than 0.44.

  7. Fabrication of patterned DNA surfaces.

    PubMed Central

    Chrisey, L A; O'Ferrall, C E; Spargo, B J; Dulcey, C S; Calvert, J M

    1996-01-01

    Two photolithographic methods are described for the formation of patterned single or multiple DNA species on SiO2 substrates. In the first approach, substrates are treated with a photochemically labile organosilane monolayer film. Irradiation of these surfaces with patterned deep UV (193 nm) light results in patterned chemically reactive groups which are then reacted with heterobifunctional crosslinking molecules. Covalent attachment of modified synthetic DNA oligomers to the crosslinker results in stable DNA patterns. Alternatively, a photoresist is spin-coated over a silane film which had been previously modified with the heterobifunctional crosslinker. Upon patterned irradiation and subsequent development, the underlying crosslinker-modified layer is revealed, and is then reacted with a chemically modified DNA. Feature dimensions to 1 micron are observed when a single fluorescent DNA is attached to the surface. By performing sequential exposures, we have successfully immobilized two distinguishable DNA oligomers on a single surface. Synthetic DNA immobilized in this manner retains the ability to hybridize to its complementary strand, suggesting that these approaches may find utility in the development of miniaturized DNA-based biosensors. PMID:8760891

  8. DNA Binding to the Silica Surface.

    PubMed

    Shi, Bobo; Shin, Yun Kyung; Hassanali, Ali A; Singer, Sherwin J

    2015-08-27

    We investigate the DNA-silica binding mechanism using molecular dynamics simulations. This system is of technological importance, and also of interest to explore how negatively charged DNA can bind to a silica surface, which is also negatively charged at pH values above its isoelectric point near pH 3. We find that the two major binding mechanisms are attractive interactions between DNA phosphate and surface silanol groups and hydrophobic bonding between DNA base and silica hydrophobic region. Umbrella sampling and the weighted histogram analysis method (WHAM) are used to calculate the free energy surface for detachment of DNA from a binding configuration to a location far from the silica surface. Several factors explain why single-stranded DNA (ssDNA) has been observed to be more strongly attracted to silica than double-stranded (dsDNA): (1) ssDNA is more flexible and therefore able to maximize the number of binding interactions. (2) ssDNA has free unpaired bases to form hydrophobic attachment to silica while dsDNA has to break hydrogen bonds with base partners to get free bases. (3) The linear charge density of dsDNA is twice that of ssDNA. We devise a procedure to approximate the atomic forces between biomolecules and amorphous silica to enable large-scale biomolecule-silica simulations as reported here. PMID:25966319

  9. Label-free and selective photoelectrochemical detection of chemical DNA methylation damage using DNA repair enzymes.

    PubMed

    Wu, Yiping; Zhang, Bintian; Guo, Liang-Hong

    2013-07-16

    Exogenous chemicals may produce DNA methylation that is potentially toxic to living systems. Methylated DNA bases are difficult to detect with biosensors because the methyl group is small and chemically inert. In this report, a label-free photoelectrochemical sensor was developed for the selective detection of chemically methylated bases in DNA films. The sensor employed two DNA repair enzymes, human alkyladenine DNA glycosylase and human apurinic/apyrimidinic endonuclease, to convert DNA methylation sites in DNA films on indium tin oxide electrodes into strand breaks. A DNA intercalator, Ru(bpy)2(dppz)(2+) (bpy=2,2'-bipyridine, dppz = dipyrido[3,2-a:2',3'-c]phenazine) was then used as the photoelectrochemical signal indicator to detect the DNA strand breaks. Its photocurrent signal was found to correlate inversely with the amount of 3-methyladenines (metAde) produced with a methylating agent, methylmethane sulfonate (MMS). The sensor detected the methylated bases produced with as low as 1 mM MMS, at which concentration the amount of metAde on the sensor surface was estimated to be 0.5 pg, or 1 metAde in 1.6 × 10(5) normal bases. Other DNA base modification products, such as 5-methylcytosine and DNA adducts with ethyl and styrene groups did not attenuate the photocurrent, demonstrating good selectivity of the sensor. This strategy can be utilized to develop sensors for the detection of other modified DNA bases with specific DNA repair enzymes. PMID:23777269

  10. Group Grammar

    ERIC Educational Resources Information Center

    Adams, Karen

    2015-01-01

    In this article Karen Adams demonstrates how to incorporate group grammar techniques into a classroom activity. In the activity, students practice using the target grammar to do something they naturally enjoy: learning about each other.

  11. DNA Microarrays

    NASA Astrophysics Data System (ADS)

    Nguyen, C.; Gidrol, X.

    Genomics has revolutionised biological and biomedical research. This revolution was predictable on the basis of its two driving forces: the ever increasing availability of genome sequences and the development of new technology able to exploit them. Up until now, technical limitations meant that molecular biology could only analyse one or two parameters per experiment, providing relatively little information compared with the great complexity of the systems under investigation. This gene by gene approach is inadequate to understand biological systems containing several thousand genes. It is essential to have an overall view of the DNA, RNA, and relevant proteins. A simple inventory of the genome is not sufficient to understand the functions of the genes, or indeed the way that cells and organisms work. For this purpose, functional studies based on whole genomes are needed. Among these new large-scale methods of molecular analysis, DNA microarrays provide a way of studying the genome and the transcriptome. The idea of integrating a large amount of data derived from a support with very small area has led biologists to call these chips, borrowing the term from the microelectronics industry. At the beginning of the 1990s, the development of DNA chips on nylon membranes [1, 2], then on glass [3] and silicon [4] supports, made it possible for the first time to carry out simultaneous measurements of the equilibrium concentration of all the messenger RNA (mRNA) or transcribed RNA in a cell. These microarrays offer a wide range of applications, in both fundamental and clinical research, providing a method for genome-wide characterisation of changes occurring within a cell or tissue, as for example in polymorphism studies, detection of mutations, and quantitative assays of gene copies. With regard to the transcriptome, it provides a way of characterising differentially expressed genes, profiling given biological states, and identifying regulatory channels.

  12. Electronic transport in methylated fragments of DNA

    NASA Astrophysics Data System (ADS)

    de Almeida, M. L.; Oliveira, J. I. N.; Lima Neto, J. X.; Gomes, C. E. M.; Fulco, U. L.; Albuquerque, E. L.; Freire, V. N.; Caetano, E. W. S.; de Moura, F. A. B. F.; Lyra, M. L.

    2015-11-01

    We investigate the electronic transport properties of methylated deoxyribonucleic-acid (DNA) strands, a biological system in which methyl groups are added to DNA (a major epigenetic modification in gene expression), sandwiched between two metallic platinum electrodes. Our theoretical simulations apply an effective Hamiltonian based on a tight-binding model to obtain current-voltage curves related to the non-methylated/methylated DNA strands. The results suggest potential applications in the development of novel biosensors for molecular diagnostics.

  13. Electronic transport in methylated fragments of DNA

    SciTech Connect

    Almeida, M. L. de; Oliveira, J. I. N.; Lima Neto, J. X.; Gomes, C. E. M.; Fulco, U. L. Albuquerque, E. L.; Freire, V. N.; Caetano, E. W. S.; Moura, F. A. B. F. de; Lyra, M. L.

    2015-11-16

    We investigate the electronic transport properties of methylated deoxyribonucleic-acid (DNA) strands, a biological system in which methyl groups are added to DNA (a major epigenetic modification in gene expression), sandwiched between two metallic platinum electrodes. Our theoretical simulations apply an effective Hamiltonian based on a tight-binding model to obtain current-voltage curves related to the non-methylated/methylated DNA strands. The results suggest potential applications in the development of novel biosensors for molecular diagnostics.

  14. Polymer microspheres carrying fluorescent DNA probes

    NASA Astrophysics Data System (ADS)

    Chen, Xiaoyu; Dai, Zhao; Zhang, Jimei; Xu, Shichao; Wu, Chunrong; Zheng, Guo

    2010-07-01

    A polymer microspheres carried DNA probe, which was based on resonance energy transfer, was presented in this paper when CdTe quantum dots(QDs) were as energy donors, Au nanoparticles were as energy accepters and poly(4- vinylpyrindine-co-ethylene glycol dimethacrylate) microspheres were as carriers. Polymer microspheres with functional group on surfaces were prepared by distillation-precipitation polymerization when ethylene glycol dimethacrylate was as crosslinker in acetonitrile. CdTe QDs were prepared when 3-mercaptopropionic acid(MPA) was as the stabilizer in aqueous solution. Because of the hydrogen-bonding between the carboxyl groups of MPA on QDs and the pyrindine groups on the microspheres, the QDs were self-assembled onto the surfaces of microspheres. Then, the other parts of DNA probe were finished according to the classic method. The DNA detection results indicated that this novel fluorescent DNA probe system could recognize the existence of complementary target DNA or not.

  15. Group dynamics.

    PubMed

    Scandiffio, A L

    1990-12-01

    Group dynamics play a significant role within any organization, culture, or unit. The important thing to remember with any of these structures is that they are made up of people--people with different ideas, motivations, background, and sometimes different agendas. Most groups, formal or informal, look for a leader in an effort to maintain cohesiveness of the unit. At times, that cultural bond must be developed; once developed, it must be nurtured. There are also times that one of the group no longer finds the culture comfortable and begins to act out behaviorally. It is these times that become trying for the leader as she or he attempts to remain objective when that which was once in the building phase of group cohesiveness starts to fall apart. At all times, the manager must continue to view the employee creating the disturbance as an integral part of the group. It is at this time that it is beneficial to perceive the employee exhibiting problem behaviors as a special employee, as one who needs the benefit of your experience and skills, as one who is still part of the group. It is also during this time that the manager should focus upon her or his own views in the area of power, communication, and the corporate culture of the unit that one has established before attempting to understand another's point of view. Once we understand our own motivation and accept ourselves, it is then that we may move on to offer assistance to another. Once we understand our insecurities recognizing staff dysfunction as a symptom of system dysfunction will not be so threatening to the concept of the manager that we perceive ourselves to be. It takes a secure person to admit that she or he favors staff before deciding to do something to change things. The important thing to know is that it can be done. The favored staff can find a new way of relating to others, the special employee can find new modes of behavior (and even find self-esteem in the process), the group can find new ways

  16. Optical DNA

    NASA Astrophysics Data System (ADS)

    Vijaywargi, Deepak; Lewis, Dave; Kirovski, Darko

    A certificate of authenticity (COA) is an inexpensive physical object with a random and unique structure S which is hard to near-exactly replicate. An inexpensive device should be able to scan object’s physical “fingerprint,” a set of features that represents S. In this paper, we explore one set of requirements that optical media such as DVDs should satisfy, to be considered as COAs. As manufacturing of such media produces inevitable errors, we use the locations and count of these errors as a “fingerprint” for each optical disc: its optical DNA. The “fingerprint” is signed using publisher’s private-key and the resulting signature is stored onto the optical medium using a post-production process. Standard DVD players with altered firmware that includes publisher’s public-key, should be able to verify the authenticity of DVDs protected with optical DNA. Our key finding is that for the proposed protocol, only DVDs with exceptional wear-and-tear characteristics would result in an inexpensive and viable anti-counterfeiting technology.

  17. Nanomechanical molecular devices made of DNA origami.

    PubMed

    Kuzuya, Akinori; Ohya, Yuichi

    2014-06-17

    CONSPECTUS: Eight years have passed since the striking debut of the DNA origami technique ( Rothemund, P. W. K. Nature 2006 , 440 , 297 - 302 ), in which long single-stranded DNA is folded into a designed nanostructure, in either 2D or 3D, with the aid of many short staple strands. The number of proposals for new design principles for DNA origami structures seems to have already reached a peak. It is apparent that DNA origami study is now entering the second phase of creating practical applications. The development of functional nanomechanical molecular devices using the DNA origami technique is one such application attracting significant interest from researchers in the field. Nanomechanical DNA origami devices, which maintain the characteristics of DNA origami structures, have various advantages over conventional DNA nanomachines. Comparatively high assembly yield, relatively large size visible via atomic force microscopy (AFM) or transmission electron microscopy (TEM), and the capability to assemble multiple functional groups with precision using multiple staple strands are some of the advantages of the DNA origami technique for constructing sophisticated molecular devices. This Account describes the recent developments of such nanomechanical DNA origami devices and reviews the emerging target of DNA origami studies. First, simple "dynamic" DNA origami structures with transformation capability, such as DNA origami boxes and a DNA origami hatch with structure control, are briefly summarized. More elaborate nanomechanical DNA origami devices are then reviewed. The first example describes DNA origami pinching devices that can be used as "single-molecule" beacons to detect a variety of biorelated molecules, from metal ions at the size of a few tens of atomic mass number units to relatively gigantic proteins with a molecular mass greater than a hundred kilodaltons, all on a single platform. Clamshell-like DNA nanorobots equipped with logic gates can discriminate

  18. DNA sequencing using fluorescence background electroblotting membrane

    DOEpatents

    Caldwell, K.D.; Chu, T.J.; Pitt, W.G.

    1992-05-12

    A method for the multiplex sequencing on DNA is disclosed which comprises the electroblotting or specific base terminated DNA fragments, which have been resolved by gel electrophoresis, onto the surface of a neutral non-aromatic polymeric microporous membrane exhibiting low background fluorescence which has been surface modified to contain amino groups. Polypropylene membranes are preferably and the introduction of amino groups is accomplished by subjecting the membrane to radio or microwave frequency plasma discharge in the presence of an aminating agent, preferably ammonia. The membrane, containing physically adsorbed DNA fragments on its surface after the electroblotting, is then treated with crosslinking means such as UV radiation or a glutaraldehyde spray to chemically bind the DNA fragments to the membrane through amino groups contained on the surface. The DNA fragments chemically bound to the membrane are subjected to hybridization probing with a tagged probe specific to the sequence of the DNA fragments. The tagging may be by either fluorophores or radioisotopes. The tagged probes hybridized to the target DNA fragments are detected and read by laser induced fluorescence detection or autoradiograms. The use of aminated low fluorescent background membranes allows the use of fluorescent detection and reading even when the available amount of DNA to be sequenced is small. The DNA bound to the membranes may be reprobed numerous times. No Drawings

  19. DNA sequencing using fluorescence background electroblotting membrane

    DOEpatents

    Caldwell, Karin D.; Chu, Tun-Jen; Pitt, William G.

    1992-01-01

    A method for the multiplex sequencing on DNA is disclosed which comprises the electroblotting or specific base terminated DNA fragments, which have been resolved by gel electrophoresis, onto the surface of a neutral non-aromatic polymeric microporous membrane exhibiting low background fluorescence which has been surface modified to contain amino groups. Polypropylene membranes are preferably and the introduction of amino groups is accomplished by subjecting the membrane to radio or microwave frequency plasma discharge in the presence of an aminating agent, preferably ammonia. The membrane, containing physically adsorbed DNA fragments on its surface after the electroblotting, is then treated with crosslinking means such as UV radiation or a glutaraldehyde spray to chemically bind the DNA fragments to the membrane through said smino groups contained on the surface thereof. The DNA fragments chemically bound to the membrane are subjected to hybridization probing with a tagged probe specific to the sequence of the DNA fragments. The tagging may be by either fluorophores or radioisotopes. The tagged probes hybridized to said target DNA fragments are detected and read by laser induced fluorescence detection or autoradiograms. The use of aminated low fluorescent background membranes allows the use of fluorescent detection and reading even when the available amount of DNA to be sequenced is small. The DNA bound to the membrances may be reprobed numerous times.

  20. Distinct DNA methylomes of newborns and centenarians

    PubMed Central

    Heyn, Holger; Li, Ning; Ferreira, Humberto J.; Moran, Sebastian; Pisano, David G.; Gomez, Antonio; Diez, Javier; Sanchez-Mut, Jose V.; Setien, Fernando; Carmona, F. Javier; Puca, Annibale A.; Sayols, Sergi; Pujana, Miguel A.; Serra-Musach, Jordi; Iglesias-Platas, Isabel; Formiga, Francesc; Fernandez, Agustin F.; Fraga, Mario F.; Heath, Simon C.; Valencia, Alfonso; Gut, Ivo G.; Wang, Jun; Esteller, Manel

    2012-01-01

    Human aging cannot be fully understood in terms of the constrained genetic setting. Epigenetic drift is an alternative means of explaining age-associated alterations. To address this issue, we performed whole-genome bisulfite sequencing (WGBS) of newborn and centenarian genomes. The centenarian DNA had a lower DNA methylation content and a reduced correlation in the methylation status of neighboring cytosine—phosphate—guanine (CpGs) throughout the genome in comparison with the more homogeneously methylated newborn DNA. The more hypomethylated CpGs observed in the centenarian DNA compared with the neonate covered all genomic compartments, such as promoters, exonic, intronic, and intergenic regions. For regulatory regions, the most hypomethylated sequences in the centenarian DNA were present mainly at CpG-poor promoters and in tissue-specific genes, whereas a greater level of DNA methylation was observed in CpG island promoters. We extended the study to a larger cohort of newborn and nonagenarian samples using a 450,000 CpG-site DNA methylation microarray that reinforced the observation of more hypomethylated DNA sequences in the advanced age group. WGBS and 450,000 analyses of middle-age individuals demonstrated DNA methylomes in the crossroad between the newborn and the nonagenarian/centenarian groups. Our study constitutes a unique DNA methylation analysis of the extreme points of human life at a single-nucleotide resolution level. PMID:22689993

  1. Compressive Sensing DNA Microarrays

    PubMed Central

    2009-01-01

    Compressive sensing microarrays (CSMs) are DNA-based sensors that operate using group testing and compressive sensing (CS) principles. In contrast to conventional DNA microarrays, in which each genetic sensor is designed to respond to a single target, in a CSM, each sensor responds to a set of targets. We study the problem of designing CSMs that simultaneously account for both the constraints from CS theory and the biochemistry of probe-target DNA hybridization. An appropriate cross-hybridization model is proposed for CSMs, and several methods are developed for probe design and CS signal recovery based on the new model. Lab experiments suggest that in order to achieve accurate hybridization profiling, consensus probe sequences are required to have sequence homology of at least 80% with all targets to be detected. Furthermore, out-of-equilibrium datasets are usually as accurate as those obtained from equilibrium conditions. Consequently, one can use CSMs in applications in which only short hybridization times are allowed. PMID:19158952

  2. Group Learning.

    ERIC Educational Resources Information Center

    Black, Susan

    1992-01-01

    Research suggests that cooperative learning works best when students are first taught group-processing skills, such as leadership, decision making, communication, trust building, and conflict management. Inadequate teacher training and boring assignments can torpedo cooperative learning efforts. Administrators should reassure teachers with…

  3. DNA mimicry by proteins.

    PubMed

    Dryden, D T F; Tock, M R

    2006-04-01

    It has been discovered recently, via structural and biophysical analyses, that proteins can mimic DNA structures in order to inhibit proteins that would normally bind to DNA. Mimicry of the phosphate backbone of DNA, the hydrogen-bonding properties of the nucleotide bases and the bending and twisting of the DNA double helix are all present in the mimics discovered to date. These mimics target a range of proteins and enzymes such as DNA restriction enzymes, DNA repair enzymes, DNA gyrase and nucleosomal and nucleoid-associated proteins. The unusual properties of these protein DNA mimics may provide a foundation for the design of targeted inhibitors of DNA-binding proteins. PMID:16545103

  4. Anthraquinones quinizarin and danthron unwind negatively supercoiled DNA and lengthen linear DNA

    SciTech Connect

    Verebová, Valéria; Adamcik, Jozef; Danko, Patrik; Podhradský, Dušan; Miškovský, Pavol; Staničová, Jana

    2014-01-31

    Highlights: • Anthraquinones quinizarin and danthron unwind negatively supercoiled DNA. • Anthraquinones quinizarin and danthron lengthen linear DNA. • Anthraquinones quinizarin and danthron possess middle binding affinity to DNA. • Anthraquinones quinizarin and danthron interact with DNA by intercalating mode. - Abstract: The intercalating drugs possess a planar aromatic chromophore unit by which they insert between DNA bases causing the distortion of classical B-DNA form. The planar tricyclic structure of anthraquinones belongs to the group of chromophore units and enables anthraquinones to bind to DNA by intercalating mode. The interactions of simple derivatives of anthraquinone, quinizarin (1,4-dihydroxyanthraquinone) and danthron (1,8-dihydroxyanthraquinone), with negatively supercoiled and linear DNA were investigated using a combination of the electrophoretic methods, fluorescence spectrophotometry and single molecule technique an atomic force microscopy. The detection of the topological change of negatively supercoiled plasmid DNA, unwinding of negatively supercoiled DNA, corresponding to appearance of DNA topoisomers with the low superhelicity and an increase of the contour length of linear DNA in the presence of quinizarin and danthron indicate the binding of both anthraquinones to DNA by intercalating mode.

  5. Underrepresented groups

    NASA Technical Reports Server (NTRS)

    Peters, David A.

    1990-01-01

    The problem with the shortage of under represented groups in science and engineering is absolutely crucial, especially considering that U.S. will experience a shortage of 560,000 science and engineering personnel by the year 2010. Most studies by the National Science Foundation also concluded that projected shortages cannot be alleviated without significant increases in the involvement of Blacks, Hispanics, Native Americans, handicapped persons, and women.

  6. Cantor Groups

    ERIC Educational Resources Information Center

    Mathes, Ben; Dow, Chris; Livshits, Leo

    2011-01-01

    The Cantor subset of the unit interval [0, 1) is "large" in cardinality and also "large" algebraically, that is, the smallest subgroup of [0, 1) generated by the Cantor set (using addition mod 1 as the group operation) is the whole of [0, 1). In this paper, we show how to construct Cantor-like sets which are "large" in cardinality but "small"…

  7. DNA Damage and Pulmonary Hypertension

    PubMed Central

    Ranchoux, Benoît; Meloche, Jolyane; Paulin, Roxane; Boucherat, Olivier; Provencher, Steeve; Bonnet, Sébastien

    2016-01-01

    Pulmonary hypertension (PH) is defined by a mean pulmonary arterial pressure over 25 mmHg at rest and is diagnosed by right heart catheterization. Among the different groups of PH, pulmonary arterial hypertension (PAH) is characterized by a progressive obstruction of distal pulmonary arteries, related to endothelial cell dysfunction and vascular cell proliferation, which leads to an increased pulmonary vascular resistance, right ventricular hypertrophy, and right heart failure. Although the primary trigger of PAH remains unknown, oxidative stress and inflammation have been shown to play a key role in the development and progression of vascular remodeling. These factors are known to increase DNA damage that might favor the emergence of the proliferative and apoptosis-resistant phenotype observed in PAH vascular cells. High levels of DNA damage were reported to occur in PAH lungs and remodeled arteries as well as in animal models of PH. Moreover, recent studies have demonstrated that impaired DNA-response mechanisms may lead to an increased mutagen sensitivity in PAH patients. Finally, PAH was linked with decreased breast cancer 1 protein (BRCA1) and DNA topoisomerase 2-binding protein 1 (TopBP1) expression, both involved in maintaining genome integrity. This review aims to provide an overview of recent evidence of DNA damage and DNA repair deficiency and their implication in PAH pathogenesis. PMID:27338373

  8. DNA Damage and Pulmonary Hypertension.

    PubMed

    Ranchoux, Benoît; Meloche, Jolyane; Paulin, Roxane; Boucherat, Olivier; Provencher, Steeve; Bonnet, Sébastien

    2016-01-01

    Pulmonary hypertension (PH) is defined by a mean pulmonary arterial pressure over 25 mmHg at rest and is diagnosed by right heart catheterization. Among the different groups of PH, pulmonary arterial hypertension (PAH) is characterized by a progressive obstruction of distal pulmonary arteries, related to endothelial cell dysfunction and vascular cell proliferation, which leads to an increased pulmonary vascular resistance, right ventricular hypertrophy, and right heart failure. Although the primary trigger of PAH remains unknown, oxidative stress and inflammation have been shown to play a key role in the development and progression of vascular remodeling. These factors are known to increase DNA damage that might favor the emergence of the proliferative and apoptosis-resistant phenotype observed in PAH vascular cells. High levels of DNA damage were reported to occur in PAH lungs and remodeled arteries as well as in animal models of PH. Moreover, recent studies have demonstrated that impaired DNA-response mechanisms may lead to an increased mutagen sensitivity in PAH patients. Finally, PAH was linked with decreased breast cancer 1 protein (BRCA1) and DNA topoisomerase 2-binding protein 1 (TopBP1) expression, both involved in maintaining genome integrity. This review aims to provide an overview of recent evidence of DNA damage and DNA repair deficiency and their implication in PAH pathogenesis. PMID:27338373

  9. Mitochondrial DNA replacement versus nuclear DNA persistence

    NASA Astrophysics Data System (ADS)

    Serva, Maurizio

    2006-10-01

    In this paper we consider two populations whose generations are not overlapping and whose size is large. The number of males and females in both populations is constant. Any generation is replaced by a new one and any individual has two parents concerning nuclear DNA and a single one (the mother) concerning mtDNA. Moreover, at any generation some individuals migrate from the first population to the second. In a finite random time T, the mtDNA of the second population is completely replaced by the mtDNA of the first. In the same time, the nuclear DNA is not completely replaced and a fraction F of the ancient nuclear DNA persists. We compute both T and F. Since this study shows that complete replacement of mtDNA in a population is compatible with the persistence of a large fraction of nuclear DNA, it may have some relevance for the 'out of Africa'/multiregional debate in palaeoanthropology.

  10. DNA modifications: Another stable base in DNA

    NASA Astrophysics Data System (ADS)

    Brazauskas, Pijus; Kriaucionis, Skirmantas

    2014-12-01

    Oxidation of 5-methylcytosine has been proposed to mediate active and passive DNA demethylation. Tracking the history of DNA modifications has now provided the first solid evidence that 5-hydroxymethylcytosine is a stable epigenetic modification.

  11. Synthesis of DNA

    DOEpatents

    Mariella, Jr., Raymond P.

    2008-11-18

    A method of synthesizing a desired double-stranded DNA of a predetermined length and of a predetermined sequence. Preselected sequence segments that will complete the desired double-stranded DNA are determined. Preselected segment sequences of DNA that will be used to complete the desired double-stranded DNA are provided. The preselected segment sequences of DNA are assembled to produce the desired double-stranded DNA.

  12. [Mitochondrial disease and mitochondrial DNA depletion syndromes].

    PubMed

    Huang, Chin-Chang; Hsu, Chang-Huang

    2009-12-01

    Mitochondria is an intracellular double membrane-bound structure and it can provide energy for intracellular metabolism. The metabolism includes Krebs cycle, beta-oxidation and lipid synthesis. The density of mitochondria is different in various tissues dependent upon the demands of oxidative phosphorylation. Mitochondrial diseases can occur by defects either in mitochondrial DNA or nuclear DNA. Human mitochondrial DNA (mtDNA) encoding for 22 tRNAs, 2 rRNAs and 13 mRNAs that are translated in the mitochondria. Mitochondrial genetic diseases are most resulted from defects in the mtDNA which may be point mutations, deletions, or mitochondrial DNA depletion. These patterns of inheritance in mitochondrial diseases include sporadic, maternally inherited, or of Mendelian inheritance. Mitochondrial DNA depletion is caused by defects in the nuclear genes that are responsible for maintenance of integrity of mtDNA or deoxyribonucelotide pools and mtDNA biogenesis. The mtDNA depletion syndrome (MDS) includes the following categories: progressive external ophthalmoplegia (PEO), predominant myopathy, mitochondrial neurogastrointestinal encephalomyopathy (MNGIE), sensory-ataxic neuropathy, dysarthria, and ophthalmoplegia (SANDO) and hepato-encephalopathy. The most common tissues or organs involved in MDS and related disorders include the brain, liver and muscles. These involved genes are divided into two groups including 1) DNA polymerase gamma (POLG, POLG2) and Twinkle genes whose products function directly at the mtDNA replication fork, and 2) adenine nucleotide translocator 1, thymidine phosphorylase, thymidine kinase 2, deoxyguanosine kinase, ADP-forming succinyl-CoA synthetase ligase, MPV17 whose products supply the mitochondria with deoxyribonucleotide triphosphate pools needed for mtDNA replication, and possible mutation in the RRM2B gene. The development has provided new information about the importance of the biosynthetic pathway of the nucleotides for mtDNA replication

  13. DNA encoding a DNA repair protein

    DOEpatents

    Petrini, John H.; Morgan, William Francis; Maser, Richard Scott; Carney, James Patrick

    2006-08-15

    An isolated and purified DNA molecule encoding a DNA repair protein, p95, is provided, as is isolated and purified p95. Also provided are methods of detecting p95 and DNA encoding p95. The invention further provides p95 knock-out mice.

  14. DNA polymerases and cancer

    PubMed Central

    Lange, Sabine S.; Takata, Kei-ichi; Wood, Richard D.

    2013-01-01

    There are fifteen different DNA polymerases encoded in mammalian genomes, which are specialized for replication, repair or the tolerance of DNA damage. New evidence is emerging for lesion-specific and tissue-specific functions of DNA polymerases. Many point mutations that occur in cancer cells arise from the error-generating activities of DNA polymerases. However, the ability of some of these enzymes to bypass DNA damage may actually defend against chromosome instability in cells and at least one DNA polymerase, POLζ, is a suppressor of spontaneous tumorigenesis. Because DNA polymerases can help cancer cells tolerate DNA damage, some of these enzymes may be viable targets for therapeutic strategies. PMID:21258395

  15. Energy transport in crystalline DNA composites

    SciTech Connect

    Xu, Zaoli; Xu, Shen; Tang, Xiaoduan; Wang, Xinwei

    2014-01-15

    This work reports on the synthesis of crystalline DNA-composited films and microfibers, and details the study of thermal energy transport in them. The transient electro-thermal technique is used to characterize the thermal transport in DNA composite microfibers, and the photothermal technique is used to explore the thermal transport in the thickness direction of DNA films. Compared with microfibers, the DNA films are found to have a higher thermal transport capacity, largely due to the carefully controlled crystallization process in film synthesis. In high NaCl concentration solutions, the bond of the Na{sup +} ion and phosphate group aligns the DNA molecules with the NaCl crystal structure during crystallization. This results in significant enhancement of thermal transport in the DNA films with aligned structure.

  16. Permanent or reversible conjugation of 2′-O- or 5′-O-aminooxymethylated nucleosides with functional groups as a convenient and efficient approach to the modification of RNA and DNA sequences

    PubMed Central

    Cieślak, Jacek; Grajkowski, Andrzej; Ausín, Cristina; Gapeev, Alexei; Beaucage, Serge L.

    2012-01-01

    2′-O-Aminooxymethyl ribonucleosides are prepared from their 3′,5′-disilylated 2′-O-phthalimidooxymethyl derivatives by treatment with NH4F in MeOH. The reaction of these novel ribonucleosides with 1-pyrenecarboxaldehyde results in the efficient formation of stable and yet reversible ribonucleoside 2′-conjugates in yields of 69–82%. Indeed, exposure of these conjugates to 0.5 M tetra-n-butylammonium fluoride (TBAF) in THF results in the cleavage of their iminoether functions to give the native ribonucleosides along with the innocuous nitrile side product. Conversely, the reaction of 5-cholesten-3-one or dansyl chloride with 2′-O-aminooxymethyl uridine provides permanent uridine 2′-conjugates, which are left essentially intact upon treatment with TBAF. Alternatively, 5′-O-aminooxymethyl thymidine is prepared by hydrazinolysis of its 3′-O-levulinyl-5′-O-phthalimidooxymethyl precursor. Pyrenylation of 5′-O-aminooxymethyl thymidine and the sensitivity of the 5′-conjugate to TBAF further exemplify the usefulness of this nucleoside for modifying DNA sequences either permanently or reversibly. Although the versatility and uniqueness of 2′-O-aminooxymethyl ribonucleosides in the preparation of modified RNA sequences is demonstrated by the single or double incorporation of a reversible pyrenylated uridine 2′-conjugate into an RNA sequence, the conjugation of 2′-O-aminooxymethyl ribonucleosides with aldehydes, including those generated from their acetals, provides reversible 2′-O-protected ribonucleosides for potential applications in the solid-phase synthesis of native RNA sequences. The synthesis of a chimeric polyuridylic acid is presented as an exemplary model. PMID:22067450

  17. DNA/RNA Detection Using DNA-Templated Few-Atom Silver Nanoclusters.

    PubMed

    Obliosca, Judy M; Liu, Cong; Batson, Robert Austin; Babin, Mark C; Werner, James H; Yeh, Hsin-Chih

    2013-01-01

    DNA-templated few-atom silver nanoclusters (DNA/Ag NCs) are a new class of organic/inorganic composite nanomaterials whose fluorescence emission can be tuned throughout the visible and near-IR range by simply programming the template sequences. Compared to organic dyes, DNA/Ag NCs can be brighter and more photostable. Compared to quantum dots, DNA/Ag NCs are smaller, less prone to blinking on long timescales, and do not have a toxic core. The preparation of DNA/Ag NCs is simple and there is no need to remove excess precursors as these precursors are non-fluorescent. Our recent discovery of the fluorogenic and color switching properties of DNA/Ag NCs have led to the invention of new molecular probes, termed NanoCluster Beacons (NCBs), for DNA detection, with the capability to differentiate single-nucleotide polymorphisms by emission colors. NCBs are inexpensive, easy to prepare, and compatible with commercial DNA synthesizers. Many other groups have also explored and taken advantage of the environment sensitivities of DNA/Ag NCs in creating new tools for DNA/RNA detection and single-nucleotide polymorphism identification. In this review, we summarize the recent trends in the use of DNA/Ag NCs for developing DNA/RNA sensors. PMID:25586126

  18. DNA/RNA Detection Using DNA-Templated Few-Atom Silver Nanoclusters

    PubMed Central

    Obliosca, Judy M.; Liu, Cong; Batson, Robert Austin; Babin, Mark C.; Werner, James H.; Yeh, Hsin-Chih

    2013-01-01

    DNA-templated few-atom silver nanoclusters (DNA/Ag NCs) are a new class of organic/inorganic composite nanomaterials whose fluorescence emission can be tuned throughout the visible and near-IR range by simply programming the template sequences. Compared to organic dyes, DNA/Ag NCs can be brighter and more photostable. Compared to quantum dots, DNA/Ag NCs are smaller, less prone to blinking on long timescales, and do not have a toxic core. The preparation of DNA/Ag NCs is simple and there is no need to remove excess precursors as these precursors are non-fluorescent. Our recent discovery of the fluorogenic and color switching properties of DNA/Ag NCs have led to the invention of new molecular probes, termed NanoCluster Beacons (NCBs), for DNA detection, with the capability to differentiate single-nucleotide polymorphisms by emission colors. NCBs are inexpensive, easy to prepare, and compatible with commercial DNA synthesizers. Many other groups have also explored and taken advantage of the environment sensitivities of DNA/Ag NCs in creating new tools for DNA/RNA detection and single-nucleotide polymorphism identification. In this review, we summarize the recent trends in the use of DNA/Ag NCs for developing DNA/RNA sensors. PMID:25586126

  19. DNA extraction techniques for use in education.

    PubMed

    Hearn, R P; Arblaster, K E

    2010-05-01

    DNA extraction provides a hands-on introduction to DNA and enables students to gain real life experience and practical knowledge of DNA. Students gain a sense of ownership and are more enthusiastic when they use their own DNA. A cost effective, simple protocol for DNA extraction and visualization was devised. Buccal mucosal epithelia provide a readily available source of cells for DNA extraction and can be harvested in a painless, noninvasive manner. Seven criteria were established to evaluate the protocol: Safety, DNA yield, DNA quality/stability, cost, user friendliness, reliability, and time. To identify the optimum conditions for each stage of the protocol (cell harvest, lysis, purification, and precipitation), each was investigated separately, and an adaptation of the fast-boiling protocol was used for the remaining stages. A validation study was undertaken with the optimized protocol to assess its performance when conducted by a group of students in a classroom setting. The optimum protocol used an isotonic Lucozade Hydro Active Fitness Water (HAFW) mouthwash. Lysis was achieved using a TE (10 mM Tris-HCl, 1 mM EDTA, pH 8) + 1% Sodium Dodecyl Sulphate (SDS) buffer. Protein was then digested using Proteinase K (Qiagen Inc., UK) at 56°C for 10 min. The DNA was then precipitated with sodium chloride and absolute ethanol. This protocol achieved an increase in DNA yield using readily available equipment and reagents at a lower per capita cost and is simple to use. PMID:21567818

  20. Cardiovascular group

    NASA Technical Reports Server (NTRS)

    Blomqvist, Gunnar

    1989-01-01

    As a starting point, the group defined a primary goal of maintaining in flight a level of systemic oxygen transport capacity comparable to each individual's preflight upright baseline. The goal of maintaining capacity at preflight levels would seem to be a reasonable objective for several different reasons, including the maintenance of good health in general and the preservation of sufficient cardiovascular reserve capacity to meet operational demands. It is also important not to introduce confounding variables in whatever other physiological studies are being performed. A change in the level of fitness is likely to be a significant confounding variable in the study of many organ systems. The principal component of the in-flight cardiovascular exercise program should be large-muscle activity such as treadmill exercise. It is desirable that at least one session per week be monitored to assure maintenance of proper functional levels and to provide guidance for any adjustments of the exercise prescription. Appropriate measurements include evaluation of the heart-rate/workload or the heart-rate/oxygen-uptake relationship. Respiratory gas analysis is helpful by providing better opportunities to document relative workload levels from analysis of the interrelationships among VO2, VCO2, and ventilation. The committee felt that there is no clear evidence that any particular in-flight exercise regimen is protective against orthostatic hypotension during the early readaptation phase. Some group members suggested that maintenance of the lower body muscle mass and muscle tone may be helpful. There is also evidence that late in-flight interventions to reexpand blood volume to preflight levels are helpful in preventing or minimizing postflight orthostatic hypotension.

  1. Preparation and characterization of imogolite/DNA hybrid hydrogels.

    PubMed

    Jiravanichanun, Nattha; Yamamoto, Kazuya; Kato, Kenichi; Kim, Jungeun; Horiuchi, Shin; Yah, Weng-On; Otsuka, Hideyuki; Takahara, Atsushi

    2012-01-01

    Imogolite is one of the clay minerals contained in volcanic ash soils. The novel hybrid hydrogels were prepared from imogolite nanofibers and DNA by utilizing strong interaction between the aluminol groups on imogolite surface and phosphate groups of DNA. The hybrid hydrogels of imogolite and DNA were prepared in various feed ratios, and their physicochemical properties and molecular aggregation states were investigated in both dispersion and gel states. The maximum DNA content in the hybrid gels was shown in equivalent molar ratio of imogolite and DNA. The physical properties of the hybrid gels were changed by varying DNA blend ratios. In the dispersion state, the hybrid gels showed a fibrous structure of imogolite, whereas a continuous network structure was observed in pure imogolite, indicating that the hybrid with DNA enhanced the dispersion of imogolite. In the gel state, DNA and imogolite nanofibers formed a 3D network structure. PMID:22148683

  2. Mouse zygotes respond to severe sperm DNA damage by delaying paternal DNA replication and embryonic development.

    PubMed

    Gawecka, Joanna E; Marh, Joel; Ortega, Michael; Yamauchi, Yasuhiro; Ward, Monika A; Ward, W Steven

    2013-01-01

    Mouse zygotes do not activate apoptosis in response to DNA damage. We previously reported a unique form of inducible sperm DNA damage termed sperm chromatin fragmentation (SCF). SCF mirrors some aspects of somatic cell apoptosis in that the DNA degradation is mediated by reversible double strand breaks caused by topoisomerase 2B (TOP2B) followed by irreversible DNA degradation by a nuclease(s). Here, we created zygotes using spermatozoa induced to undergo SCF (SCF zygotes) and tested how they responded to moderate and severe paternal DNA damage during the first cell cycle. We found that the TUNEL assay was not sensitive enough to identify the breaks caused by SCF in zygotes in either case. However, paternal pronuclei in both groups stained positively for γH2AX, a marker for DNA damage, at 5 hrs after fertilization, just before DNA synthesis, while the maternal pronuclei were negative. We also found that both pronuclei in SCF zygotes with moderate DNA damage replicated normally, but paternal pronuclei in the SCF zygotes with severe DNA damage delayed the initiation of DNA replication by up to 12 hrs even though the maternal pronuclei had no discernable delay. Chromosomal analysis of both groups confirmed that the paternal DNA was degraded after S-phase while the maternal pronuclei formed normal chromosomes. The DNA replication delay caused a marked retardation in progression to the 2-cell stage, and a large portion of the embryos arrested at the G2/M border, suggesting that this is an important checkpoint in zygotic development. Those embryos that progressed through the G2/M border died at later stages and none developed to the blastocyst stage. Our data demonstrate that the zygote responds to sperm DNA damage through a non-apoptotic mechanism that acts by slowing paternal DNA replication and ultimately leads to arrest in embryonic development. PMID:23431372

  3. DNA Binding Hydroxyl Radical Probes

    PubMed Central

    Tang, Vicky J; Konigsfeld, Katie M; Aguilera, Joe A; Milligan, Jamie R

    2011-01-01

    The hydroxyl radical is the primary mediator of DNA damage by the indirect effect of ionizing radiation. It is a powerful oxidizing agent produced by the radiolysis of water and is responsible for a significant fraction of the DNA damage associated with ionizing radiation. There is therefore an interest in the development of sensitive assays for its detection. The hydroxylation of aromatic groups to produce fluorescent products has been used for this purpose. We have examined four different chromophores which produce fluorescent products when hydroxylated. Of these, the coumarin system suffers from the fewest disadvantages. We have therefore examined its behavior when linked to a cationic peptide ligand designed to bind strongly to DNA. PMID:22125376

  4. Mechanism of DNA loading by the DNA repair helicase XPD.

    PubMed

    Constantinescu-Aruxandei, Diana; Petrovic-Stojanovska, Biljana; Penedo, J Carlos; White, Malcolm F; Naismith, James H

    2016-04-01

    The xeroderma pigmentosum group D (XPD) helicase is a component of the transcription factor IIH complex in eukaryotes and plays an essential role in DNA repair in the nucleotide excision repair pathway. XPD is a 5' to 3' helicase with an essential iron-sulfur cluster. Structural and biochemical studies of the monomeric archaeal XPD homologues have aided a mechanistic understanding of this important class of helicase, but several important questions remain open. In particular, the mechanism for DNA loading, which is assumed to require large protein conformational change, is not fully understood. Here, DNA binding by the archaeal XPD helicase fromThermoplasma acidophilumhas been investigated using a combination of crystallography, cross-linking, modified substrates and biochemical assays. The data are consistent with an initial tight binding of ssDNA to helicase domain 2, followed by transient opening of the interface between the Arch and 4FeS domains, allowing access to a second binding site on helicase domain 1 that directs DNA through the pore. A crystal structure of XPD fromSulfolobus acidocaldiariusthat lacks helicase domain 2 has an otherwise unperturbed structure, emphasizing the stability of the interface between the Arch and 4FeS domains in XPD. PMID:26896802

  5. Quantitative DNA fiber mapping

    DOEpatents

    Gray, Joe W.; Weier, Heinz-Ulrich G.

    1998-01-01

    The present invention relates generally to the DNA mapping and sequencing technologies. In particular, the present invention provides enhanced methods and compositions for the physical mapping and positional cloning of genomic DNA. The present invention also provides a useful analytical technique to directly map cloned DNA sequences onto individual stretched DNA molecules.

  6. Characterization of group A Streptococcus strains recovered from Mexican children with pharyngitis by automated DNA sequencing of virulence-related genes: unexpectedly large variation in the gene (sic) encoding a complement-inhibiting protein.

    PubMed

    Mejia, L M; Stockbauer, K E; Pan, X; Cravioto, A; Musser, J M

    1997-12-01

    Sequence variation was studied in several target genes in 54 strains of group A Streptococcus (GAS) cultured from children with pharyngitis in Mexico City. Although 16 distinct emm alleles were identified, only 4 had not been previously described. Virtually all bacteria (31 of 33 [94%] with the streptococcal pyrogenic exotoxin gene (speA) had emm1-related, emm3, or emm6 alleles. The gene (sic) encoding an extracellular GAS protein that inhibits complement function was unusually variable among isolates with the emm1 family of alleles, with a total of seven variants identified. The data suggest that many GAS strains infecting Mexican children are genetically similar to organisms commonly encountered in the United States and western Europe. Sequence variation in the sic gene is useful for rapid differentiation among GAS isolates with the emm1 family of alleles. PMID:9399523

  7. Characterization of group A Streptococcus strains recovered from Mexican children with pharyngitis by automated DNA sequencing of virulence-related genes: unexpectedly large variation in the gene (sic) encoding a complement-inhibiting protein.

    PubMed Central

    Mejia, L M; Stockbauer, K E; Pan, X; Cravioto, A; Musser, J M

    1997-01-01

    Sequence variation was studied in several target genes in 54 strains of group A Streptococcus (GAS) cultured from children with pharyngitis in Mexico City. Although 16 distinct emm alleles were identified, only 4 had not been previously described. Virtually all bacteria (31 of 33 [94%] with the streptococcal pyrogenic exotoxin gene (speA) had emm1-related, emm3, or emm6 alleles. The gene (sic) encoding an extracellular GAS protein that inhibits complement function was unusually variable among isolates with the emm1 family of alleles, with a total of seven variants identified. The data suggest that many GAS strains infecting Mexican children are genetically similar to organisms commonly encountered in the United States and western Europe. Sequence variation in the sic gene is useful for rapid differentiation among GAS isolates with the emm1 family of alleles. PMID:9399523

  8. Group evaporation

    NASA Technical Reports Server (NTRS)

    Shen, Hayley H.

    1991-01-01

    Liquid fuel combustion process is greatly affected by the rate of droplet evaporation. The heat and mass exchanges between gas and liquid couple the dynamics of both phases in all aspects: mass, momentum, and energy. Correct prediction of the evaporation rate is therefore a key issue in engineering design of liquid combustion devices. Current analytical tools for characterizing the behavior of these devices are based on results from a single isolated droplet. Numerous experimental studies have challenged the applicability of these results in a dense spray. To account for the droplets' interaction in a dense spray, a number of theories have been developed in the past decade. Herein, two tasks are examined. One was to study how to implement the existing theoretical results, and the other was to explore the possibility of experimental verifications. The current theoretical results of group evaporation are given for a monodispersed cluster subject to adiabatic conditions. The time evolution of the fluid mechanic and thermodynamic behavior in this cluster is derived. The results given are not in the form of a subscale model for CFD codes.

  9. Binding of HIV-1 Vpr Protein to the Human Homolog of the Yeast DNA Repair Protein RAD23 (hHR23A) Requires Its Xeroderma Pigmentosum Complementation Group C Binding (XPCB) Domain as Well as the Ubiquitin-associated 2 (UBA2) Domain*

    PubMed Central

    Jung, Jinwon; Byeon, In-Ja L.; DeLucia, Maria; Koharudin, Leonardus M. I.; Ahn, Jinwoo; Gronenborn, Angela M.

    2014-01-01

    The human homolog of the yeast DNA repair protein RAD23, hHR23A, has been found previously to interact with the human immunodeficiency virus, type 1 accessory protein Vpr. hHR23A is a modular protein containing an N-terminal ubiquitin-like (UBL) domain and two ubiquitin-associated domains (UBA1 and UBA2) separated by a xeroderma pigmentosum complementation group C binding (XPCB) domain. All domains are connected by flexible linkers. hHR23A binds ubiquitinated proteins and acts as a shuttling factor to the proteasome. Here, we show that hHR23A utilizes both the UBA2 and XPCB domains to form a stable complex with Vpr, linking Vpr directly to cellular DNA repair pathways and their probable exploitation by the virus. Detailed structural mapping of the Vpr contacts on hHR23A, by NMR, revealed substantial contact surfaces on the UBA2 and XPCB domains. In addition, Vpr binding disrupts an intramolecular UBL-UBA2 interaction. We also show that Lys-48-linked di-ubiquitin, when binding to UBA1, does not release the bound Vpr from the hHR23A-Vpr complex. Instead, a ternary hHR23A·Vpr·di-UbK48 complex is formed, indicating that Vpr does not necessarily abolish hHR23A-mediated shuttling to the proteasome. PMID:24318982

  10. Photochemical modulation of DNA conformation by organic dications.

    PubMed

    Zinchenko, Anatoly A; Tanahashi, Mao; Murata, Shizuaki

    2012-01-01

    A group of azobenzene derivatives containing two quaternary ammonium groups with various intercharge distances between them was synthesised and used to control photochemically the conformation of genomic DNA by switching the distance between cationic ammonium groups in the dications. It was found that isomerisation of either dication from the trans form to cis resulted in an increase in the dication's efficiency for DNA compaction; this is associated with a decrease in intercharge distance between ammonium groups and leads to a better match of the binder's cationic groups to adjacent phosphate groups of DNA. Ammonium dications have several important advantages over the photosensitive surfactant type of diazobenzene reported earlier: they can be used at significantly lower (>100-fold) concentrations than photosensitive surfactants, and DNA conformation control can be performed over a broader concentration range of dications. The influence of intercharge distance in photosensitive dications on photo-induced DNA binding discrimination is discussed, and the molecular mechanism is proposed. PMID:22109974

  11. Melanesian mtDNA Complexity

    PubMed Central

    Friedlaender, Jonathan S.; Friedlaender, Françoise R.; Hodgson, Jason A.; Stoltz, Matthew; Koki, George; Horvat, Gisele; Zhadanov, Sergey; Schurr, Theodore G.; Merriwether, D. Andrew

    2007-01-01

    Melanesian populations are known for their diversity, but it has been hard to grasp the pattern of the variation or its underlying dynamic. Using 1,223 mitochondrial DNA (mtDNA) sequences from hypervariable regions 1 and 2 (HVR1 and HVR2) from 32 populations, we found the among-group variation is structured by island, island size, and also by language affiliation. The more isolated inland Papuan-speaking groups on the largest islands have the greatest distinctions, while shore dwelling populations are considerably less diverse (at the same time, within-group haplotype diversity is less in the most isolated groups). Persistent differences between shore and inland groups in effective population sizes and marital migration rates probably cause these differences. We also add 16 whole sequences to the Melanesian mtDNA phylogenies. We identify the likely origins of a number of the haplogroups and ancient branches in specific islands, point to some ancient mtDNA connections between Near Oceania and Australia, and show additional Holocene connections between Island Southeast Asia/Taiwan and Island Melanesia with branches of haplogroup E. Coalescence estimates based on synonymous transitions in the coding region suggest an initial settlement and expansion in the region at ∼30–50,000 years before present (YBP), and a second important expansion from Island Southeast Asia/Taiwan during the interval ∼3,500–8,000 YBP. However, there are some important variance components in molecular dating that have been overlooked, and the specific nature of ancestral (maternal) Austronesian influence in this region remains unresolved. PMID:17327912

  12. Functionalizing Designer DNA Crystals

    NASA Astrophysics Data System (ADS)

    Chandrasekaran, Arun Richard

    nucleotides is usually pH dependent (pH < 6) four different TFOs were examined: TFO-1 was unmodified while TFOs 2-4 contained additional stabilizing analogues capable of extending triplex formation to pH 7. In addition, each of the TFOs contained a Cy5 dye at the 5'-end of the oligonucleotide to aid in characterization of TFO binding - crystals were obtained with all four variations of TFOs. Formation of DNA triplex in the motif was characterized by an electrophoretic mobility shift assay (EMSA), UV melting studies and FRET. Crystals containing TFO-1 (unmodified) and TFO-2 (with 2'-amino ethoxy modification) were isolated and flash-frozen in liquid nitrogen for X-ray data collection at beam line NSLS-X25. X-ray data was also collected for crystals of the 3-turn triangle without any TFO bound to it. Difference maps were done between the crystals with TFO against the one without to identify any additional electron density corresponding to the third strand in the triplex binding region. The data from the crystal containing TFO-2 was used to further analyze if the additional density can match the expected position of the TFO on the triangle motif. Since the additional density did not correspond to the entire binding region, 2Fo-Fc, 3Fo-2Fc and 4Fo-3Fc maps were done to check for missing pieces of the electron density. From the resulting 2Fo-Fc map, the asymmetric unit from the 3-turn triangle (31-bp duplex model based on previous structure 3UBI) was inserted into the density as a reference. However, the electron density corresponding to the TFO was still not continuous throughout the 13-nt triplex binding region and allowed only a partial fit of the TFO. The third nucleotide in positions 1, 3, 4, 6, 7 were fit into the density in the major groove of the underlying duplex with proper triplex configuration. The third chapter describes the triplex approach to position a functional group (the UV cross-linking agent psoralen) within a pre-formed DNA motif. Triplex formation and

  13. Tumorigenic DNA viruses

    SciTech Connect

    Klein, G.

    1989-01-01

    The eighth volume of Advances in Viral Oncology focuses on the three major DNA virus groups with a postulated or proven tumorigenic potential: papillomaviruses, animal hepatitis viruses, and the Epstein-Bar virus. In the opening chapters, the contributors analyze the evidence that papillomaviruses and animal hepatitis viruses are involved in tumorigenesis and describe the mechanisms that trigger virus-host cell interactions. A detailed section on the Epstein-Barr virus (EBV) - comprising more than half the book - examines the transcription and mRNA processing patterns of the virus genome; the mechanisms by which EBV infects lymphoid and epithelial cells; the immunological aspects of the virus; the actions of EBV in hosts with Acquired Immune Deficiency Syndrome; and the involvement of EBV in the etiology of Burkitt's lymphoma.

  14. newDNA-Prot: Prediction of DNA-binding proteins by employing support vector machine and a comprehensive sequence representation.

    PubMed

    Zhang, Yanping; Xu, Jun; Zheng, Wei; Zhang, Chen; Qiu, Xingye; Chen, Ke; Ruan, Jishou

    2014-10-01

    Identification of DNA-binding proteins is essential in studying cellular activities as the DNA-binding proteins play a pivotal role in gene regulation. In this study, we propose newDNA-Prot, a DNA-binding protein predictor that employs support vector machine classifier and a comprehensive feature representation. The sequence representation are categorized into 6 groups: primary sequence based, evolutionary profile based, predicted secondary structure based, predicted relative solvent accessibility based, physicochemical property based and biological function based features. The mRMR, wrapper and two-stage feature selection methods are employed for removing irrelevant features and reducing redundant features. Experiments demonstrate that the two-stage method performs better than the mRMR and wrapper methods. We also perform a statistical analysis on the selected features and results show that more than 95% of the selected features are statistically significant and they cover all 6 feature groups. The newDNA-Prot method is compared with several state of the art algorithms, including iDNA-Prot, DNAbinder and DNA-Prot. The results demonstrate that newDNA-Prot method outperforms the iDNA-Prot, DNAbinder and DNA-Prot methods. More specific, newDNA-Prot improves the runner-up method, DNA-Prot for around 10% on several evaluation measures. The proposed newDNA-Prot method is available at http://sourceforge.net/projects/newdnaprot/ PMID:25240115

  15. Conserved Sequences at the Origin of Adenovirus DNA Replication

    PubMed Central

    Stillman, Bruce W.; Topp, William C.; Engler, Jeffrey A.

    1982-01-01

    The origin of adenovirus DNA replication lies within an inverted sequence repetition at either end of the linear, double-stranded viral DNA. Initiation of DNA replication is primed by a deoxynucleoside that is covalently linked to a protein, which remains bound to the newly synthesized DNA. We demonstrate that virion-derived DNA-protein complexes from five human adenovirus serological subgroups (A to E) can act as a template for both the initiation and the elongation of DNA replication in vitro, using nuclear extracts from adenovirus type 2 (Ad2)-infected HeLa cells. The heterologous template DNA-protein complexes were not as active as the homologous Ad2 DNA, most probably due to inefficient initiation by Ad2 replication factors. In an attempt to identify common features which may permit this replication, we have also sequenced the inverted terminal repeated DNA from human adenovirus serotypes Ad4 (group E), Ad9 and Ad10 (group D), and Ad31 (group A), and we have compared these to previously determined sequences from Ad2 and Ad5 (group C), Ad7 (group B), and Ad12 and Ad18 (group A) DNA. In all cases, the sequence around the origin of DNA replication can be divided into two structural domains: a proximal A · T-rich region which is partially conserved among these serotypes, and a distal G · C-rich region which is less well conserved. The G · C-rich region contains sequences similar to sequences present in papovavirus replication origins. The two domains may reflect a dual mechanism for initiation of DNA replication: adenovirus-specific protein priming of replication, and subsequent utilization of this primer by host replication factors for completion of DNA synthesis. Images PMID:7143575

  16. Molecular Mechanisms of DNA Polymerase Clamp Loaders

    NASA Astrophysics Data System (ADS)

    Kelch, Brian; Makino, Debora; Simonetta, Kyle; O'Donnell, Mike; Kuriyan, John

    Clamp loaders are ATP-driven multiprotein machines that couple ATP hydrolysis to the opening and closing of a circular protein ring around DNA. This ring-shaped clamp slides along DNA, and interacts with numerous proteins involved in DNA replication, DNA repair and cell cycle control. Recently determined structures of clamp loader complexes from prokaryotic and eukaryotic DNA polymerases have revealed exciting new details of how these complex AAA+ machines perform this essential clamp loading function. This review serves as background to John Kuriyan's lecture at the 2010 Erice School, and is not meant as a comprehensive review of the contributions of the many scientists who have advanced this field. These lecture notes are derived from recent reviews and research papers from our groups.

  17. LCAT DNA shearing.

    PubMed

    Okabe, Yuka; Lee, Abraham P

    2014-04-01

    We present a novel method to fragment DNA by using lateral cavity acoustic transducers (LCATs). DNA solution is placed within a microfluidic device containing LCATs. The LCATs cause microstreaming, which fragments DNA within the solution without any need for purification or downstream processing. The LCAT-based DNA fragmentation method offers an easy-to-use, low-cost, low-energy way to fragment DNA that is amenable to integration on microfluidic platforms to further automate DNA processing. Furthermore, the LCAT microdevice requires less than 10 µL of sample, and no external equipment is needed besides a piezoelectric transducer. PMID:23850863

  18. Chemical method for introducing haptens on to DNA probes

    SciTech Connect

    Keller, G.H.; Cumming, C.U.; Huang, D.P.; Manak, M.M.; Ting, R.

    1988-05-01

    The authors developed a versatile chemical method of attaching hapten moieties onto DNA, for the construction of nonisotopic DNA probes. The DNA is reacted with N-bromosuccinimide at alkaline pH, resulting in bromination of a fraction of the thymine, guanine, and cytosine residues, with adenine modified to a lesser extent. The bromine is subsequently displaced by a primary amino group, attached to a linker arm. The other end of the linker arm has a detectable group preattached to it. They have labeled cloned hepatitis B viral (HBV) DNA with the hapten 2,4-dinitrophenyl (DNP) and used it in combination with a high affinity rabbit anti-DNP antibody, for the detection of hepatitis B DNA by slot blotting. This probe was sensitive enough to specifically detect 1 x 10/sup -17/ mol (1 x 10/sup 6/ copies) of HBV DNA in total DNA from human serum.

  19. Effect of DNA type on response of DNA biosensor for carcinogens

    NASA Astrophysics Data System (ADS)

    Sani, Nor Diyana bt. Md.; Heng, Lee Yook; Surif, Salmijah; Lazim, Azwani Mat

    2013-11-01

    Carcinogens are cancer causing chemicals that can bind to DNA and cause damage to the DNA. These chemicals are available everywhere including in water, air, soil and food. Therefore, a sensor that can detect the presence of these chemicals will be a very useful tool. Since carcinogens bind to DNA, DNA can be used as the biological element in a biosensor. This study has utilized different types of DNA in a biosensor for carcinogen detection. The DNAs include double stranded calf thymus DNA, single stranded calf thymus DNA and guanine rich single stranded DNA. The modified SPE was exposed to a carcinogen followed by interaction with methylene blue which acts as the electroactive indicator. The SPE was then analysed using differential pulse voltammetry (DPV). Optimization studies were conducted for MB concentration and accumulation time, DNA concentration, as well as effect of buffer concentration, buffer pH and ionic strength. The performance of the biosensor was tested on a group 1 carcinogen, formaldehyde. The results indicated that the usage of guanine rich single stranded DNA also gives higher response as carcinogens prefer to bind with guanine compared to other bases.

  20. Crossing borders: the DNA of physics

    NASA Astrophysics Data System (ADS)

    Beijerinck, H. C. W.

    2015-01-01

    In cell culture, the physical environment plays an important role: "Everything is everywhere, but the environment selects"[1]. The education of physicists can be viewed within this framework. The Petri dish for the reproduction of physicists is a university research group. The full professor is its DNA. The selection process of new professors - new DNA - is a determining step in creating the right culture. [1] M.W. Beijerinck and L.G.M. Baas Becking, en.wikipedia.org.

  1. Structural Organization of DNA.

    ERIC Educational Resources Information Center

    Banfalvi, Gaspar

    1986-01-01

    Explains the structural organization of DNA by providing information on the primary, secondary, tertiary, and higher organization levels of the molecule. Also includes illustrations and descriptions of sign-inversion and rotating models for supercoiling of DNA. (ML)

  2. Modeling DNA Replication.

    ERIC Educational Resources Information Center

    Bennett, Joan

    1998-01-01

    Recommends the use of a model of DNA made out of Velcro to help students visualize the steps of DNA replication. Includes a materials list, construction directions, and details of the demonstration using the model parts. (DDR)

  3. HPV DNA test

    MedlinePlus

    The HPV DNA test is used to check for high-risk HPV infection in women. HPV infection around the genitals is ... warts spread when you have sex. The HPV-DNA test is generally not recommended for detecting low- ...

  4. DNA tagged microparticles

    DOEpatents

    Farquar, George Roy; Leif, Roald N; Wheeler, Elizabeth

    2015-05-05

    A simulant that includes a carrier and DNA encapsulated in the carrier. Also a method of making a simulant including the steps of providing a carrier and encapsulating DNA in the carrier to produce the simulant.

  5. Oxidation of DNA: damage to nucleobases.

    PubMed

    Kanvah, Sriram; Joseph, Joshy; Schuster, Gary B; Barnett, Robert N; Cleveland, Charles L; Landman, Uzi

    2010-02-16

    All organisms store the information necessary to maintain life in their DNA. Any process that damages DNA, causing a loss or corruption of that information, jeopardizes the viability of the organism. One-electron oxidation is such a process. In this Account, we address three of the central features of one-electron oxidation of DNA: (i) the migration of the radical cation away from the site of its formation; (ii) the electronic and structural factors that determine the nucleobases at which irreversible reactions most readily occur; (iii) the mechanism of reaction for nucleobase radical cations. The loss of an electron (ionization) from DNA generates an electron "hole" (a radical cation), located most often on its nucleobases, that migrates reversibly through duplex DNA by hopping until it is trapped in an irreversible chemical reaction. The particular sequence of nucleobases in a DNA oligomer determines both the efficiency of hopping and the specific location and nature of the damaging chemical reaction. In aqueous solution, DNA is a polyanion because of the negative charge carried by its phosphate groups. Counterions to the phosphate groups (typically Na(+)) play an important role in facilitating both hopping and the eventual reaction of the radical cation with H(2)O. Irreversible reaction of a radical cation with H(2)O in duplex DNA occurs preferentially at the most reactive site. In normal DNA, comprising the four common DNA nucleobases G, C, A, and T, reaction occurs most commonly at a guanine, resulting in its conversion primarily to 8-oxo-7,8-dihydroguanine (8-OxoG). Both electronic and steric effects control the outcome of this process. If the DNA oligomer does not contain a suitable guanine, then reaction of the radical cation occurs at the thymine of a TT step, primarily by a tandem process. The oxidative damage of DNA is a complex process, influenced by charge transport and reactions that are controlled by a combination of enthalpic, entropic, steric, and

  6. The logic of DNA replication in double-stranded DNA viruses: insights from global analysis of viral genomes

    PubMed Central

    Kazlauskas, Darius; Krupovic, Mart; Venclovas, Česlovas

    2016-01-01

    Genomic DNA replication is a complex process that involves multiple proteins. Cellular DNA replication systems are broadly classified into only two types, bacterial and archaeo-eukaryotic. In contrast, double-stranded (ds) DNA viruses feature a much broader diversity of DNA replication machineries. Viruses differ greatly in both completeness and composition of their sets of DNA replication proteins. In this study, we explored whether there are common patterns underlying this extreme diversity. We identified and analyzed all major functional groups of DNA replication proteins in all available proteomes of dsDNA viruses. Our results show that some proteins are common to viruses infecting all domains of life and likely represent components of the ancestral core set. These include B-family polymerases, SF3 helicases, archaeo-eukaryotic primases, clamps and clamp loaders of the archaeo-eukaryotic type, RNase H and ATP-dependent DNA ligases. We also discovered a clear correlation between genome size and self-sufficiency of viral DNA replication, the unanticipated dominance of replicative helicases and pervasive functional associations among certain groups of DNA replication proteins. Altogether, our results provide a comprehensive view on the diversity and evolution of replication systems in the DNA virome and uncover fundamental principles underlying the orchestration of viral DNA replication. PMID:27112572

  7. The logic of DNA replication in double-stranded DNA viruses: insights from global analysis of viral genomes.

    PubMed

    Kazlauskas, Darius; Krupovic, Mart; Venclovas, Česlovas

    2016-06-01

    Genomic DNA replication is a complex process that involves multiple proteins. Cellular DNA replication systems are broadly classified into only two types, bacterial and archaeo-eukaryotic. In contrast, double-stranded (ds) DNA viruses feature a much broader diversity of DNA replication machineries. Viruses differ greatly in both completeness and composition of their sets of DNA replication proteins. In this study, we explored whether there are common patterns underlying this extreme diversity. We identified and analyzed all major functional groups of DNA replication proteins in all available proteomes of dsDNA viruses. Our results show that some proteins are common to viruses infecting all domains of life and likely represent components of the ancestral core set. These include B-family polymerases, SF3 helicases, archaeo-eukaryotic primases, clamps and clamp loaders of the archaeo-eukaryotic type, RNase H and ATP-dependent DNA ligases. We also discovered a clear correlation between genome size and self-sufficiency of viral DNA replication, the unanticipated dominance of replicative helicases and pervasive functional associations among certain groups of DNA replication proteins. Altogether, our results provide a comprehensive view on the diversity and evolution of replication systems in the DNA virome and uncover fundamental principles underlying the orchestration of viral DNA replication. PMID:27112572

  8. Is DNA a language?

    PubMed

    Tsonis, A A; Elsner, J B; Tsonis, P A

    1997-01-01

    DNA sequences usually involve local construction rules that affect different scales. As such their "dictionary" may not follow Zipf's law (a power law) which is followed in every natural language. Indeed, analysis of many DNA sequences suggests that no linguistics connections to DNA exist and that even though it has structure DNA is not a language. Computer simulations and a biological approach to this problem further support these results. PMID:9039397

  9. Photoelectrochemical synthesis of DNA microarrays

    PubMed Central

    Chow, Brian Y.; Emig, Christopher J.; Jacobson, Joseph M.

    2009-01-01

    Optical addressing of semiconductor electrodes represents a powerful technology that enables the independent and parallel control of a very large number of electrical phenomena at the solid-electrolyte interface. To date, it has been used in a wide range of applications including electrophoretic manipulation, biomolecule sensing, and stimulating networks of neurons. Here, we have adapted this approach for the parallel addressing of redox reactions, and report the construction of a DNA microarray synthesis platform based on semiconductor photoelectrochemistry (PEC). An amorphous silicon photoconductor is activated by an optical projection system to create virtual electrodes capable of electrochemically generating protons; these PEC-generated protons then cleave the acid-labile dimethoxytrityl protecting groups of DNA phosphoramidite synthesis reagents with the requisite spatial selectivity to generate DNA microarrays. Furthermore, a thin-film porous glass dramatically increases the amount of DNA synthesized per chip by over an order of magnitude versus uncoated glass. This platform demonstrates that PEC can be used toward combinatorial bio-polymer and small molecule synthesis. PMID:19706433

  10. DNAzymes in DNA Nanomachines and DNA Analysis

    NASA Astrophysics Data System (ADS)

    He, Yu; Tian, Ye; Chen, Yi; Mao, Chengde

    This chapter discusses our efforts in using DNAzymes in DNA nano-machines and DNA analysis systems. 10-23 DNAzymes can cleave specific phos-phodiester bonds in RNA. We use them to construct an autonomous DNA-RNA chimera nanomotor, which constantly extracts chemical energy from RNA substrates and transduces the energy into a mechanical motion: cycles of contraction and extension. The motor's motion can be reversibly turned on and off by a DNA analogue (brake) of the RNA substrate. Addition and removal of the brake stops and restarts, respectively, the motor's motion. Furthermore, when the RNA substrates are preorganized into a one-dimensional track, a DNAzyme can continuously move along the track so long as there are substrates available ahead. Based on a similar mechanism, a novel DNA detection system has been developed. A target DNA activates a DNAzyme to cleave RNA-containing molecular beacons (MB), which generates an enhanced fluorescence signal. A following work integrates two steps of signal amplifications: a rolling-circle amplification (RCA) to synthesize multiple copies of DNAzymes, and the DNAzymes catalyze a chemical reaction to generate a colorimetric signal. This method allows detection of DNA analytes whose concentration is as low as 1 pM.

  11. The Many Sides of DNA.

    ERIC Educational Resources Information Center

    Flannery, Maura C.

    1997-01-01

    Explores the meaning of DNA. Discusses histories of DNA, literature on DNA, the contributions of Max Delbruck and Barbara McClintock, life, views of control, current research, and the language of DNA. Contains 24 references. (JRH)

  12. Small Molecules, Inhibitors of DNA-PK, Targeting DNA Repair, and Beyond

    PubMed Central

    Davidson, David; Amrein, Lilian; Panasci, Lawrence; Aloyz, Raquel

    2012-01-01

    design will not only assist in identifying novel functional moieties to replace the metabolically labile morpholino group but will also facilitate the design of molecules to target the DNA-PKcs/Ku80 interface or one of the autophosphorylation sites. PMID:23386830

  13. Synthesis of site-specific DNA-protein conjugates and their effects on DNA replication.

    PubMed

    Yeo, Jung Eun; Wickramaratne, Susith; Khatwani, Santoshkumar; Wang, Yen-Chih; Vervacke, Jeffrey; Distefano, Mark D; Tretyakova, Natalia Y

    2014-08-15

    DNA-protein cross-links (DPCs) are bulky, helix-distorting DNA lesions that form in the genome upon exposure to common antitumor drugs, environmental/occupational toxins, ionizing radiation, and endogenous free-radical-generating systems. As a result of their considerable size and their pronounced effects on DNA-protein interactions, DPCs can interfere with DNA replication, transcription, and repair, potentially leading to mutagenesis, genotoxicity, and cytotoxicity. However, the biological consequences of these ubiquitous lesions are not fully understood due to the difficulty of generating DNA substrates containing structurally defined, site-specific DPCs. In the present study, site-specific cross-links between the two biomolecules were generated by copper-catalyzed [3 + 2] Huisgen cycloaddition (click reaction) between an alkyne group from 5-(octa-1,7-diynyl)-uracil in DNA and an azide group within engineered proteins/polypeptides. The resulting DPC substrates were subjected to in vitro primer extension in the presence of human lesion bypass DNA polymerases η, κ, ν, and ι. We found that DPC lesions to the green fluorescent protein and a 23-mer peptide completely blocked DNA replication, while the cross-link to a 10-mer peptide was bypassed. These results indicate that the polymerases cannot read through the larger DPC lesions and further suggest that proteolytic degradation may be required to remove the replication block imposed by bulky DPC adducts. PMID:24918113

  14. DNA Sequencing apparatus

    DOEpatents

    Tabor, Stanley; Richardson, Charles C.

    1992-01-01

    An automated DNA sequencing apparatus having a reactor for providing at least two series of DNA products formed from a single primer and a DNA strand, each DNA product of a series differing in molecular weight and having a chain terminating agent at one end; separating means for separating the DNA products to form a series bands, the intensity of substantially all nearby bands in a different series being different, band reading means for determining the position an This invention was made with government support including a grant from the U.S. Public Health Service, contract number AI-06045. The U.S. government has certain rights in the invention.

  15. Functionalizing Designer DNA Crystals

    NASA Astrophysics Data System (ADS)

    Chandrasekaran, Arun Richard

    nucleotides is usually pH dependent (pH < 6) four different TFOs were examined: TFO-1 was unmodified while TFOs 2-4 contained additional stabilizing analogues capable of extending triplex formation to pH 7. In addition, each of the TFOs contained a Cy5 dye at the 5'-end of the oligonucleotide to aid in characterization of TFO binding - crystals were obtained with all four variations of TFOs. Formation of DNA triplex in the motif was characterized by an electrophoretic mobility shift assay (EMSA), UV melting studies and FRET. Crystals containing TFO-1 (unmodified) and TFO-2 (with 2'-amino ethoxy modification) were isolated and flash-frozen in liquid nitrogen for X-ray data collection at beam line NSLS-X25. X-ray data was also collected for crystals of the 3-turn triangle without any TFO bound to it. Difference maps were done between the crystals with TFO against the one without to identify any additional electron density corresponding to the third strand in the triplex binding region. The data from the crystal containing TFO-2 was used to further analyze if the additional density can match the expected position of the TFO on the triangle motif. Since the additional density did not correspond to the entire binding region, 2Fo-Fc, 3Fo-2Fc and 4Fo-3Fc maps were done to check for missing pieces of the electron density. From the resulting 2Fo-Fc map, the asymmetric unit from the 3-turn triangle (31-bp duplex model based on previous structure 3UBI) was inserted into the density as a reference. However, the electron density corresponding to the TFO was still not continuous throughout the 13-nt triplex binding region and allowed only a partial fit of the TFO. The third nucleotide in positions 1, 3, 4, 6, 7 were fit into the density in the major groove of the underlying duplex with proper triplex configuration. The third chapter describes the triplex approach to position a functional group (the UV cross-linking agent psoralen) within a pre-formed DNA motif. Triplex formation and

  16. DNA barcoding amphibians and reptiles.

    PubMed

    Vences, Miguel; Nagy, Zoltán T; Sonet, Gontran; Verheyen, Erik

    2012-01-01

    Only a few major research programs are currently targeting COI barcoding of amphibians and reptiles (including chelonians and crocodiles), two major groups of tetrapods. Amphibian and reptile species are typically old, strongly divergent, and contain deep conspecific lineages which might lead to problems in species assignment with incomplete reference databases. As far as known, there is no single pair of COI primers that will guarantee a sufficient rate of success across all amphibian and reptile taxa, or within major subclades of amphibians and reptiles, which means that the PCR amplification strategy needs to be adjusted depending on the specific research question. In general, many more amphibian and reptile taxa have been sequenced for 16S rDNA, which for some purposes may be a suitable complementary marker, at least until a more comprehensive COI reference database becomes available. DNA barcoding has successfully been used to identify amphibian larval stages (tadpoles) in species-rich tropical assemblages. Tissue sampling, DNA extraction, and amplification of COI is straightforward in amphibians and reptiles. Single primer pairs are likely to have a failure rate between 5 and 50% if taxa of a wide taxonomic range are targeted; in such cases the use of primer cocktails or subsequent hierarchical usage of different primer pairs is necessary. If the target group is taxonomically limited, many studies have followed a strategy of designing specific primers which then allow an easy and reliable amplification of all samples. PMID:22684953

  17. Sequence-specific DNA nicking endonucleases.

    PubMed

    Xu, Shuang-yong

    2015-08-01

    A group of small HNH nicking endonucleases (NEases) was discovered recently from phage or prophage genomes that nick double-stranded DNA sites ranging from 3 to 5 bp in the presence of Mg2+ or Mn2+. The cosN site of phage HK97 contains a gp74 nicking site AC↑CGC, which is similar to AC↑CGR (R=A/G) of N.ϕGamma encoded by Bacillus phage Gamma. A minimal nicking domain of 76 amino acid residues from N.ϕGamma could be fused to other DNA binding partners to generate chimeric NEases with new specificities. The biological roles of a few small HNH endonucleases (HNHE, gp74 of HK97, gp37 of ϕSLT, ϕ12 HNHE) have been demonstrated in phage and pathogenicity island DNA packaging. Another group of NEases with 3- to 7-bp specificities are either natural components of restriction systems or engineered from type IIS restriction endonucleases. A phage group I intron-encoded HNH homing endonucleases, I-PfoP3I was found to nick DNA sites of 14-16 bp. I-TslI encoded by T7-like ΦI appeared to nick DNA sites with a 9-bp core sequence. DNA nicking and labeling have been applied to optical mapping to aid genome sequence assembly and detection of large insertion/deletion mutations in genomic DNA of cancer cells. Nicking enzyme-mediated amplification reaction has been applied to rapid diagnostic testing of influenza A and B in clinical setting and for construction of DNA-based Boolean logic gates. The clustered regularly interspaced short palindromic repeats-ribonucleoprotein complex consisting of engineered Cas9 nickases in conjunction with tracerRNA:crRNA or a single-guide RNA have been successfully used in genome modifications. PMID:26352356

  18. DNA in Nanoscale Electronics

    NASA Astrophysics Data System (ADS)

    Slinker, Jason

    2012-10-01

    DNA, the quintessential molecule of life, possesses a number of attractive properties for use in nanoscale circuits. Charge transport (CT) through DNA itself is of both fundamental and practical interest. Fundamentally, DNA has a unique configuration of π-stacked bases in a well ordered, double helical structure. Given its unparalleled importance to life processes and its arrangement of conjugated subunits, DNA has been a compelling target of conductivity studies. In addition, further understanding of DNA CT will elucidate the biological implications of this process and advance its use in sensing technologies. We have investigated the fundamentals of DNA CT by measuring the electrochemistry of DNA monolayers under biologically-relevant conditions. We have uncovered both fundamental kinetic parameters to distinguish between competing models of operation as well as the practical implications of DNA CT for sensing. Furthermore, we are leveraging our studies of DNA conductivity for the manufacture of nanoscale circuits. We are investigating the electrical properties and self-assembly of DNA nanowires containing artificial base pair surrogates, which can be prepared through low cost and high throughput automated DNA synthesis. This unique and economically viable approach will establish a new paradigm for the scalable manufacture of nanoscale semiconductor devices.

  19. DNA structure and function.

    PubMed

    Travers, Andrew; Muskhelishvili, Georgi

    2015-06-01

    The proposal of a double-helical structure for DNA over 60 years ago provided an eminently satisfying explanation for the heritability of genetic information. But why is DNA, and not RNA, now the dominant biological information store? We argue that, in addition to its coding function, the ability of DNA, unlike RNA, to adopt a B-DNA structure confers advantages both for information accessibility and for packaging. The information encoded by DNA is both digital - the precise base specifying, for example, amino acid sequences - and analogue. The latter determines the sequence-dependent physicochemical properties of DNA, for example, its stiffness and susceptibility to strand separation. Most importantly, DNA chirality enables the formation of supercoiling under torsional stress. We review recent evidence suggesting that DNA supercoiling, particularly that generated by DNA translocases, is a major driver of gene regulation and patterns of chromosomal gene organization, and in its guise as a promoter of DNA packaging enables DNA to act as an energy store to facilitate the passage of translocating enzymes such as RNA polymerase. PMID:25903461

  20. DNA binding and recognition by binuclear transition metal complexes

    NASA Astrophysics Data System (ADS)

    Liu, Changlin; Yan, Rui; Xu, Yan; Yu, Siwang; Liao, Zhanru; Li, Dongfeng; Xu, Hui-Bie F.

    2001-09-01

    The development of small molecules that can bind and recognize DNA with sequence- or stereo-specificity under physiological conditions has been attracting a great interest in chemistry and biochemistry. Here, spectroscopic characterization and gel electrophoresis methods have been utilized to investigate the DNA binding and recognition by a variety of binuclear transition metal complexes. The result indicate that the structures and charges of binuclear transition metal complexes, compositions of coordination spheres, central metal ions and their coordination unsaturation, and separations between two central metal atoms can exert significant effects on the DNA binding and recognition. If there are not intercalative ligands into DNA base pairs or kinetically substitutable ligands by DNA phosphate groups within coordination sphere, the coordination saturation and compact binuclear transition metal complexes weaker bind to DNA than the coordination unsaturation and extended ones to DNA. Since the different transtiometal ions exhibit different affinities to DNA phosphate oxygen atoms, the binding interactions between their binuclear complexes and DNA are controlled by the affinity. He binuclear complexes with one or more negative charges lead to a consequence that they can not efficient associate with DNA, because DNA phosphodiester backbone is negatively charged. Whenthe separations between two central transition metal atoms is more than the distance between two DNA base pairs, the binuclear complexes could bind and recognize the DNA sequence with two or more base pairs. The protonated and positively charged ligands can strengthen the DNA binding and recognition by these binuclear metal complexes. Based on such DNA binding and recognition principles, the binuclear zinc complex designed in the study preferentially bind and recognize the following DNA sequence on pBR322 DNA with binding constant K.

  1. Human DNA repair genes.

    PubMed

    Wood, R D; Mitchell, M; Sgouros, J; Lindahl, T

    2001-02-16

    Cellular DNA is subjected to continual attack, both by reactive species inside cells and by environmental agents. Toxic and mutagenic consequences are minimized by distinct pathways of repair, and 130 known human DNA repair genes are described here. Notable features presently include four enzymes that can remove uracil from DNA, seven recombination genes related to RAD51, and many recently discovered DNA polymerases that bypass damage, but only one system to remove the main DNA lesions induced by ultraviolet light. More human DNA repair genes will be found by comparison with model organisms and as common folds in three-dimensional protein structures are determined. Modulation of DNA repair should lead to clinical applications including improvement of radiotherapy and treatment with anticancer drugs and an advanced understanding of the cellular aging process. PMID:11181991

  2. DNA Repair Gene Polymorphisms and Their Relation With DNA Damage, DNA Repair, and Total Antioxidant Capacity in Childhood Acute Lymphoblastic Leukemia Survivors.

    PubMed

    Dincer, Yildiz; Yüksel, Selin; Batar, Bahadir; Güven, Mehmet; Onaran, Ilhan; Celkan, Tiraje

    2015-07-01

    Oxidative stress and defective DNA repair are major contributory factors in the initiation and progression of carcinogenesis. Chemotherapy and radiotherapy cause oxidative DNA damage, consume antioxidant capacity, and impair DNA repair activity. These effects of chemotherapy and radiotherapy may be contributory factors in the development of secondary malignancy in cancer survivors. Basal, H2O2-induced, and postrepair DNA damage; urinary 8-hydroxydeoxyguanosine level as a marker of oxidatively damaged DNA; and serum total antioxidant capacity were measured; XPD Lys751Gln, XRCC1 Arg399Gln, and XRCC1 Arg194Trp polymorphisms were analyzed in childhood acute lymphoblastic leukemia (ALL) survivors. Basal and H2O2-induced DNA damage were found to be higher in the ALL survivor group versus the control group, however, there was no significant difference between the other parameters. No association was found between the examined parameters and polymorphisms of XPD 751 and XRCC1 399 and both the groups. XRCC1 194Trp allele was found to be associated with a low level of postrepair DNA damage in the ALL survivors. In conclusion, basal DNA damage and susceptibility to oxidation are high in childhood ALL survivors. This situation which may easily lead to occurrence of a secondary cancer does not seem to be a result of deficient DNA repair. PMID:24577548

  3. Photocross-linking of an oriented DNA repair complex. Ku bound at a single DNA end.

    PubMed

    Yoo, S; Kimzey, A; Dynan, W S

    1999-07-01

    Ku protein binds broken DNA ends, triggering a double-strand DNA break repair pathway. The spatial arrangement of the two Ku subunits in the initial Ku-DNA complex, when the Ku protein first approaches the broken DNA end, is not well defined. We have investigated the geometry of the complex using a novel set of photocross-linking probes that force Ku protein to be constrained in position and orientation, relative to a single free DNA end. Results suggest that this complex is roughly symmetric and that both Ku subunits make contact with an approximately equal area of the DNA. The complex has a strongly preferred orientation, with Ku70-DNA backbone contacts located proximal and Ku80-DNA backbone contacts located distal to the free end. Ku70 also contacts functional groups in the major groove proximal to the free end. Ku80 apparently does not make major groove contacts. Results are consistent with a model where the Ku70 and Ku80 subunits contact the major and minor grooves of DNA, respectively. PMID:10391954

  4. Impacts of degraded DNA on restriction enzyme associated DNA sequencing (RADSeq).

    PubMed

    Graham, Carly F; Glenn, Travis C; McArthur, Andrew G; Boreham, Douglas R; Kieran, Troy; Lance, Stacey; Manzon, Richard G; Martino, Jessica A; Pierson, Todd; Rogers, Sean M; Wilson, Joanna Y; Somers, Christopher M

    2015-11-01

    Degraded DNA from suboptimal field sampling is common in molecular ecology. However, its impact on techniques that use restriction site associated next-generation DNA sequencing (RADSeq, GBS) is unknown. We experimentally examined the effects of in situDNA degradation on data generation for a modified double-digest RADSeq approach (3RAD). We generated libraries using genomic DNA serially extracted from the muscle tissue of 8 individual lake whitefish (Coregonus clupeaformis) following 0-, 12-, 48- and 96-h incubation at room temperature posteuthanasia. This treatment of the tissue resulted in input DNA that ranged in quality from nearly intact to highly sheared. All samples were sequenced as a multiplexed pool on an Illumina MiSeq. Libraries created from low to moderately degraded DNA (12-48 h) performed well. In contrast, the number of RADtags per individual, number of variable sites, and percentage of identical RADtags retained were all dramatically reduced when libraries were made using highly degraded DNA (96-h group). This reduction in performance was largely due to a significant and unexpected loss of raw reads as a result of poor quality scores. Our findings remained consistent after changes in restriction enzymes, modified fold coverage values (2- to 16-fold), and additional read-length trimming. We conclude that starting DNA quality is an important consideration for RADSeq; however, the approach remains robust until genomic DNA is extensively degraded. PMID:25783180

  5. Comparative studies of UV-induced DNA cleavage by structural isomers of an iodinated DNA ligand

    SciTech Connect

    Martin, R.F.; Green, A.; Denison, L.; Pardee, M.; Kelly, D.P.; Roberts, M.; Rose, M.; Reum, M.

    1994-06-15

    The purpose was to evaluate the importance of the position of the halogen atom in iodinated DNA-binding bibenzimidazoles, with respect to sensitization of UV-A-induced DNA breakage. Three analogues of iodoHoechst 33258, denoted ortho-, meta- and paraiodoHoechst, according to the site of iodine substitution, were synthesized. Plasmid DNA (pBR322) was used to assay UV-A-induced DNA single-strand breaks (ssbs). The location of the sites of strand breakage was determined by DNA sequencing gel analysis, using a [sup 32]P-endlabelled oligoDNA with a single binding site for the ligands. A clear trend in decreasing activity of sensitization of UV-induced DNA ssbs was established: Ortho- > meta-, para- > iodoHoechst 33258. The sequencing gel studies showed that orthoiodoHoechst was distinct from the other three compounds, with respect to the sites of DNA strand breakage and the chemistry of the cleavage reaction. The position of iodine substitution in iodinated bibenzimidazoles determines the location of the carbon-centered radical on the ligand in the minor groove of DNA. DNA strand cleavage is mediated by abstraction of a nearby deoxyribosyl H-atom. Hence, the position of the radical species determines: which deoxyribosyl group is attacked (i.e., site of cleavage relative to the ligand binding site); which H-atom is abstracted, more specifically which of the five deoxyribosyl carbons is involved (i.e., the chemistry of the cleavage reaction), and the stereochemistry of the transition state for the H-atom abstraction (and hence the efficiency or extent of strand breakage). The ortho-compound represents the best example to date of iodinated DNA ligands designed as potential radiation sensitizers, as an extension of the well-established sensitization by halogenated DNA precursors. 30 refs., 3 figs.

  6. DNA hybridization activity of single-stranded DNA-conjugated gold nanoparticles used as probes for DNA detection

    NASA Astrophysics Data System (ADS)

    Kira, Atsushi; Matsuo, Kosuke; Nakajima, Shin-ichiro

    2016-02-01

    Colloidal nanoparticles (NPs) have potential applications in bio-sensing technologies as labels or signal enhancers. In order to meet demands for a development of biomolecular assays by a quantitative understanding of single-molecule, it is necessary to regulate accuracy of the NPs probes modified with biomolecules to optimize the characteristics of NPs. However, to our knowledge, there is little information about the structural effect of conjugated biomolecules to the NPs. In this study, we investigated the contribution of a density of single-stranded DNA (ssDNA) conjugating gold NP to hybridization activity. Hybridization activity decreased in accordance with increases in the density of attached ssDNAs, likely due to electrostatic repulsion generated by negatively charged phosphate groups in the ssDNA backbone. These results highlight the importance of controlling the density of ssDNAs attached to the surface of NPs used as DNA detection probes.

  7. DNA methylation in plants.

    PubMed

    Vanyushin, B F

    2006-01-01

    DNA in plants is highly methylated, containing 5-methylcytosine (m5C) and N6-methyladenine (m6A); m5C is located mainly in symmetrical CG and CNG sequences but it may occur also in other non-symmetrical contexts. m6A but not m5C was found in plant mitochondrial DNA. DNA methylation in plants is species-, tissue-, organelle- and age-specific. It is controlled by phytohormones and changes on seed germination, flowering and under the influence of various pathogens (viral, bacterial, fungal). DNA methylation controls plant growth and development, with particular involvement in regulation of gene expression and DNA replication. DNA replication is accompanied by the appearance of under-methylated, newly formed DNA strands including Okazaki fragments; asymmetry of strand DNA methylation disappears until the end of the cell cycle. A model for regulation of DNA replication by methylation is suggested. Cytosine DNA methylation in plants is more rich and diverse compared with animals. It is carried out by the families of specific enzymes that belong to at least three classes of DNA methyltransferases. Open reading frames (ORF) for adenine DNA methyltransferases are found in plant and animal genomes, and a first eukaryotic (plant) adenine DNA methyltransferase (wadmtase) is described; the enzyme seems to be involved in regulation of the mitochondria replication. Like in animals, DNA methylation in plants is closely associated with histone modifications and it affects binding of specific proteins to DNA and formation of respective transcription complexes in chromatin. The same gene (DRM2) in Arabidopsis thaliana is methylated both at cytosine and adenine residues; thus, at least two different, and probably interdependent, systems of DNA modification are present in plants. Plants seem to have a restriction-modification (R-M) system. RNA-directed DNA methylation has been observed in plants; it involves de novo methylation of almost all cytosine residues in a region of siRNA-DNA

  8. Risk stratification based on change in plasma Epstein-Barr virus DNA load after treatment in nasopharyngeal carcinoma

    PubMed Central

    Mao, Yan-Ping; Guo, Rui; Tang, Ling-Long; Peng, Hao; Sun, Ying; Liu, Qing; Chen, Lei; Ma, Jun

    2016-01-01

    Background: Nasopharyngeal carcinoma is associated with Epstein-Barr virus (EBV). The current study investigated change in the plasma EBV DNA load in the first 3 months after treatment and its clinical significance in NPC. Methods: A total of 273 patients with non-metastatic, histologically-proven NPC treated with radiotherapy or chemoradiotherapy were retrospectively reviewed. Results: EBV DNA was detectable in 19/273 (7.0%) patients at the end of therapy (end-DNA). Three months later, 16/273 (5.9%) patients had detectable EBV DNA (3-month-DNA). To investigate risk stratified by the pattern of change in post-treatment EBV-DNA, we divided patients into four subgroups: Group 1, undetectable end-DNA and 3-month-DNA (n = 244); Group 2, detectable end-DNA and undetectable 3-month-DNA (n = 13); Group 3, undetectable end-DNA and detectable 3-month-DNA (n = 7); and Group 4, detectable end-DNA and 3-month-DNA (n = 2). Patients with delayed remission of EBV DNA after treatment (Group 2) had significantly poorer 3-year DFS (48.6% vs. 89.7%, P < 0.001), DMFS (48.6% vs. 94.6%, P < 0.001) and OS (91.7% vs. 97.5%, P < 0.001) than those with persistently undetectable EBV DNA post-treatment (Group 1). Five of the seven patients with re-emergent EBV DNA (Group 3) and both patients with persistent EBV DNA post-treatment (Group 4) developed disease failure. Conclusion: Plasma EBV DNA load continues to change during the first 3 months after treatment. The pattern of change in EBV DNA load post-treatment could help identify patients with different prognoses. PMID:26840023

  9. Syndromes associated with mitochondrial DNA depletion

    PubMed Central

    2014-01-01

    Mitochondrial dysfunction accounts for a large group of inherited metabolic disorders most of which are due to a dysfunctional mitochondrial respiratory chain (MRC) and, consequently, deficient energy production. MRC function depends on the coordinated expression of both nuclear (nDNA) and mitochondrial (mtDNA) genomes. Thus, mitochondrial diseases can be caused by genetic defects in either the mitochondrial or the nuclear genome, or in the cross-talk between the two. This impaired cross-talk gives rise to so-called nuclear-mitochondrial intergenomic communication disorders, which result in loss or instability of the mitochondrial genome and, in turn, impaired maintenance of qualitative and quantitative mtDNA integrity. In children, most MRC disorders are associated with nuclear gene defects rather than alterations in the mtDNA itself. The mitochondrial DNA depletion syndromes (MDSs) are a clinically heterogeneous group of disorders with an autosomal recessive pattern of transmission that have onset in infancy or early childhood and are characterized by a reduced number of copies of mtDNA in affected tissues and organs. The MDSs can be divided into least four clinical presentations: hepatocerebral, myopathic, encephalomyopathic and neurogastrointestinal. The focus of this review is to offer an overview of these syndromes, listing the clinical phenotypes, together with their relative frequency, mutational spectrum, and possible insights for improving diagnostic strategies. PMID:24708634

  10. Mitochondrial DNA hypomethylation in chrome plating workers.

    PubMed

    Yang, Linqing; Xia, Bo; Yang, Xueqin; Ding, Hong; Wu, Desheng; Zhang, Huimin; Jiang, Gaofeng; Liu, Jianjun; Zhuang, Zhixiong

    2016-01-22

    A matched case-control study was conducted to examine the relationship between chromium (Cr) exposure and variation in mitochondrial (mt) DNA methylation. We enrolled 29 pairs of subjects in this study; Cr exposure was confirmed in the cases by detecting blood Cr and other metal ion concentrations. DNA damage caused by Cr exposure was determined in terms of binucleated micronucleus frequency (BNMN) and mtDNA copy number. Finally, a Sequenom MassARRAY platform was applied to inspect the DNA methylation levels of mitochondrially encoded tRNA phenylalanine (MT-TF), mitochondrially encoded 12S RNA (MT-RNR1), and long interspersed nucleotide element-1 (LINE-1) genes. The blood Cr ion concentration and micronucleus frequency of the Cr-exposed group were higher than those of the control group, whereas the mtDNA copy number remained unchanged. The methylation levels of MT-TF and MT-RNR1 but not LINE-1 were significantly lower in Cr-exposed workers. Pearson correlation analysis showed that workers with higher blood Cr ion concentrations exhibited lower MT-TF and MT-RNR1 gene methylation, and multiple linear regression analysis indicated that CpG sites 1 and 2 in MT-TF and CpG site 6 in MT-RNR1 were affected. These results suggested that methylation level of mtDNA has the possibility of acting as an alternative effect biomarker for Cr exposure. PMID:26656300

  11. Forensic DNA analysis.

    PubMed

    McDonald, Jessica; Lehman, Donald C

    2012-01-01

    Before the routine use of DNA profiling, blood typing was an important forensic tool. However, blood typing was not very discriminating. For example, roughly 30% of the United States population has type A-positive blood. Therefore, if A-positive blood were found at a crime scene, it could have come from 30% of the population. DNA profiling has a much better ability for discrimination. Forensic laboratories no longer routinely determine blood type. If blood is found at a crime scene, DNA profiling is performed. From Jeffrey's discovery of DNA fingerprinting to the development of PCR of STRs to the formation of DNA databases, our knowledge of DNA and DNA profiling have expanded greatly. Also, the applications for which we use DNA profiling have increased. DNA profiling is not just used for criminal case work, but it has expanded to encompass paternity testing, disaster victim identification, monitoring bone marrow transplants, detecting fetal cells in a mother's blood, tracing human history, and a multitude of other areas. The future of DNA profiling looks expansive with the development of newer instrumentation and techniques. PMID:22693781

  12. Circulating Cell Free DNA in the Diagnosis of Trophoblastic Tumors

    PubMed Central

    Openshaw, Mark R.; Harvey, Richard A.; Sebire, Neil J.; Kaur, Baljeet; Sarwar, Naveed; Seckl, Michael J.; Fisher, Rosemary A.

    2015-01-01

    Gestational trophoblastic neoplasia (GTN) represents a group of diseases characterized by production of human chorionic gonadotropin (hCG). Since non-gestational tumors may occasionally secrete hCG, histopathological diagnosis is important for appropriate clinical management. However, a histopathological diagnosis is not always available. We therefore investigated the feasibility of extracting cell free DNA (cfDNA) from the plasma of women with GTN for use as a “liquid biopsy” in patients without histopathological diagnosis. cfDNA was prepared from the plasma of 20 women with a diagnosis of GTN and five with hCG-secreting tumors of unknown origin. Genotyping of cfDNA from the patient, genomic DNA from her and her partner and DNA from the tumor tissue identified circulating tumor DNA (ctDNA) (from 9% to 53% of total cfDNA) in 12 of 20 patients with GTN. In one case without a tissue diagnosis, ctDNA enabled a diagnosis of GTN originating in a non-molar conception and in another a diagnosis of non-gestational tumor, based on the high degree of allelic instability and loss of heterozygosity in the ctDNA. In summary ctDNA can be detected in the plasma of women with GTN and can facilitate the diagnosis of both gestational and non-gestational trophoblastic tumors in cases without histopathological diagnosis. PMID:26981554

  13. DNA–DNA hybridization study of strains of Chryseobacterium, Elizabethkingia and Empedobacter and of other usually indole-producing non-fermenters of CDC groups IIc, IIe, IIh and IIi, mostly from human clinical sources, and proposals of Chryseobacterium bernardetii sp. nov., Chryseobacterium carnis sp. nov., Chryseobacterium lactis sp. nov., Chryseobacterium nakagawai sp. nov. and Chryseobacterium taklimakanense comb. nov

    PubMed Central

    Holmes, B.; Steigerwalt, A. G.; Nicholson, A. C.

    2015-01-01

    The taxonomic classification of 182 phenotypically similar isolates was evaluated using DNA–DNA hybridization and 16S rRNA gene sequence analysis. These bacterial isolates were mainly derived from clinical sources; all were Gram-negative non-fermenters and most were indoleproducing. Phenotypically, they resembled species from the genera Chryseobacterium, Elizabethkingia or Empedobacter or belonged to CDC groups IIc, IIe, IIh and IIi. Based on these analyses, four novel species are described: Chryseobacterium bernardetii sp. nov. (type strain NCTC 13530T=CCUG 60564T=CDC G229T), Chryseobacterium carnis sp. nov. (type strain NCTC 13525T=CCUG 60559T=CDC G81T), Chryseobacterium lactis sp. nov. (type strain NCTC 11390T=CCUG 60566T=CDC KC1864T) and Chryseobacterium nakagawai sp. nov. (type strain NCTC 13529T=CCUG 60563T=CDC G41T). The new combination Chryseobacterium taklimakanense comb. nov. (type strain NCTC 13490T=X-65T=CCTCC AB 208154T=NRRL B-51322T) is also proposed to accommodate the reclassified Planobacterium taklimakanense. PMID:23934253

  14. Interaction between DNA and cationic surfactants: effect of DNA conformation and surfactant headgroup.

    PubMed

    Dias, Rita S; Magno, Luís M; Valente, Artur J M; Das, Dibyendu; Das, Prasanta K; Maiti, Souvik; Miguel, Maria G; Lindman, Björn

    2008-11-20

    The interactions between DNA and a number of different cationic surfactants, differing in headgroup polarity, were investigated by electric conductivity measurements and fluorescence microscopy. It was observed that, the critical association concentration (cac), characterizing the onset of surfactant binding to DNA, does not vary significantly with the architecture of the headgroup. However, comparing with the critical micelle concentration (cmc) in the absence of DNA, it can be inferred that the micelles of a surfactant with a simple quaternary ammonium headgroup are much more stabilized by the presence of DNA than those of surfactants with hydroxylated head-groups. In line with previous studies of polymer-surfactant association, the cac does not vary significantly with either the DNA concentration or its chain length. On the other hand, a novel observation is that the cac is much lower when DNA is denaturated and in the single-stranded conformation, than for the double-helix DNA. This is contrary to expectation for a simple electrostatically driven association. Thus previous studies of polyelectrolyte-surfactant systems have shown that the cac decreases strongly with increasing linear charge density of the polyion. Since double-stranded DNA (dsDNA) has twice as large linear charge density as single-stranded DNA (ssDNA), the stronger binding in the latter case indicates an important role of nonelectrostatic effects. Both a higher flexibility of ssDNA and a higher hydrophobicity due to the exposed bases are found to play a role, with the hydrophobic interaction argued to be more important. The significance of hydrophobic DNA-surfactant interaction is in line with other observations. The significance of nonelectrostatic effects is also indicated in significant differences in cac between different surfactants for ssDNA but not for dsDNA. For lower concentrations of DNA, the conductivity measurements presented an "anomalous" feature, i.e., a second inflection point

  15. All that is gold does not glitter? Age, taxonomy, and ancient plant DNA quality

    PubMed Central

    Choi, JinHee; Lee, HyeJi

    2015-01-01

    More than 600 herbarium samples from four distantly related groups of flowering plants were used for DNA extraction and subsequent measurements of DNA purity and concentration. We did not find any significant relation between DNA purity and the age of the sample. However, DNA yields were different between plant groups studied. We believe that there there should be no reservations about “old” samples if the goal is to extract more DNA of better purity. We argue that the older herbarium samples are the mine for the future DNA studies, and have the value not less than the “fresh” specimens. PMID:26244108

  16. All that is gold does not glitter? Age, taxonomy, and ancient plant DNA quality.

    PubMed

    Choi, JinHee; Lee, HyeJi; Shipunov, Alexey

    2015-01-01

    More than 600 herbarium samples from four distantly related groups of flowering plants were used for DNA extraction and subsequent measurements of DNA purity and concentration. We did not find any significant relation between DNA purity and the age of the sample. However, DNA yields were different between plant groups studied. We believe that there there should be no reservations about "old" samples if the goal is to extract more DNA of better purity. We argue that the older herbarium samples are the mine for the future DNA studies, and have the value not less than the "fresh" specimens. PMID:26244108

  17. Simulation of Dna-Nanotube Interactions

    NASA Astrophysics Data System (ADS)

    Gao, Huajian; Kong, Yong

    2004-08-01

    Carbon nanotubes functionalized with biological molecules (such as protein peptides and nucleic acids) show great potential for application in bioengineering and nanotechnology. Fundamental understanding, description, and regulation of such bio-nano-systems will ultimately lead to a new generation of integrated systems that combine unique properties of the carbon nanotube (CNT) with biological recognition capabilities. In this review, we describe recent advances in understanding the interactions between deoxyribonucleic acids (DNA) and CNT, as well as relevant simulation techniques. We also review progress in simulating DNA noncovalent interactions with CNTs in an aqueous environment. Molecular dynamics simulations indicate that DNA molecules may be encapsulated inside or wrap around CNT owing to van der Waals attraction between DNA and CNT. We focus on the dynamics and energetics of DNA encapsulation inside nanotubes and discuss the mechanism of encapsulation and the effects of nanotube size, nanotube end-group, DNA base sequence, solvent temperature and pressure on the encapsulation process. Finally, we discuss the likely impact of DNA encapsulation on bioengineering and nanotechnology, as well as other potential applications.

  18. Group Work Publication-1991.

    ERIC Educational Resources Information Center

    Zimpfer, David G.

    1992-01-01

    Lists 21 new publications in group work, of which 9 are reviewed. Those discussed include publications on group counseling and psychotherapy, structured groups, support groups, psychodrama, and social group work. (Author/NB)

  19. Method for producing intact plants containing foreign DNA

    SciTech Connect

    Simpson, R.B.; Margossian, L.J.

    1987-04-14

    An in vivo method is described for transforming and regenerating whole dicotyledenous plants comprising: infecting a dicotyledenous plant (P) with Rhizobiaceae bacteria containing virulence functions; a first plasmid containing T-DNA terminal sequences flanking oncogenic factors capable of inducing a shoot-bearing shooty tumor on plant (P), and a second plasmid containing T-DNA terminal sequences flanking heterologous transfer DNA capable of being integrated into nuclear DNA of plant (P) cells wherein the second plasmid does not contain any oncogenic factors; maintaining the infected dicotyledenous plant (P) until shoot-bearing shooty tumor develops on the dicotyledenous plant (P); selecting those shoots, or progeny wherein the progeny are selected from the group consisting of seeds or tubers, that contain transformed cells having heterologous transfer DNA but not any tumorous DNA integrated into their genomes; and utilizing the selected shoots, or progeny to produce whole plants that contain cells having heterologous transfer DNA integrated into their genomes.

  20. Getting Ready for the Dance: FANCJ Irons Out DNA Wrinkles

    PubMed Central

    Bharti, Sanjay Kumar; Awate, Sanket; Banerjee, Taraswi; Brosh, Robert M.

    2016-01-01

    Mounting evidence indicates that alternate DNA structures, which deviate from normal double helical DNA, form in vivo and influence cellular processes such as replication and transcription. However, our understanding of how the cellular machinery deals with unusual DNA structures such as G-quadruplexes (G4), triplexes, or hairpins is only beginning to emerge. New advances in the field implicate a direct role of the Fanconi Anemia Group J (FANCJ) helicase, which is linked to a hereditary chromosomal instability disorder and important for cancer suppression, in replication past unusual DNA obstacles. This work sets the stage for significant progress in dissecting the molecular mechanisms whereby replication perturbation by abnormal DNA structures leads to genomic instability. In this review, we focus on FANCJ and its role to enable efficient DNA replication when the fork encounters vastly abundant naturally occurring DNA obstacles, which may have implications for targeting rapidly dividing cancer cells. PMID:27376332

  1. Getting Ready for the Dance: FANCJ Irons Out DNA Wrinkles.

    PubMed

    Bharti, Sanjay Kumar; Awate, Sanket; Banerjee, Taraswi; Brosh, Robert M

    2016-01-01

    Mounting evidence indicates that alternate DNA structures, which deviate from normal double helical DNA, form in vivo and influence cellular processes such as replication and transcription. However, our understanding of how the cellular machinery deals with unusual DNA structures such as G-quadruplexes (G4), triplexes, or hairpins is only beginning to emerge. New advances in the field implicate a direct role of the Fanconi Anemia Group J (FANCJ) helicase, which is linked to a hereditary chromosomal instability disorder and important for cancer suppression, in replication past unusual DNA obstacles. This work sets the stage for significant progress in dissecting the molecular mechanisms whereby replication perturbation by abnormal DNA structures leads to genomic instability. In this review, we focus on FANCJ and its role to enable efficient DNA replication when the fork encounters vastly abundant naturally occurring DNA obstacles, which may have implications for targeting rapidly dividing cancer cells. PMID:27376332

  2. Sequence specificity of DNA cleavage by Micrococcus luteus. gamma. endonuclease

    SciTech Connect

    Hentosh, P.; Henner, W.D.; Reynolds, R.J.

    1985-04-01

    DNA fragments of defined sequence have been used to determine the sites of cleavage by ..gamma..-endonuclease activity in extracts prepared from Micrococcus luteus. End-labeled DNA restriction fragments of pBR322 DNA that had been irradiated under nitrogen in the presence of potassium iodide or t-butanol were treated with M. luteus ..gamma.. endonuclease and analyzed on irradiated DNA preferentially at the positions of cytosines and thymines. DNA cleavage occurred immediately to the 3' side of pyrimidines in irradiated DNA and resulted in fragments that terminate in a 5'-phosphoryl group. These studies indicate that both altered cytosines and thymines may be important DNA lesions requiring repair after exposure to ..gamma.. radiation.

  3. Viroid-induced DNA methylation in plants.

    PubMed

    Dalakouras, Athanasios; Dadami, Elena; Wassenegger, Michael

    2013-12-01

    In eukaryotes, DNA methylation refers to the addition of a methyl group to the fifth atom in the six-atom ring of cytosine residues. At least in plants, DNA regions that become de novo methylated can be defined by homologous RNA molecules in a process termed RNA-directed DNA methylation (RdDM). RdDM was first discovered in viroid-infected plants. Viroids are pathogenic circular, non-coding, single-stranded RNA molecules. Members of the Pospiviroidae family replicate in the nucleus through double-stranded RNA intermediates, attracting the host RNA silencing machinery. The recruitment of this machinery results in the production of viroid-derived small RNAs (vd-sRNAs) that mediate RNA degradation and DNA methylation of cognate sequences. Here, we provide an overview of the cumulative data on the field of viroid-induced RdDM and discuss three possible scenarios concerning the mechanistic details of its establishment. PMID:25436756

  4. Group Composition, Group Interaction and Achievement in Cooperative Small Groups.

    ERIC Educational Resources Information Center

    Webb, Noreen M.

    This study investigated interaction and achievement in cooperative small groups in four junior high school mathematics classrooms. Ninety-six students learned a one-week unit on consumer mathematics in mixed-ability or uniform-ability groups. Students in mixed-ability groups scored higher on a problem-solving test than students in uniform-ability…

  5. Group Cohesion in Experiential Growth Groups

    ERIC Educational Resources Information Center

    Steen, Sam; Vasserman-Stokes, Elaina; Vannatta, Rachel

    2014-01-01

    This article explores the effect of web-based journaling on changes in group cohesion within experiential growth groups. Master's students were divided into 2 groups. Both used a web-based platform to journal after each session; however, only 1 of the groups was able to read each other's journals. Quantitative data collected before and…

  6. Evaluation of DNAstable for DNA storage at ambient temperature.

    PubMed

    Howlett, Susanne E; Castillo, Hilda S; Gioeni, Lora J; Robertson, James M; Donfack, Joseph

    2014-01-01

    Preserving DNA is important for validation of prospective and retrospective analyses, requiring many expensive types of equipment (e.g., freezers and back-up generators) and energy. While freezing is the most common method for storing extracted DNA evidence or well-characterized DNA samples for validation studies, DNAstable (Biomatrica), a commercially available medium for room temperature storage of DNA extracts was evaluated in this study. Two groups of samples consisting of different DNA quantities were investigated, one ranging from 20 to 400 ng (group 1) and the other one ranging from 1.4 to 20 ng (group 2). The DNA samples with and without DNAstable were stored at four different temperatures [∼25 °C (room temperature), -20 °C, 37 °C or 50 °C]. DNA degradation over several months was monitored by SYBR Green-based qPCR assays and by PCR amplification of the core CODIS STR markers for group 1 and 2 DNA samples, respectively. For the time points tested in this study (up to 365 days), the findings indicate that the -20 °C controls and the DNAstable protected samples at room temperature provided similar DNA recoveries that were higher compared to the unprotected controls kept at RT, 37 °C or 50 °C. These results suggest that DNAstable can protect DNA samples with effectiveness similar to that of the traditional -20 °C freezing method. In addition, extrapolations from accelerated aging experiments conducted at high temperatures support that DNAstable is an effective technology for preserving purified DNA at room temperature with a larger protective impact on DNA samples of low quantity (<20 ng). PMID:24315605

  7. An overview of the structures of protein-DNA complexes

    PubMed Central

    Luscombe, Nicholas M; Austin, Susan E; Berman , Helen M; Thornton, Janet M

    2000-01-01

    On the basis of a structural analysis of 240 protein-DNA complexes contained in the Protein Data Bank (PDB), we have classified the DNA-binding proteins involved into eight different structural/functional groups, which are further classified into 54 structural families. Here we present this classification and review the functions, structures and binding interactions of these protein-DNA complexes. PMID:11104519

  8. Detection and quantitation of single nucleotide polymorphisms, DNA sequence variations, DNA mutations, DNA damage and DNA mismatches

    DOEpatents

    McCutchen-Maloney, Sandra L.

    2002-01-01

    DNA mutation binding proteins alone and as chimeric proteins with nucleases are used with solid supports to detect DNA sequence variations, DNA mutations and single nucleotide polymorphisms. The solid supports may be flow cytometry beads, DNA chips, glass slides or DNA dips sticks. DNA molecules are coupled to solid supports to form DNA-support complexes. Labeled DNA is used with unlabeled DNA mutation binding proteins such at TthMutS to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by binding which gives an increase in signal. Unlabeled DNA is utilized with labeled chimeras to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by nuclease activity of the chimera which gives a decrease in signal.

  9. Class Numbers and Groups of Algebraic Groups

    NASA Astrophysics Data System (ADS)

    Platonov, V. P.; Bondarenko, A. A.; Rapinčuk, A. S.

    1980-06-01

    The class number of an algebraic group G defined over a global field is the number of double cosets of the adele group GA with respect to the subgroups of integral and principal adeles. In most cases the set of double cosets has the natural structure of an abelian group, called the class group of G. In this article the class number of a semisimple group G is computed, and it is proved that any finite abelian group can be realized as a class group.Bibliography: 24 titles.

  10. DNA profiles from fingermarks.

    PubMed

    Templeton, Jennifer E L; Linacre, Adrian

    2014-11-01

    Criminal investigations would be considerably improved if DNA profiles could be routinely generated from single fingermarks. Here we report a direct DNA profiling method that was able to generate interpretable profiles from 71% of 170 fingermarks. The data are based on fingermarks from all 5 digits of 34 individuals. DNA was obtained from the fingermarks using a swab moistened with Triton-X, and the fibers were added directly to one of two commercial DNA profiling kits. All profiles were obtained without increasing the number of amplification cycles; therefore, our method is ideally suited for adoption by the forensic science community. We indicate the use of the technique in a criminal case in which a DNA profile was generated from a fingermark on tape that was wrapped around a drug seizure. Our direct DNA profiling approach is rapid and able to generate profiles from touched items when current forensic practices have little chance of success. PMID:25391915

  11. Electrocatalysis in DNA Sensors.

    PubMed

    Furst, Ariel; Hill, Michael G; Barton, Jacqueline K

    2014-12-14

    Electrocatalysis is often thought of solely in the inorganic realm, most often applied to energy conversion in fuel cells. However, the ever-growing field of bioelectrocatalysis has made great strides in advancing technology for both biofuel cells as well as biological detection platforms. Within the context of bioelectrocatalytic detection systems, DNA-based platforms are especially prevalent. One subset of these platforms, the one we have developed, takes advantage of the inherent charge transport properties of DNA. Electrocatalysis coupled with DNA-mediated charge transport has enabled specific and sensitive detection of lesions, mismatches and DNA-binding proteins. Even greater signal amplification from these platforms is now being achieved through the incorporation of a secondary electrode to the platform both for patterning DNA arrays and for detection. Here, we describe the evolution of this new DNA sensor technology. PMID:25435647

  12. DNA Media Storage

    PubMed Central

    Bogard, Christy M.; Rouchka, Eric C.

    2010-01-01

    In 1994, University of Southern California computer scientist Dr. Leonard Adelman solved the Hamiltonian Path Problem using DNA as a computational mechanism. He proved the principle that DNA computing could be used to solve computationally complex problems. Because of the limitations in discovery time, resource requirements, and sequence mismatches, DNA computing has not yet become a commonly accepted practice. However, advancements are continually being discovered that are evolving the field of DNA Computing. Practical applications of DNA are not restricted to computation alone. This research presents a novel approach in which DNA could be used as a means of storing files. Through the use of Multiple Sequence Alignment combined with intelligent heuristics, the most probabilistic file contents can be determined with minimal errors. PMID:20622994

  13. DNA Media Storage.

    PubMed

    Bogard, Christy M; Rouchka, Eric C

    2007-09-01

    In 1994, University of Southern California computer scientist Dr. Leonard Adelman solved the Hamiltonian Path Problem using DNA as a computational mechanism. He proved the principle that DNA computing could be used to solve computationally complex problems. Because of the limitations in discovery time, resource requirements, and sequence mismatches, DNA computing has not yet become a commonly accepted practice. However, advancements are continually being discovered that are evolving the field of DNA Computing. Practical applications of DNA are not restricted to computation alone. This research presents a novel approach in which DNA could be used as a means of storing files. Through the use of Multiple Sequence Alignment combined with intelligent heuristics, the most probabilistic file contents can be determined with minimal errors. PMID:20622994

  14. Ribonucleotides in Bacterial DNA

    PubMed Central

    Schroeder, Jeremy W.; Randall, Justin R.; Matthews, Lindsay A.; Simmons, Lyle A.

    2014-01-01

    In all living cells, DNA is the storage medium for genetic information. Being quite stable, DNA is well-suited for its role in storage and propagation of information, but RNA is also covalently included in DNA through various mechanisms. Recent studies also demonstrate useful aspects of including ribonucleotides in the genome during repair. Therefore, our understanding of the consequences of RNA inclusion into bacterial genomic DNA is just beginning, but with its high frequency of occurrence the consequences and potential benefits are likely to be numerous and diverse. In this review, we discuss the processes that cause ribonucleotide inclusion in genomic DNA, the pathways important for ribonucleotide removal and the consequences that arise should ribonucleotides remain nested in genomic DNA. PMID:25387798

  15. Electrocatalysis in DNA Sensors

    PubMed Central

    Furst, Ariel; Hill, Michael G.; Barton, Jacqueline K.

    2014-01-01

    Electrocatalysis is often thought of solely in the inorganic realm, most often applied to energy conversion in fuel cells. However, the ever-growing field of bioelectrocatalysis has made great strides in advancing technology for both biofuel cells as well as biological detection platforms. Within the context of bioelectrocatalytic detection systems, DNA-based platforms are especially prevalent. One subset of these platforms, the one we have developed, takes advantage of the inherent charge transport properties of DNA. Electrocatalysis coupled with DNA-mediated charge transport has enabled specific and sensitive detection of lesions, mismatches and DNA-binding proteins. Even greater signal amplification from these platforms is now being achieved through the incorporation of a secondary electrode to the platform both for patterning DNA arrays and for detection. Here, we describe the evolution of this new DNA sensor technology. PMID:25435647

  16. DNA-based machines.

    PubMed

    Wang, Fuan; Willner, Bilha; Willner, Itamar

    2014-01-01

    The base sequence in nucleic acids encodes substantial structural and functional information into the biopolymer. This encoded information provides the basis for the tailoring and assembly of DNA machines. A DNA machine is defined as a molecular device that exhibits the following fundamental features. (1) It performs a fuel-driven mechanical process that mimics macroscopic machines. (2) The mechanical process requires an energy input, "fuel." (3) The mechanical operation is accompanied by an energy consumption process that leads to "waste products." (4) The cyclic operation of the DNA devices, involves the use of "fuel" and "anti-fuel" ingredients. A variety of DNA-based machines are described, including the construction of "tweezers," "walkers," "robots," "cranes," "transporters," "springs," "gears," and interlocked cyclic DNA structures acting as reconfigurable catenanes, rotaxanes, and rotors. Different "fuels", such as nucleic acid strands, pH (H⁺/OH⁻), metal ions, and light, are used to trigger the mechanical functions of the DNA devices. The operation of the devices in solution and on surfaces is described, and a variety of optical, electrical, and photoelectrochemical methods to follow the operations of the DNA machines are presented. We further address the possible applications of DNA machines and the future perspectives of molecular DNA devices. These include the application of DNA machines as functional structures for the construction of logic gates and computing, for the programmed organization of metallic nanoparticle structures and the control of plasmonic properties, and for controlling chemical transformations by DNA machines. We further discuss the future applications of DNA machines for intracellular sensing, controlling intracellular metabolic pathways, and the use of the functional nanostructures for drug delivery and medical applications. PMID:24647836

  17. Markovian language model of the DNA and its information content

    PubMed Central

    Srivastava, S.; Baptista, M. S.

    2016-01-01

    This work proposes a Markovian memoryless model for the DNA that simplifies enormously the complexity of it. We encode nucleotide sequences into symbolic sequences, called words, from which we establish meaningful length of words and groups of words that share symbolic similarities. Interpreting a node to represent a group of similar words and edges to represent their functional connectivity allows us to construct a network of the grammatical rules governing the appearance of groups of words in the DNA. Our model allows us to predict the transition between groups of words in the DNA with unprecedented accuracy, and to easily calculate many informational quantities to better characterize the DNA. In addition, we reduce the DNA of known bacteria to a network of only tens of nodes, show how our model can be used to detect similar (or dissimilar) genes in different organisms, and which sequences of symbols are responsible for most of the information content of the DNA. Therefore, the DNA can indeed be treated as a language, a Markovian language, where a ‘word’ is an element of a group, and its grammar represents the rules behind the probability of transitions between any two groups. PMID:26909179

  18. DNA ELECTROPHORESIS AT SURFACES

    SciTech Connect

    RAFAILOVICH, MIRIAM; SOKOLOV, JONATHAN; GERSAPPE, DILIP

    2003-09-01

    During this year we performed two major projects: I. We developed a detailed theoretical model which complements our experiments on surface DNA electrophoresis. We found that it was possible to enhance the separation of DNA chains by imposing a chemical nanoscale pattern on the surface. This approach utilized the surface interaction effect of the DNA chains with the substrate and is a refinement to our previous method in which DNA chains were separated on homogeneous flat surfaces. By introducing the nano-patterns on the surface, the conformational changes of DNA chains of different lengths can be amplified, which results in the different friction strengths with the substrate surface. Our results also show that, when compared to the DNA electrophoresis performed on homogeneous flat surfaces, nanopatterned surfaces offer a larger window in choosing different surface interactions to achieve separation. II. In collaboration with a large international manufacturer of skin care products we also embarked on a project involving photo toxicity of titanium dioxide nanoparticles, which are a key ingredient in sunscreen and cosmetic lotions. The results clearly implicated the nanoparticles in catalyzing damage to chromosomal DNA. We then used this knowledge to develop a polymer/anti-oxidant coating which prevented the photocatalytic reaction on DNA while still retaining the UV absorptive properties of the nanoparticles. The standard gel electrophoresis was not sufficient in determining the extent of the DNA damage. The conclusions of this study were based predominantly on analysis obtained with the surface electrophoresis method.

  19. Tiny telomere DNA

    PubMed Central

    Ren, Jinsong; Qu, Xiaogang; Trent, John O.; Chaires, Jonathan B.

    2002-01-01

    We describe the design, synthesis and biophysical characterization of a novel DNA construct in which a folded quadruplex structure is joined to a standard double helix. Circular dichroism, gel electrophoresis, three-dimensional UV melting and differential scanning calorimetry were all used to characterize the structure. Rigorous molecular dynamics simulations were used to build a plausible atomic-level structural model of the DNA construct. This novel DNA construct provides a model for the duplex–quadruplex junction region at the end of chromosomal DNA and offers a system for the study of structure-selective ligand binding. PMID:12034817

  20. DNA Damage Response

    PubMed Central

    Giglia-Mari, Giuseppina; Zotter, Angelika; Vermeulen, Wim

    2011-01-01

    Structural changes to DNA severely affect its functions, such as replication and transcription, and play a major role in age-related diseases and cancer. A complicated and entangled network of DNA damage response (DDR) mechanisms, including multiple DNA repair pathways, damage tolerance processes, and cell-cycle checkpoints safeguard genomic integrity. Like transcription and replication, DDR is a chromatin-associated process that is generally tightly controlled in time and space. As DNA damage can occur at any time on any genomic location, a specialized spatio-temporal orchestration of this defense apparatus is required. PMID:20980439

  1. Whole DNA methylome profiling in mice exposed to secondhand smoke

    PubMed Central

    Tommasi, Stella; Zheng, Albert; Yoon, Jae-In; Li, Arthur Xuejun; Wu, Xiwei; Besaratinia, Ahmad

    2012-01-01

    Aberration of DNA methylation is a prime epigenetic mechanism of carcinogenesis. Aberrant DNA methylation occurs frequently in lung cancer, with exposure to secondhand smoke (SHS) being an established risk factor. The causal role of SHS in the genesis of lung cancer, however, remains elusive. To investigate whether SHS can cause aberrant DNA methylation in vivo, we have constructed the whole DNA methylome in mice exposed to SHS for a duration of 4 mo, both after the termination of exposure and at ensuing intervals post-exposure (up to 10 mo). Our genome-wide and gene-specific profiling of DNA methylation in the lung of SHS-exposed mice revealed that all groups of SHS-exposed mice and controls share a similar pattern of DNA methylation. Furthermore, the methylation status of major repetitive DNA elements, including long-interspersed nuclear elements (LINE L1), intracisternal A particle long-terminal repeat retrotransposons (IAP-LTR), and short-interspersed nuclear elements (SINE B1), in the lung of all groups of SHS-exposed mice and controls remains comparable. The absence of locus-specific gain of DNA methylation and global loss of DNA methylation in the lung of SHS-exposed mice within a timeframe that precedes neoplastic-lesion formation underscore the challenges of lung cancer biomarker development. Identifying the initiating events that cause aberrant DNA methylation in lung carcinogenesis may help improve future strategies for prevention, early detection and treatment of this highly lethal disease. PMID:23051858

  2. Recombinant DNA technology in apple.

    PubMed

    Gessler, Cesare; Patocchi, Andrea

    2007-01-01

    This review summarizes the achievements of almost 20 years of recombinant DNA technology applied to apple, grouping the research results into the sections: developing the technology, insect resistance, fungal disease resistance, self-incompatibility, herbicide resistance, fire blight resistance, fruit ripening, allergens, rooting ability, and acceptance and risk assessment. The diseases fire blight, caused by Erwinia amylovora, and scab, caused by Venturia inaequalis, were and still are the prime targets. Shelf life improvement and rooting ability of rootstocks are also relevant research areas. The tools to create genetically modified apples of added value to producers, consumers, and the environment are now available. PMID:17522823

  3. Low-Dose Formaldehyde Delays DNA Damage Recognition and DNA Excision Repair in Human Cells

    PubMed Central

    Luch, Andreas; Frey, Flurina C. Clement; Meier, Regula; Fei, Jia; Naegeli, Hanspeter

    2014-01-01

    Objective Formaldehyde is still widely employed as a universal crosslinking agent, preservative and disinfectant, despite its proven carcinogenicity in occupationally exposed workers. Therefore, it is of paramount importance to understand the possible impact of low-dose formaldehyde exposures in the general population. Due to the concomitant occurrence of multiple indoor and outdoor toxicants, we tested how formaldehyde, at micromolar concentrations, interferes with general DNA damage recognition and excision processes that remove some of the most frequently inflicted DNA lesions. Methodology/Principal Findings The overall mobility of the DNA damage sensors UV-DDB (ultraviolet-damaged DNA-binding) and XPC (xeroderma pigmentosum group C) was analyzed by assessing real-time protein dynamics in the nucleus of cultured human cells exposed to non-cytotoxic (<100 μM) formaldehyde concentrations. The DNA lesion-specific recruitment of these damage sensors was tested by monitoring their accumulation at local irradiation spots. DNA repair activity was determined in host-cell reactivation assays and, more directly, by measuring the excision of DNA lesions from chromosomes. Taken together, these assays demonstrated that formaldehyde obstructs the rapid nuclear trafficking of DNA damage sensors and, consequently, slows down their relocation to DNA damage sites thus delaying the excision repair of target lesions. A concentration-dependent effect relationship established a threshold concentration of as low as 25 micromolar for the inhibition of DNA excision repair. Conclusions/Significance A main implication of the retarded repair activity is that low-dose formaldehyde may exert an adjuvant role in carcinogenesis by impeding the excision of multiple mutagenic base lesions. In view of this generally disruptive effect on DNA repair, we propose that formaldehyde exposures in the general population should be further decreased to help reducing cancer risks. PMID:24722772

  4. Group theories: relevance to group safety studies.

    PubMed

    Benevento, A L

    1998-01-01

    Promoting safety in the workplace has been attempted in a variety of ways. Increasingly, industries are using groups such as safety teams and quality circles to promote worker safety. Group influences on individual behavior and attitudes have long been studied in the social psychology literature, but the theories have not been commonly found outside the psychology arena. This paper describes the group theories of group polarization, risky shift, social loafing, groupthink and team think and attempts to apply these theories to existing studies that examine work group influences on safety. Interesting parallels were found but only one study examined group influences as their primary focus of research. Since groups are increasingly used for safety promotion, future research on safety that studies group influences with respect to current group theories is recommended. PMID:24441299

  5. Sequencing mitochondrial DNA polymorphisms by hybridization

    SciTech Connect

    Chee, M.S.; Lockhart, D.J.; Hubbell, E.

    1994-09-01

    We have investigated the use of DNA chips for genetic analysis, using human mitochondrial DNA (mtDNA) as a model. The DNA chips are made up of ordered arrays of DNA oligonucleotide probes, synthesized on a glass substrate using photolithographic techniques. The synthesis site for each different probe is specifically addressed by illumination of the substrate through a photolithographic mask, achieving selective deprotection Nucleoside phosphoramidites bearing photolabile protecting groups are coupled only to exposed sites. Repeated cycles of deprotection and coupling generate all the probes in parallel. The set of 4{sup N} N-mer probes can be synthesized in only 4N steps. Any subset can be synthesized in 4N steps. Any subset can be synthesized in 4N or fewer steps. Sequences amplified from the D-loop region of human mitochondrial DNA (mtDNA) were fluorescently labelled and hybridized to DNA chips containing probes specific for mtDNA. Each nucleotide of a 1.3 kb region spanning the D loop is represented by four probes on the chip. Each probe has a different base at the position of interest: together they comprise a set of A, C, G and T probes which are otherwise identical. In principle, only one probe-target hybrid will be a perfect match. The other three will be single base mismatches. Fluorescence imaging of the hybridized chip allows quantification of hybridization signals. Heterozygous mixtures of sequences can also be characterized. We have developed software to quantitate and interpret the hybridization signals, and to call the sequence automatically. Results of sequence analysis of human mtDNAs will be presented.

  6. DNA Targeting Sequence Improves Magnetic Nanoparticle-Based Plasmid DNA Transfection Efficiency in Model Neurons

    PubMed Central

    Vernon, Matthew M.; Dean, David A.; Dobson, Jon

    2015-01-01

    Efficient non-viral plasmid DNA transfection of most stem cells, progenitor cells and primary cell lines currently presents an obstacle for many applications within gene therapy research. From a standpoint of efficiency and cell viability, magnetic nanoparticle-based DNA transfection is a promising gene vectoring technique because it has demonstrated rapid and improved transfection outcomes when compared to alternative non-viral methods. Recently, our research group introduced oscillating magnet arrays that resulted in further improvements to this novel plasmid DNA (pDNA) vectoring technology. Continued improvements to nanomagnetic transfection techniques have focused primarily on magnetic nanoparticle (MNP) functionalization and transfection parameter optimization: cell confluence, growth media, serum starvation, magnet oscillation parameters, etc. Noting that none of these parameters can assist in the nuclear translocation of delivered pDNA following MNP-pDNA complex dissociation in the cell’s cytoplasm, inclusion of a cassette feature for pDNA nuclear translocation is theoretically justified. In this study incorporation of a DNA targeting sequence (DTS) feature in the transfecting plasmid improved transfection efficiency in model neurons, presumably from increased nuclear translocation. This observation became most apparent when comparing the response of the dividing SH-SY5Y precursor cell to the non-dividing and differentiated SH-SY5Y neuroblastoma cells. PMID:26287182

  7. Quantitative analysis of cell-free DNA in ovarian cancer

    PubMed Central

    SHAO, XUEFENG; He, YAN; JI, MIN; CHEN, XIAOFANG; QI, JING; SHI, WEI; HAO, TIANBO; JU, SHAOQING

    2015-01-01

    The aim of the present study was to investigate the association between cell-free DNA (cf-DNA) levels and clinicopathological characteristics of patients with ovarian cancer using a branched DNA (bDNA) technique, and to determine the value of quantitative cf-DNA detection in assisting with the diagnosis of ovarian cancer. Serum specimens were collected from 36 patients with ovarian cancer on days 1, 3 and 7 following surgery, and additional serum samples were also collected from 22 benign ovarian tumor cases, and 19 healthy, non-cancerous ovaries. bDNA techniques were used to detect serum cf-DNA concentrations. All data were analyzed using SPSS version 18.0. The cf-DNA levels were significantly increased in the ovarian cancer group compared with those of the benign ovarian tumor group and healthy ovarian group (P<0.01). Furthermore, cf-DNA levels were significantly increased in stage III and IV ovarian cancer compared with those of stages I and II (P<0.01). In addition, cf-DNA levels were significantly increased on the first day post-surgery (P<0.01), and subsequently demonstrated a gradual decrease. In the ovarian cancer group, the area under the receiver operating characteristic curve of cf-DNA and the sensitivity were 0.917 and 88.9%, respectively, which was higher than those of cancer antigen 125 (0.724, 75%) and human epididymis protein 4 (0.743, 80.6%). There was a correlation between the levels of serum cf-DNA and the occurrence and development of ovarian cancer in the patients evaluated. bDNA techniques possessed higher sensitivity and specificity than other methods for the detection of serum cf-DNA in patients exhibiting ovarian cancer, and bDNA techniques are more useful for detecting cf-DNA than other factors. Thus, the present study demonstrated the potential value for the use of bDNA as an adjuvant diagnostic method for ovarian cancer. PMID:26788153

  8. Aberrant DNA methylation reprogramming in bovine SCNT preimplantation embryos

    PubMed Central

    Zhang, Sheng; Chen, Xin; Wang, Fang; An, Xinglan; Tang, Bo; Zhang, Xueming; Sun, Liguang; Li, Ziyi

    2016-01-01

    DNA methylation reprogramming plays important roles in mammalian embryogenesis. Mammalian somatic cell nuclear transfer (SCNT) embryos with reprogramming defects fail to develop. Thus, we compared DNA methylation reprogramming in preimplantation embryos from bovine SCNT and in vitro fertilization (IVF) and analyzed the influence of vitamin C (VC) on the reprogramming of DNA methylation. The results showed that global DNA methylation followed a typical pattern of demethylation and remethylation in IVF preimplantation embryos; however, the global genome remained hypermethylated in SCNT preimplantation embryos. Compared with the IVF group, locus DNA methylation reprogramming showed three patterns in the SCNT group. First, some pluripotency genes (POU5F1 and NANOG) and repeated elements (satellite I and α-satellite) showed insufficient demethylation and hypermethylation in the SCNT group. Second, a differentially methylated region (DMR) of an imprint control region (ICR) in H19 exhibited excessive demethylation and hypomethylation. Third, some pluripotency genes (CDX2 and SOX2) were hypomethylated in both the IVF and SCNT groups. Additionally, VC improved the DNA methylation reprogramming of satellite I, α-satellite and H19 but not that of POU5F1 and NANOG in SCNT preimplantation embryos. These results indicate that DNA methylation reprogramming was aberrant and that VC influenced DNA methylation reprogramming in SCNT embryos in a locus-specific manner. PMID:27456302

  9. ASSESSING THE EFFECTS OF HIGH METHIONINE INTAKE ON DNA METHYLATION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Methylation of DNA occurs at cytosines within CpG (cytosine-guanine) dinucleotides and is one of several epigenetic mechanisms that serve to establish and maintain tissue-specific patterns of gene expression. The methyl groups transferred in mammalian DNA methylation reactions are ultimately derived...

  10. Replicative DNA polymerases.

    PubMed

    Johansson, Erik; Dixon, Nicholas

    2013-06-01

    In 1959, Arthur Kornberg was awarded the Nobel Prize for his work on the principles by which DNA is duplicated by DNA polymerases. Since then, it has been confirmed in all branches of life that replicative DNA polymerases require a single-stranded template to build a complementary strand, but they cannot start a new DNA strand de novo. Thus, they also depend on a primase, which generally assembles a short RNA primer to provide a 3'-OH that can be extended by the replicative DNA polymerase. The general principles that (1) a helicase unwinds the double-stranded DNA, (2) single-stranded DNA-binding proteins stabilize the single-stranded DNA, (3) a primase builds a short RNA primer, and (4) a clamp loader loads a clamp to (5) facilitate the loading and processivity of the replicative polymerase, are well conserved among all species. Replication of the genome is remarkably robust and is performed with high fidelity even in extreme environments. Work over the last decade or so has confirmed (6) that a common two-metal ion-promoted mechanism exists for the nucleotidyltransferase reaction that builds DNA strands, and (7) that the replicative DNA polymerases always act as a key component of larger multiprotein assemblies, termed replisomes. Furthermore (8), the integrity of replisomes is maintained by multiple protein-protein and protein-DNA interactions, many of which are inherently weak. This enables large conformational changes to occur without dissociation of replisome components, and also means that in general replisomes cannot be isolated intact. PMID:23732474

  11. Many Ways to Loop DNA

    PubMed Central

    Griffith, Jack D.

    2013-01-01

    In the 1960s, I developed methods for directly visualizing DNA and DNA-protein complexes using an electron microscope. This made it possible to examine the shape of DNA and to visualize proteins as they fold and loop DNA. Early applications included the first visualization of true nucleosomes and linkers and the demonstration that repeating tracts of adenines can cause a curvature in DNA. The binding of DNA repair proteins, including p53 and BRCA2, has been visualized at three- and four-way junctions in DNA. The trombone model of DNA replication was directly verified, and the looping of DNA at telomeres was discovered. PMID:24005675

  12. DNA Barcodes for Nearctic Auchenorrhyncha (Insecta: Hemiptera)

    PubMed Central

    Foottit, Robert G.; Maw, Eric; Hebert, P. D. N.

    2014-01-01

    Background Many studies have shown the suitability of sequence variation in the 5′ region of the mitochondrial cytochrome c oxidase I (COI) gene as a DNA barcode for the identification of species in a wide range of animal groups. We examined 471 species in 147 genera of Hemiptera: Auchenorrhyncha drawn from specimens in the Canadian National Collection of Insects to assess the effectiveness of DNA barcoding in this group. Methodology/Principal Findings Analysis of the COI gene revealed less than 2% intra-specific divergence in 93% of the taxa examined, while minimum interspecific distances exceeded 2% in 70% of congeneric species pairs. Although most species are characterized by a distinct sequence cluster, sequences for members of many groups of closely related species either shared sequences or showed close similarity, with 25% of species separated from their nearest neighbor by less than 1%. Conclusions/Significance This study, although preliminary, provides DNA barcodes for about 8% of the species of this hemipteran suborder found in North America north of Mexico. Barcodes can enable the identification of many species of Auchenorrhyncha, but members of some species groups cannot be discriminated. Future use of DNA barcodes in regulatory, pest management, and environmental applications will be possible as the barcode library for Auchenorrhyncha expands to include more species and broader geographic coverage. PMID:25004106

  13. Escherichia coli DnaB Helicase–DnaC Protein Complex: Allosteric Effects of the Nucleotides on the Nucleic Acid Binding and the Kinetic Mechanism of NTP Hydrolysis. 3†

    PubMed Central

    Roychowdhury, Anasuya; Szymanski, Michal R.; Jezewska, Maria J.; Bujalowski, Wlodzimierz

    2011-01-01

    Allosteric interactions between the DNA- and NTP-binding sites of the Escherichia coli DnaB helicase engaged in the DnaB–DnaC complex and the mechanism of NTP hydrolysis by the complex have been examined using the fluorescence titration, analytical ultracentrifugation, and rapid quench-flow technique. Surprisingly, the ssDNA affinity of the DnaB–DnaC complex is independent of the structure of the phosphate group of the cofactor bound to the helicase. Thus, the DnaC protein eliminates the antagonistic allosteric effect of NTP and NDP on the ssDNA affinity of the enzyme. The protein changes the engagement of the DNA-binding subsites of the helicase in interactions with the nucleic acid, depending on the structure of the phosphate group of the present nucleotide cofactor and profoundly affects the structure of the bound DNA. Moreover, the ssDNA affinity of the helicase in the DnaB–DnaC complex is under the control of the nucleotide-binding site of the DnaC protein. The protein does not affect the NTP hydrolysis mechanism of the helicase. Nevertheless, the rate of the chemical step is diminished in the DnaB–DnaC complex. In the tertiary DnaB–DnaC–ssDNA complex, the ssDNA changes the internal dynamics between intermediates of the pyrimidine cofactor, in a manner independent of the base composition of the DNA, while the hydrolysis step of the purine cofactor is specifically stimulated by the homoadenosine ssDNA. The significance of these results for functional activities of the DnaB–DnaC complex is discussed. PMID:19432487

  14. The interplay of primer-template DNA phosphorylation status and single-stranded DNA binding proteins in directing clamp loaders to the appropriate polarity of DNA

    PubMed Central

    Hayner, Jaclyn N.; Douma, Lauren G.; Bloom, Linda B.

    2014-01-01

    Sliding clamps are loaded onto DNA by clamp loaders to serve the critical role of coordinating various enzymes on DNA. Clamp loaders must quickly and efficiently load clamps at primer/template (p/t) junctions containing a duplex region with a free 3′OH (3′DNA), but it is unclear how clamp loaders target these sites. To measure the Escherichia coli and Saccharomyces cerevisiae clamp loader specificity toward 3′DNA, fluorescent β and PCNA clamps were used to measure clamp closing triggered by DNA substrates of differing polarity, testing the role of both the 5′phosphate (5′P) and the presence of single-stranded binding proteins (SSBs). SSBs inhibit clamp loading by both clamp loaders on the incorrect polarity of DNA (5′DNA). The 5′P groups contribute selectivity to differing degrees for the two clamp loaders, suggesting variations in the mechanism by which clamp loaders target 3′DNA. Interestingly, the χ subunit of the E. coli clamp loader is not required for SSB to inhibit clamp loading on phosphorylated 5′DNA, showing that χ·SSB interactions are dispensable. These studies highlight a common role for SSBs in directing clamp loaders to 3′DNA, as well as uncover nuances in the mechanisms by which SSBs perform this vital role. PMID:25159615

  15. Structural features of DNA interaction with caffeine and theophylline

    NASA Astrophysics Data System (ADS)

    Nafisi, Shohreh; Manouchehri, Firouzeh; Tajmir-Riahi, Heidar-Ali; Varavipour, Maryam

    2008-03-01

    Caffeine and theophylline are strong antioxidants that prevent DNA damage. The anticancer and antiviral activities of these natural products are implicated in their mechanism of actions. However, there has been no information on the interactions of these xanthine derivatives with individual DNA at molecular level. The aim of this study was to examine the stability and structural features of calf-thymus DNA complexes with caffeine and theophylline in aqueous solution, using constant DNA concentration (6.25 mM) and various caffeine or theophylline/DNA(P) ratios of 1/80, 1/40, 1/20, 1/10, 1/5, 1/2 and 1/1. FTIR, UV-visible spectroscopic methods were used to determine the ligand external binding modes, the binding constant and the stability of caffeine, theophylline-DNA complexes in aqueous solution. Spectroscopic evidence showed that the complexation of caffeine and theophylline with DNA occurred via G-C and A-T and PO 2 group with overall binding constants of K(caffeine-DNA) = 9.7 × 10 3 M -1 and K(theophylline-DNA) = 1.7 × 10 4 M -1. The affinity of ligand-DNA binding is in the order of theophylline > caffeine. A partial B to A-DNA transition occurs upon caffeine and theophylline complexation.

  16. Conformational changes of the phenyl and naphthyl isocyanate-DNA adducts during DNA replication and by minor groove binding molecules

    PubMed Central

    Nakano, Shu-ichi; Uotani, Yuuki; Sato, Yuichi; Oka, Hirohito; Fujii, Masayuki; Sugimoto, Naoki

    2013-01-01

    DNA lesions produced by aromatic isocyanates have an extra bulky group on the nucleotide bases, with the capability of forming stacking interaction within a DNA helix. In this work, we investigated the conformation of the 2′-deoxyadenosine and 2′-deoxycytidine derivatives tethering a phenyl or naphthyl group, introduced in a DNA duplex. The chemical modification experiments using KMnO4 and 1-cyclohexyl-3 -(2-morpholinoethyl) carbodiimide metho-p-toluenesulfonate have shown that the 2′-deoxycytidine lesions form the base pair with guanine while the 2′-deoxyadenosine lesions have less ability of forming the base pair with thymine in solution. Nevertheless, the kinetic analysis shows that these DNA lesions are compatible with DNA ligase and DNA polymerase reactions, as much as natural DNA bases. We suggest that the adduct lesions have a capability of adopting dual conformations, depending on the difference in their interaction energies between stacking of the attached aromatic group and base pairing through hydrogen bonds. It is also presented that the attached aromatic groups change their orientation by interacting with the minor groove binding netropsin, distamycin and synthetic polyamide. The nucleotide derivatives would be useful for enhancing the phenotypic diversity of DNA molecules and for exploring new non-natural nucleotides. PMID:23873956

  17. Curating DNA specimens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    DNA data are used in a variety of ethnobiological disciplines including archaeology, conservation, ecology, medicinal plants and natural products research, taxonomy and systematics, crop evolution and domestication, and genetic diversity. It frequently is convenient to store and share DNA among coop...

  18. Routine DNA testing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Routine DNA testing. It’s done once you’ve Marker-Assisted Breeding Pipelined promising Qantitative Trait Loci within your own breeding program and thereby established the performance-predictive power of each DNA test for your germplasm under your conditions. By then you are ready to screen your par...

  19. Recombinant DNA for Teachers.

    ERIC Educational Resources Information Center

    Duvall, James G., III

    1992-01-01

    A science teacher describes his experience at a workshop to learn to teach the Cold Spring Harbor DNA Science Laboratory Protocols. These protocols lead students through processes for taking E. coli cells and transforming them into a new antibiotic resistant strain. The workshop featured discussions of the role of DNA recombinant technology in…

  20. MICROWAVE RESONANCES IN DNA

    EPA Science Inventory

    This report describes spectroscopic studies of DNA which were undertaken to better understand a physical basis for microwave absorption by this molecule. hree types of studies are described. ) The low frequency scattered light spectrum of DNA was studied by two methods. irst, Ram...

  1. Replicating repetitive DNA.

    PubMed

    Tognetti, Silvia; Speck, Christian

    2016-05-27

    The function and regulation of repetitive DNA, the 'dark matter' of the genome, is still only rudimentarily understood. Now a study investigating DNA replication of repetitive centromeric chromosome segments has started to expose a fascinating replication program that involves suppression of ATR signalling, in particular during replication stress. PMID:27230530

  2. Hydrogels: DNA bulks up

    NASA Astrophysics Data System (ADS)

    Labean, Thom

    2006-10-01

    Since the 1940s DNA has been known as the genetic material connected to heredity, and from the early 1980s it has also been considered as a potential structural material for nanoscale construction. Now, a hydrogel made entirely of DNA brings this molecule into the realm of bulk materials.

  3. Translesion DNA synthesis

    PubMed Central

    Vaisman, Alexandra; McDonald, John P.; Woodgate, Roger

    2014-01-01

    All living organisms are continually exposed to agents that damage their DNA, which threatens the integrity of their genome. As a consequence, cells are equipped with a plethora of DNA repair enzymes to remove the damaged DNA. Unfortunately, situations nevertheless arise where lesions persist, and these lesions block the progression of the cell’s replicase. Under these situations, cells are forced to choose between recombination-mediated “damage avoidance” pathways, or use a specialized DNA polymerase (pol) to traverse the blocking lesion. The latter process is referred to as Translesion DNA Synthesis (TLS). As inferred by its name, TLS not only results in bases being (mis)incorporated opposite DNA lesions, but also downstream of the replicase-blocking lesion, so as to ensure continued genome duplication and cell survival. Escherichia coli and Salmonella typhimurium possess five DNA polymerases, and while all have been shown to facilitate TLS under certain experimental conditions, it is clear that the LexA-regulated and damage-inducible pols II, IV and V perform the vast majority of TLS under physiological conditions. Pol V can traverse a wide range of DNA lesions and performs the bulk of mutagenic TLS, whereas pol II and pol IV appear to be more specialized TLS polymerases. PMID:26442823

  4. Characterization of muntjac DNA

    SciTech Connect

    Davis, R.C.

    1981-05-27

    Sister chromatid exchange (SCE) in muntjac chromosomes is generally proportional to the chromosomal DNA content, but the SCE frequency is reduced in the heterochromatic neck region of the X chromosome. The physical properties of muntjac DNA and the kinetics of repair of UV damage in muntjac heterochromatin and euchromatin were examined and compared with the distribution of sister chromatid exchange.

  5. DNA as information.

    PubMed

    Wills, Peter R

    2016-03-13

    This article reviews contributions to this theme issue covering the topic 'DNA as information' in relation to the structure of DNA, the measure of its information content, the role and meaning of information in biology and the origin of genetic coding as a transition from uninformed to meaningful computational processes in physical systems. PMID:26857666

  6. A radioimmunoassay method for detection of DNA based on chemical immobilization of anti-DNA antibody.

    PubMed

    Yoo, S K; Yoon, M; Park, U J; Han, H S; Kim, J H; Hwang, H J

    1999-09-30

    High selectivity provided by biomolecules such as antibodies and enzymes has been exploited during the last two decades for development of biosensors. Of particular importance are efficient immobilization methods for biomolecules in order to preserve their biological activities. In this study, we have evaluated immobilization strategies for an anti-DNA antibody on a self-assembled monolayer of omega-functionalized thiols. The antibody was immobilized via peptide bond formation between the primary amines in the antibody and the carboxyl groups on the self-assembled monolayer. The peptide bond coupling was achieved by activating COOH groups on the surface through N-Hydroxysuccimide (NHS)-ester formation, followed by acylation of NH2 group in the antibody. DNA binding activity of the immobilized antibody was examined by counting beta emission from 35S-labeled DNA. PMID:10551259

  7. Group B Strep Infection

    MedlinePlus

    ... Return to Web version Group B Strep Infection Overview What is group B strep? Group B streptococcus, or group B strep for short, is a certain kind of bacteria (germ) that lives in the intestine, rectum, and ...

  8. Premeltons in DNA.

    PubMed

    Sobell, Henry M

    2016-03-01

    Premeltons are examples of emergent-structures (i.e., structural-solitons) that arise spontaneously in DNA due to the presence of nonlinear-excitations in its structure. They are of two kinds: B-B (or A-A) premeltons form at specific DNA-regions to nucleate site-specific DNA melting. These are stationary and, being globally-nontopological, undergo breather-motions that allow drugs and dyes to intercalate into DNA. B-A (or A-B) premeltons, on the other hand, are mobile, and being globally-topological, act as phase-boundaries transforming B- into A-DNA during the structural phase-transition. They are not expected to undergo breather motions. A key feature of both types of premeltons is the presence of an intermediate structural-form in their central regions (proposed as being a transition-state intermediate in DNA-melting and in the B- to A-transition), which differs from either A- or B-DNA. Called beta-DNA, this is both metastable and hyperflexible--and contains an alternating sugar-puckering pattern along the polymer backbone combined with the partial unstacking (in its lower energy-forms) of every-other base-pair. Beta-DNA is connected to either B- or to A-DNA on either side by boundaries possessing a gradation of nonlinear structural-change, these being called the kink and the antikink regions. The presence of premeltons in DNA leads to a unifying theory to understand much of DNA physical chemistry and molecular biology. In particular, premeltons are predicted to define the 5' and 3' ends of genes in naked-DNA and DNA in active-chromatin, this having important implications for understanding physical aspects of the initiation, elongation and termination of RNA-synthesis during transcription. For these and other reasons, the model will be of broader interest to the general-audience working in these areas. The model explains a wide variety of data, and carries with it a number of experimental predictions--all readily testable--as will be described in this review

  9. Group Dynamic Processes in Email Groups

    ERIC Educational Resources Information Center

    Alpay, Esat

    2005-01-01

    Discussion is given on the relevance of group dynamic processes in promoting decision-making in email discussion groups. General theories on social facilitation and social loafing are considered in the context of email groups, as well as the applicability of psychodynamic and interaction-based models. It is argued that such theories may indeed…

  10. Interagency mechanical operations group numerical systems group

    SciTech Connect

    1997-09-01

    This report consists of the minutes of the May 20-21, 1971 meeting of the Interagency Mechanical Operations Group (IMOG) Numerical Systems Group. This group looks at issues related to numerical control in the machining industry. Items discussed related to the use of CAD and CAM, EIA standards, data links, and numerical control.

  11. Advances in DNA photonics

    NASA Astrophysics Data System (ADS)

    Heckman, Emily M.; Aga, Roberto S.; Fehrman Cory, Emily M.; Ouchen, Fahima; Lesko, Alyssa; Telek, Brian; Lombardi, Jack; Bartsch, Carrie M.; Grote, James G.

    2012-10-01

    In this paper we present our current research in exploring a DNA biopolymer for photonics applications. A new processing technique has been adopted that employs a modified soxhlet-dialysis (SD) rinsing technique to completely remove excess ionic contaminants from the DNA biopolymer, resulting in a material with greater mechanical stability and enhanced performance reproducibility. This newly processed material has been shown to be an excellent material for cladding layers in poled polymer electro-optic (EO) waveguide modulator applications. Thin film poling results are reported for materials using the DNA biopolymer as a cladding layer, as are results for beam steering devices also using the DNA biopolymer. Finally, progress on fabrication of a Mach Zehnder EO modulator with DNA biopolymer claddings using nanoimprint lithography techniques is reported.

  12. Archaeal DNA replication.

    PubMed

    Kelman, Lori M; Kelman, Zvi

    2014-01-01

    DNA replication is essential for all life forms. Although the process is fundamentally conserved in the three domains of life, bioinformatic, biochemical, structural, and genetic studies have demonstrated that the process and the proteins involved in archaeal DNA replication are more similar to those in eukaryal DNA replication than in bacterial DNA replication, but have some archaeal-specific features. The archaeal replication system, however, is not monolithic, and there are some differences in the replication process between different species. In this review, the current knowledge of the mechanisms governing DNA replication in Archaea is summarized. The general features of the replication process as well as some of the differences are discussed. PMID:25421597

  13. One-electron oxidation of DNA: mechanism and consequences.

    PubMed

    Schuster, Gary B

    2009-01-01

    All living organisms store the information necessary to maintain life in their DNA. Any process that damages DNA and causes loss or corruption of that information threatens the viability of the organism. One-electron oxidation is such a process. Loss of an electron from DNA generates a radical cation that is located primarily on its nucleobases. The radical cation migrates reversibly through duplex DNA by hopping until it is eventually trapped in an irreversible chemical reaction. The particular sequence of nucleobases in a DNA oligomer determines both the efficiency of hopping and the specific location and nature of the damaging chemical reaction. In its normal aqueous solutions, DNA is a polyanion because of the negative charge carried by its phosphate groups. Counter ions (typically Na(+)) to the phosphate groups play an important role in facilitating both the migration of the radical cation and in its eventual reaction with H(2)O. Irreversible reaction of a radical cation with H(2)O in duplex DNA occurs preferentially at the most reactive site. In normal DNA that is comprised of the four common DNA nucleobases, reaction occurs most commonly at a guanine and results in its conversion primarily to 8-oxo-7,8-dihydroguanine (8-OxoG). Both electronic and steric effects control the outcome of this process. If the DNA oligomer does not contain a suitable guanine, then reaction of the radical cation occurs at the thymine of a TT step primarily by a tandem process. The general outcomes revealed in the one-electron oxidation of DNA oligomers in solution appear to be generally valid also for more complex DNA structures and for the cellular DNA of living organisms. PMID:19749272

  14. DNA Align Editor: DNA Alignment Editor Tool

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The SNPAlignEditor is a DNA sequence alignment editor that runs on Windows platforms. The purpose of the program is to provide an intuitive, user-friendly tool for manual editing of multiple sequence alignments by providing functions for input, editing, and output of nucleotide sequence alignments....

  15. Developmental validation of the ParaDNA(®) Intelligence System-A novel approach to DNA profiling.

    PubMed

    Blackman, Stephen; Dawnay, Nick; Ball, Glyn; Stafford-Allen, Beccy; Tribble, Nicholas; Rendell, Paul; Neary, Kelsey; Hanson, Erin K; Ballantyne, Jack; Kallifatidis, Beatrice; Mendel, Julian; Mills, DeEtta K; Wells, Simon

    2015-07-01

    DNA profiling through the analysis of STRs remains one of the most widely used tools in human identification across the world. Current laboratory STR analysis is slow, costly and requires expert users and interpretation which can lead to instances of delayed investigations or non-testing of evidence on budget grounds. The ParaDNA(®) Intelligence System has been designed to provide a simple, fast and robust way to profile DNA samples in a lab or field-deployable manner. The system analyses 5-STRs plus amelogenin to deliver a DNA profile that enables users to gain rapid investigative leads and intelligent prioritisation of samples in human identity testing applications. Utilising an innovative sample collector, minimal training is required to enable both DNA analysts and nonspecialist personnel to analyse biological samples directly, without prior processing, in approximately 75min. The test uses direct PCR with fluorescent HyBeacon(®) detection of STR allele lengths to provide a DNA profile. The developmental validation study described here followed the Scientific Working Group on DNA Analysis Methods (SWGDAM) guidelines and tested the sensitivity, reproducibility, accuracy, inhibitor tolerance, and performance of the ParaDNA Intelligence System on a range of mock evidence items. The data collected demonstrate that the ParaDNA Intelligence System displays useful DNA profiles when sampling a variety of evidence items including blood, saliva, semen and touch DNA items indicating the potential to benefit a number of applications in fields such as forensic, military and disaster victim identification (DVI). PMID:25980941

  16. Activation of an Mg2+-dependent DNA endonuclease of avian myeloblastosis virus alpha beta DNA polymerase by in vitro proteolytic cleavage.

    PubMed Central

    Grandgenett, D P; Golomb, M; Vora, A C

    1980-01-01

    Partial chymotryptic digestion of purified avian myeloblastosis virus alpha beta DNA polymerase resulted in the activation of a Mg2+-dependent DNA endonuclease activity. Incubation of the polymerase-protease mixture in the presence of super-coiled DNA and Mg2+ permitted detection of the cleaved polymerase fragment possessing DNA nicking activity. Protease digestion conditions were established permitting selective cleavage of beta to alpha, which contained DNA polymerase and RNase H activity and to a family of polypeptides ranging in size from 30,000 to 34,000 daltons. These latter beta-unique fragments were purified by polyuridylate-Sepharose 4B chromatography and were shown to contain both DNA binding and DNA endonuclease activities. We have demonstrated that this group of polymerase fragments derived by chymotryptic digestion of alpha beta DNA polymerase is similar to the in vivo-isolated avian myeloblastosis virus p32pol in size, sequence, and DNA endonuclease activity. Images PMID:6154149

  17. A DNA structural atlas for Escherichia coli.

    PubMed

    Pedersen, A G; Jensen, L J; Brunak, S; Staerfeldt, H H; Ussery, D W

    2000-06-16

    location of promoters at the apex of plectonemically supercoiled DNA. We have also analyzed the structural similarities between groups of genes by clustering all RNA and protein-encoding genes in E. coli, based on the average structural parameters. We find that most ribosomal genes (protein-encoding as well as rRNA genes) cluster together, and we suggest that DNA structure may play a role in the transcription of these highly expressed genes. PMID:10843847

  18. Paternal DNA damage resulting from various sperm treatments persists after fertilization and is similar before and after DNA replication.

    PubMed

    Yamauchi, Yasuhiro; Riel, Jonathan M; Ward, Monika A

    2012-01-01

    In spite of its highly condensed state, sperm DNA is vulnerable to damage that can originate from oxidative stress, the activity of sperm-specific nucleases, or both. After fertilization, in the oocyte, paternal chromatin undergoes dramatic changes, and during this extensive remodeling, it can be both repaired and degraded, and these processes can be linked to DNA synthesis. Here, we analyzed sperm response to damage-inducing treatments both before and after fertilization and before or after zygotic DNA replication. Epididymal mouse spermatozoa were either frozen without cryoprotection (FT) or treated with detergent Triton X-100 coupled with dithiothreitol (TX+DTT) to induce DNA damage. Fresh, untreated sperm served as control. Immediately after preparation, spermatozoa from 3 groups were taken for comet assay, or for intracytoplasmic sperm injection into prometaphase I oocytes to visualize prematurely condensed single-chromatid chromosomes, or into mature metaphase II oocytes to visualize chromosomes after DNA replication. Comet assay revealed increased DNA fragmentation in treated sperm when compared with control, with FT sperm more severely affected. Chromosome analysis demonstrated paternal DNA damage in oocytes injected with treated, but not with fresh, sperm, with FT and TX+DTT groups now yielding similar damage. There were no differences in the incidence of abnormal paternal karyoplates before and after DNA synthesis in all examined groups. This study provides evidence that subjecting sperm to DNA damage-inducing treatments results in degradation of highly condensed sperm chromatin when it is still packed within the sperm head, and that this DNA damage persists after fertilization. The difference in DNA damage in sperm subjected to 2 treatments was ameliorated in the fertilized oocytes, suggesting that some chromatin repair might have occurred. This process, however, was independent of DNA synthesis and took place during oocyte maturation. PMID:21546611

  19. Introduction to Sporadic Groups

    NASA Astrophysics Data System (ADS)

    Boya, Luis J.

    2011-01-01

    This is an introduction to finite simple groups, in particular sporadic groups, intended for physicists. After a short review of group theory, we enumerate the 1+1+16=18 families of finite simple groups, as an introduction to the sporadic groups. These are described next, in three levels of increasing complexity, plus the six isolated ''pariah'' groups. The (old) five Mathieu groups make up the first, smallest order level. The seven groups related to the Leech lattice, including the three Conway groups, constitute the second level. The third and highest level contains the Monster group M, plus seven other related groups. Next a brief mention is made of the remaining six pariah groups, thus completing the 5+7+8+6=26 sporadic groups. The review ends up with a brief discussion of a few of physical applications of finite groups in physics, including a couple of recent examples which use sporadic groups.

  20. Studying DNA in the Classroom.

    ERIC Educational Resources Information Center

    Zarins, Silja

    1993-01-01

    Outlines a workshop for teachers that illustrates a method of extracting DNA and provides instructions on how to do some simple work with DNA without sophisticated and expensive equipment. Provides details on viscosity studies and breaking DNA molecules. (DDR)

  1. Simple & Safe Genomic DNA Isolation.

    ERIC Educational Resources Information Center

    Moss, Robert; Solomon, Sondra

    1991-01-01

    A procedure for purifying DNA using either bacteria or rat liver is presented. Directions for doing a qualitative DNA assay using diphenylamine and a quantitative DNA assay using spectroscopy are included. (KR)

  2. A search for specificity in DNA-drug interactions.

    PubMed

    Cruciani, G; Goodford, P J

    1994-06-01

    The GRID force field and a principal component analysis have been used in order to predict the interactions of small chemical groups with all 64 different triplet sequences of B-DNA. Factors that favor binding to guanine-cytosine base pairs have been identified, and a dictionary of ligand groups and their locations is presented as a guide to the design of specific DNA ligands. PMID:7918250

  3. Toxoplasma gondii infection can induce retinal DNA damage: an experimental study

    PubMed Central

    El-Sayed, Nagwa Mostafa; Aly, Eman Mohamed

    2014-01-01

    AIM To detect whether Toxoplasma gondii (T. gondii) infection of mice can induce retinal DNA damage. METHODS A total of 20 laboratory-bred male Swiss albino mice were used and divided into four groups: control group (non-infected animals); T. gondii infected group; immunosuppressed infected group; and infected group treated with sulfadiazine and pyrimethamine. Mice eyes were collected 6wk post infection and retinas were obtained. Each retina was immediately processed for comet assay and the frequency of tailed nuclei (DNA damage) was calculated. In addition, retinal DNA damage was revealed by various comet assay parameters that were provided by the image analysis software including tail length, percentage of DNA in the tail, percentage of tailed cells and tail moment. RESULTS The obtained results showed that T. gondii infection induced a statistically significant increase in the frequency of tailed nuclei, tail length, percentage of DNA in the tail, and tail moment in mice retinal cells compared to the control group (which showed some degree of DNA damage). In immunosuppressed infected group, retinal DNA damage was severing and there was significant increase in various comet assay parameters compared to both control and infected groups. After treatment with sulfadiazine and pyrimethamine, retinal DNA damage decreased and all comet assay parameters showed a statistical significant decrease compared to infected groups. CONCLUSION T. gondii infection can induce DNA damage in mice retinal cells. PMID:24967186

  4. DNA methyltransferase 1 mutations and mitochondrial pathology: is mtDNA methylated?

    PubMed Central

    Maresca, Alessandra; Zaffagnini, Mirko; Caporali, Leonardo; Carelli, Valerio; Zanna, Claudia

    2015-01-01

    Autosomal dominant cerebellar ataxia-deafness and narcolepsy (ADCA-DN) and Hereditary sensory neuropathy with dementia and hearing loss (HSN1E) are two rare, overlapping neurodegenerative syndromes that have been recently linked to allelic dominant pathogenic mutations in the DNMT1 gene, coding for DNA (cytosine-5)-methyltransferase 1 (DNMT1). DNMT1 is the enzyme responsible for maintaining the nuclear genome methylation patterns during the DNA replication and repair, thus regulating gene expression. The mutations responsible for ADCA-DN and HSN1E affect the replication foci targeting sequence domain, which regulates DNMT1 binding to chromatin. DNMT1 dysfunction is anticipated to lead to a global alteration of the DNA methylation pattern with predictable downstream consequences on gene expression. Interestingly, ADCA-DN and HSN1E phenotypes share some clinical features typical of mitochondrial diseases, such as optic atrophy, peripheral neuropathy, and deafness, and some biochemical evidence of mitochondrial dysfunction. The recent discovery of a mitochondrial isoform of DNMT1 and its proposed role in methylating mitochondrial DNA (mtDNA) suggests that DNMT1 mutations may directly affect mtDNA and mitochondrial physiology. On the basis of this latter finding the link between DNMT1 abnormal activity and mitochondrial dysfunction in ADCA-DN and HSN1E appears intuitive, however, mtDNA methylation remains highly debated. In the last years several groups demonstrated the presence of 5-methylcytosine in mtDNA by different approaches, but, on the other end, the opposite evidence that mtDNA is not methylated has also been published. Since over 1500 mitochondrial proteins are encoded by the nuclear genome, the altered methylation of these genes may well have a critical role in leading to the mitochondrial impairment observed in ADCA-DN and HSN1E. Thus, many open questions still remain unanswered, such as why mtDNA should be methylated, and how this process is regulated and

  5. Short thio-multi-walled carbon nanotubes and Au nanoparticles enhanced electrochemical DNA biosensor for DNA hybridization detection

    NASA Astrophysics Data System (ADS)

    Guo, Feng; Zhang, Jimei; Dai, Zhao; Zheng, Guo

    2010-07-01

    A novel and sensitive electrochemical DNA biosensor based on multi-walled carbon nanotubes functionalized with a thio group (MWNTs-SH) and gold nanoparticles (GNPs) for covalent DNA immobilization and enhanced hybridization detection is described. The key step for developing this novel DNA biosensor is to cut the pristine MWNT into short and generate lots of active sites simultaneously. With this approach, the target DNA could be quantified in a linear range from 8.5×10-10 to 1.5×10-5 mol/L, with a detection limit of 1.67×10-11 mol/L by 3σ.

  6. Molecular structure of r/GCG/d/TATACGC/ - A DNA-RNA hybrid helix joined to double helical DNA

    NASA Technical Reports Server (NTRS)

    Wang, A. H.-J.; Fujii, S.; Rich, A.; Van Boom, J. H.; Van Der Marel, G. A.; Van Boeckel, S. A. A.

    1982-01-01

    The molecule r(GCG)d(TATACGC) is self-complementary and forms two DNA-RNA hybrid segments surrounding a central region of double helical DNA; its molecular structure has been solved by X-ray analysis. All three parts of the molecule adopt a conformation which is close to that seen in the 11-fold RNA double helix. The conformation of the ribonucleotides is partly determined by water molecules bridging between the ribose O2' hydroxyl group and cytosine O2. The hybrid-DNA duplex junction contains no structural discontinuities. However, the central DNA TATA sequence has some structural irregularities.

  7. Dietary and lifestyle factors of DNA methylation.

    PubMed

    Lim, Unhee; Song, Min-Ae

    2012-01-01

    Lifestyle factors, such as diet, smoking, physical activity, and body weight management, are known to constitute the majority of cancer causes. Epigenetics has been widely proposed as a main mechanism that mediates the reversible effects of dietary and lifestyle factors on carcinogenesis. This chapter reviews human studies on potential dietary and lifestyle determinants of DNA methylation. Apart from a few prospective investigations and interventions of limited size and duration, evidence mostly comes from cross-sectional observational studies and supports some associations. Studies to date suggest that certain dietary components may alter genomic and gene-specific DNA methylation levels in systemic and target tissues, affecting genomic stability and transcription of tumor suppressors and oncogenes. Most data and supportive evidence exist for folate, a key nutritional factor in one-carbon metabolism that supplies the methyl units for DNA methylation. Other candidate bioactive food components include alcohol and other key nutritional factors of one-carbon metabolism, polyphenols and flavonoids in green tea, phytoestrogen, and lycopene. Some data also support a link of DNA methylation with physical activity and energy balance. Effects of dietary and lifestyle exposures on DNA methylation may be additionally modified by common genetic variants, environmental carcinogens, and infectious agents, an aspect that remains largely unexplored. In addition, growing literature supports that the environmental conditions during critical developmental stages may influence later risk of metabolic disorders in part through persistent programming of DNA methylation. Further research of these modifiable determinants of DNA methylation will improve our understanding of cancer etiology and may present certain DNA methylation markers as attractive surrogate endpoints for prevention research. Considering the plasticity of epigenetic marks and correlated nature of lifestyle factors, more

  8. DNA damage induction of ribonucleotide reductase.

    PubMed Central

    Elledge, S J; Davis, R W

    1989-01-01

    RNR2 encodes the small subunit of ribonucleotide reductase, the enzyme that catalyzes the first step in the pathway for the production of deoxyribonucleotides needed for DNA synthesis. RNR2 is a member of a group of genes whose activities are cell cycle regulated and that are transcriptionally induced in response to the stress of DNA damage. An RNR2-lacZ fusion was used to further characterize the regulation of RNR2 and the pathway responsible for its response to DNA damage. beta-Galactosidase activity in yeast strains containing the RNR2-lacZ fusion was inducible in response to DNA-damaging agents (UV light, 4-nitroquinoline-1-oxide [4-NQO], and methyl methanesulfonate [MMS]) and agents that block DNA replication (hydroxyurea [HU] and methotrexate) but not heat shock. When MATa cells were arrested in G1 by alpha-factor, RNR2 mRNA was still inducible by DNA damage, indicating that the observed induction can occur outside of S phase. In addition, RNR2 induction was not blocked by the presence of cycloheximide and is therefore likely to be independent of protein synthesis. A mutation, rnr2-314, was found to confer hypersensitivity to HU and increased sensitivity to MMS. In rnr2-314 mutant strains, the DNA damage stress response was found to be partially constitutive as well as hypersensitive to induction by HU but not MMS. The induction properties of RNR2 were examined in a rad4-2 mutant background; in this genetic background, RNR2 was hypersensitive to induction by 4-NQO but not MMS. Induction of the RNR2-lacZ fusion in a RAD(+) strain in response to 4-NQO was not enhanced by the presence of an equal number of rad4-2 cells that lacked the fusion, implying that the DNA damage stress response in cell autonomous. Images PMID:2513480

  9. Quantitive DNA Fiber Mapping

    SciTech Connect

    Lu, Chun-Mei; Wang, Mei; Greulich-Bode, Karin M.; Weier, Jingly F.; Weier, Heinz-Ulli G.

    2008-01-28

    Several hybridization-based methods used to delineate single copy or repeated DNA sequences in larger genomic intervals take advantage of the increased resolution and sensitivity of free chromatin, i.e., chromatin released from interphase cell nuclei. Quantitative DNA fiber mapping (QDFM) differs from the majority of these methods in that it applies FISH to purified, clonal DNA molecules which have been bound with at least one end to a solid substrate. The DNA molecules are then stretched by the action of a receding meniscus at the water-air interface resulting in DNA molecules stretched homogeneously to about 2.3 kb/{micro}m. When non-isotopically, multicolor-labeled probes are hybridized to these stretched DNA fibers, their respective binding sites are visualized in the fluorescence microscope, their relative distance can be measured and converted into kilobase pairs (kb). The QDFM technique has found useful applications ranging from the detection and delineation of deletions or overlap between linked clones to the construction of high-resolution physical maps to studies of stalled DNA replication and transcription.

  10. Reversible DNA compaction.

    PubMed

    González-Pérez, Alfredo

    2014-01-01

    In this review we summarize and discuss the different methods we can use to achieve reversible DNA compaction in vitro. Reversible DNA compaction is a natural process that occurs in living cells and viruses. As a result these process long sequences of DNA can be concentrated in a small volume (compacted) to be decompacted only when the information carried by the DNA is needed. In the current work we review the main artificial compacting agents looking at their suitability for decompaction. The different approaches used for decompaction are strongly influenced by the nature of the compacting agent that determines the mechanism of compaction. We focus our discussion on two main artificial compacting agents: multivalent cations and cationic surfactants that are the best known compacting agents. The reversibility of the process can be achieved by adding chemicals like divalent cations, alcohols, anionic surfactants, cyclodextrins or by changing the chemical nature of the compacting agents via pH modifications, light induced conformation changes or by redox-reactions. We stress the relevance of electrostatic interactions and self-assembly as a main approach in order to tune up the DNA conformation in order to create an on-off switch allowing a transition between coil and compact states. The recent advances to control DNA conformation in vitro, by means of molecular self-assembly, result in a better understanding of the fundamental aspects involved in the DNA behavior in vivo and serve of invaluable inspiration for the development of potential biomedical applications. PMID:24444152

  11. A new structural insight into XPA–DNA interactions

    PubMed Central

    Hilton, Benjamin; Shkriabai, Nick; Musich, Phillip R.; Kvaratskhelia, Mamuka; Shell, Steven; Zou, Yue

    2014-01-01

    XPA (xeroderma pigmentosum group A) protein is an essential factor for NER (nucleotide excision repair) which is believed to be involved in DNA damage recognition/verification, NER factor recruiting and stabilization of repair intermediates. Past studies on the structure of XPA have focused primarily on XPA interaction with damaged DNA. However, how XPA interacts with other DNA structures remains unknown though recent evidence suggest that these structures could be important for its roles in both NER and non-NER activities. Previously, we reported that XPA recognizes undamaged DNA ds/ssDNA (double-strand/single-strandDNA) junctions with a binding affinity much higher than its ability to bind bulky DNA damage. To understand how this interaction occurs biochemically we implemented a structural determination of the interaction using a MS-based protein footprinting method and limited proteolysis. By monitoring surface accessibility of XPA lysines to NHS-biotin modification in the free protein and the DNA junction-bound complex we show that XPA physically interacts with the DNA junctions via two lysines, K168 and K179, located in the previously known XPA(98–219) DBD (DNA-binding domain). Importantly, we also uncovered new lysine residues, outside of the known DBD, involved in the binding. We found that residues K221, K222, K224 and K236 in the C-terminal domain are involved in DNA binding. Limited proteolysis analysis of XPA–DNA interactions further confirmed this observation. Structural modelling with these data suggests a clamp-like DBD for the XPA binding to ds/ssDNA junctions. Our results provide a novel structure-function view of XPA–DNA junction interactions. PMID:25385088

  12. Tracking Mitochondrial DNA In Situ.

    PubMed

    Ligasová, Anna; Koberna, Karel

    2016-01-01

    The methods of the detection of (1) non-labeled and (2) BrdU-labeled mitochondrial DNA (mtDNA) are described. They are based on the production of singlet oxygen by monovalent copper ions and the subsequent induction of DNA gaps. The ends of interrupted DNA serve as origins for the labeling of mtDNA by DNA polymerase I or they are utilized by exonuclease that degrades DNA strands, unmasking BrdU in BrdU-labeled DNA. Both methods are sensitive approaches without the need of additional enhancement of the signal or the use of highly sensitive optical systems. PMID:26530676

  13. Chimeric proteins for detection and quantitation of DNA mutations, DNA sequence variations, DNA damage and DNA mismatches

    DOEpatents

    McCutchen-Maloney, Sandra L.

    2002-01-01

    Chimeric proteins having both DNA mutation binding activity and nuclease activity are synthesized by recombinant technology. The proteins are of the general formula A-L-B and B-L-A where A is a peptide having DNA mutation binding activity, L is a linker and B is a peptide having nuclease activity. The chimeric proteins are useful for detection and identification of DNA sequence variations including DNA mutations (including DNA damage and mismatches) by binding to the DNA mutation and cutting the DNA once the DNA mutation is detected.

  14. The DNA-bending protein HMG-1 enhances progesterone receptor binding to its target DNA sequences.

    PubMed Central

    Oñate, S A; Prendergast, P; Wagner, J P; Nissen, M; Reeves, R; Pettijohn, D E; Edwards, D P

    1994-01-01

    Steroid hormone receptors are ligand-dependent transcriptional activators that exert their effects by binding as dimers to cis-acting DNA sequences termed hormone response elements. When human progesterone receptor (PR), expressed as a full-length protein in a baculovirus system, was purified to homogeneity, it retained its ability to bind hormonal ligand and to dimerize but exhibited a dramatic loss in DNA binding activity for specific progesterone response elements (PREs). Addition of nuclear extracts from several cellular sources restored DNA binding activity, suggesting that PR requires a ubiquitous accessory protein for efficient interaction with specific DNA sequences. Here we have demonstrated that the high-mobility-group chromatin protein HMG-1, as a highly purified protein, dramatically enhanced binding of purified PR to PREs in gel mobility shift assays. This effect appeared to be highly selective for HMG-1, since a number of other nonspecific proteins failed to enhance PRE binding. Moreover, HMG-1 was effective when added in stoichiometric amounts with receptor, and it was capable of enhancing the DNA binding of both the A and B amino-terminal variants of PR. The presence of HMG-1 measurably increased the binding affinity of purified PR by 10-fold when a synthetic palindromic PRE was the target DNA. The increase in binding affinity for a partial palindromic PRE present in natural target genes was greater than 10-fold. Coimmunoprecipitation assays using anti-PR or anti-HMG-1 antibodies demonstrated that both PR and HMG-1 are present in the enhanced complex with PRE. HMG-1 protein has two conserved DNA binding domains (A and B), which recognize DNA structure rather than specific sequences. The A- or B-box domain expressed and purified from Escherichia coli independently stimulated the binding of PR to PRE, and the B box was able to functionally substitute for HMG-1 in enhancing PR binding. DNA ligase-mediated ring closure assays demonstrated that both the

  15. DNA-PK assay

    DOEpatents

    Anderson, Carl W.; Connelly, Margery A.

    2004-10-12

    The present invention provides a method for detecting DNA-activated protein kinase (DNA-PK) activity in a biological sample. The method includes contacting a biological sample with a detectably-labeled phosphate donor and a synthetic peptide substrate defined by the following features to provide specific recognition and phosphorylation by DNA-PK: (1) a phosphate-accepting amino acid pair which may include serine-glutamine (Ser-Gln) (SQ), threonine-glutamine (Thr-Gln) (TQ), glutamine-serine (Gln-Ser) (QS), or glutamine-threonine (Gln-Thr) (QT); (2) enhancer amino acids which may include glutamic acid or glutamine immediately adjacent at the amino- or carboxyl- side of the amino acid pair and forming an amino acid pair-enhancer unit; (3) a first spacer sequence at the amino terminus of the amino acid pair-enhancer unit; (4) a second spacer sequence at the carboxyl terminus of the amino acid pair-enhancer unit, which spacer sequences may include any combination of amino acids that does not provide a phosphorylation site consensus sequence motif; and, (5) a tag moiety, which may be an amino acid sequence or another chemical entity that permits separating the synthetic peptide from the phosphate donor. A compostion and a kit for the detection of DNA-PK activity are also provided. Methods for detecting DNA, protein phosphatases and substances that alter the activity of DNA-PK are also provided. The present invention also provides a method of monitoring protein kinase and DNA-PK activity in living cells. -A composition and a kit for monitoring protein kinase activity in vitro and a composition and a kit for monitoring DNA-PK activities in living cells are also provided. A method for identifying agents that alter protein kinase activity in vitro and a method for identifying agents that alter DNA-PK activity in living cells are also provided.

  16. DNA Damage among Wood Workers Assessed with the Comet Assay.

    PubMed

    Bruschweiler, Evin Danisman; Wild, Pascal; Huynh, Cong Khanh; Savova-Bianchi, Dessislava; Danuser, Brigitta; Hopf, Nancy B

    2016-01-01

    Exposure to wood dust, a human carcinogen, is common in wood-related industries, and millions of workers are occupationally exposed to wood dust worldwide. The comet assay is a rapid, simple, and sensitive method for determining DNA damage. The objective of this study was to investigate the DNA damage associated with occupational exposure to wood dust using the comet assay (peripheral blood samples) among nonsmoking wood workers (n = 31, furniture and construction workers) and controls (n = 19). DNA damage was greater in the group exposed to composite wood products compared to the group exposed to natural woods and controls (P < 0.001). No difference in DNA damage was observed between workers exposed to natural woods and controls (P = 0.13). Duration of exposure and current dust concentrations had no effect on DNA damage. In future studies, workers' exposures should include cumulative dust concentrations and exposures originating from the binders used in composite wood products. PMID:27398027

  17. DNA Damage among Wood Workers Assessed with the Comet Assay

    PubMed Central

    Bruschweiler, Evin Danisman; Wild, Pascal; Huynh, Cong Khanh; Savova-Bianchi, Dessislava; Danuser, Brigitta; Hopf, Nancy B.

    2016-01-01

    Exposure to wood dust, a human carcinogen, is common in wood-related industries, and millions of workers are occupationally exposed to wood dust worldwide. The comet assay is a rapid, simple, and sensitive method for determining DNA damage. The objective of this study was to investigate the DNA damage associated with occupational exposure to wood dust using the comet assay (peripheral blood samples) among nonsmoking wood workers (n = 31, furniture and construction workers) and controls (n = 19). DNA damage was greater in the group exposed to composite wood products compared to the group exposed to natural woods and controls (P < 0.001). No difference in DNA damage was observed between workers exposed to natural woods and controls (P = 0.13). Duration of exposure and current dust concentrations had no effect on DNA damage. In future studies, workers’ exposures should include cumulative dust concentrations and exposures originating from the binders used in composite wood products. PMID:27398027

  18. Diet-related DNA adduct formation in relation to carcinogenesis.

    PubMed

    Hemeryck, Lieselot Y; Vanhaecke, Lynn

    2016-08-01

    The human diet contributes significantly to the initiation and promotion of carcinogenesis. It has become clear that the human diet contains several groups of natural foodborne chemicals that are at least in part responsible for the genotoxic, mutagenic, and carcinogenic potential of certain foodstuffs. Electrophilic chemicals are prone to attack nucleophilic sites in DNA, resulting in the formation of altered nucleobases, also known as DNA adducts. Since DNA adduct formation is believed to signal the onset of chemically induced carcinogenesis, the DNA adduct-inducing potential of certain foodstuffs has been investigated to gain more insight into diet-related pathways of carcinogenesis. Many studies have investigated diet-related DNA adduct formation. This review summarizes work on known or suspected dietary carcinogens and the role of DNA adduct formation in hypothesized carcinogenesis pathways. PMID:27330144

  19. DNA Mismatch Repair

    PubMed Central

    MARINUS, M. G.

    2014-01-01

    DNA mismatch repair functions to correct replication errors in newly synthesized DNA and to prevent recombination between related, but not identical (homeologous), DNA sequences. The mechanism of mismatch repair is best understood in Escherichia coli and is the main focus of this review. The early genetic studies of mismatch repair are described as a basis for the subsequent biochemical characterization of the system. The effects of mismatch repair on homologous and homeologous recombination are described. The relationship of mismatch repair to cell toxicity induced by various drugs is included. The VSP (Very Short Patch) repair system is described in detail. PMID:26442827

  20. Focus: DNA probes

    SciTech Connect

    Not Available

    1986-11-01

    Progress in the development of DNA probes for the identification and quantitation of specific genetic sequences in biological samples is reviewed. Current research efforts in the development of DNA probes for the diagnosis of a wide variety of bacterial, viral, and other infectious diseases, such as herpes simplex and cytomegalovirus, and inherited genetic diseases such as cystic fibrosis and sickle cell anemia are discussed. Progress in development of DNA probe assays for cancer diagnosis, detection of Salmonella food poisoning, tissue typing (detection of histocompatibility antigens), mutagen screening, and animal diseases, among other applications is included.

  1. Structure of large dsDNA viruses

    PubMed Central

    Klose, Thomas; Rossmann, Michael G.

    2015-01-01

    Nucleocytoplasmic large dsDNA viruses (NCLDVs) encompass an ever-increasing group of large eukaryotic viruses, infecting a wide variety of organisms. The set of core genes shared by all these viruses includes a major capsid protein with a double jelly-roll fold forming an icosahedral capsid, which surrounds a double layer membrane that contains the viral genome. Furthermore, some of these viruses, such as the members of the Mimiviridae and Phycodnaviridae have a unique vertex that is used during infection to transport DNA into the host. PMID:25003382

  2. Phylogenetic Analysis of Shewanella Strains by DNA Relatedness Derived from Whole Genome Microarray DNA-DNA Hybridization and Comparison with Other Methods

    SciTech Connect

    Wu, Liyou; Yi, T. Y.; Van Nostrand, Joy; Zhou, Jizhong

    2010-05-17

    Phylogenetic analyses were done for the Shewanella strains isolated from Baltic Sea (38 strains), US DOE Hanford Uranium bioremediation site [Hanford Reach of the Columbia River (HRCR), 11 strains], Pacific Ocean and Hawaiian sediments (8 strains), and strains from other resources (16 strains) with three out group strains, Rhodopseudomonas palustris, Clostridium cellulolyticum, and Thermoanaerobacter ethanolicus X514, using DNA relatedness derived from WCGA-based DNA-DNA hybridizations, sequence similarities of 16S rRNA gene and gyrB gene, and sequence similarities of 6 loci of Shewanella genome selected from a shared gene list of the Shewanella strains with whole genome sequenced based on the average nucleotide identity of them (ANI). The phylogenetic trees based on 16S rRNA and gyrB gene sequences, and DNA relatedness derived from WCGA hybridizations of the tested Shewanella strains share exactly the same sub-clusters with very few exceptions, in which the strains were basically grouped by species. However, the phylogenetic analysis based on DNA relatedness derived from WCGA hybridizations dramatically increased the differentiation resolution at species and strains level within Shewanella genus. When the tree based on DNA relatedness derived from WCGA hybridizations was compared to the tree based on the combined sequences of the selected functional genes (6 loci), we found that the resolutions of both methods are similar, but the clustering of the tree based on DNA relatedness derived from WMGA hybridizations was clearer. These results indicate that WCGA-based DNA-DNA hybridization is an idea alternative of conventional DNA-DNA hybridization methods and it is superior to the phylogenetics methods based on sequence similarities of single genes. Detailed analysis is being performed for the re-classification of the strains examined.

  3. DNA as a Binary Code: How the Physical Structure of Nucleotide Bases Carries Information

    ERIC Educational Resources Information Center

    McCallister, Gary

    2005-01-01

    The DNA triplet code also functions as a binary code. Because double-ring compounds cannot bind to double-ring compounds in the DNA code, the sequence of bases classified simply as purines or pyrimidines can encode for smaller groups of possible amino acids. This is an intuitive approach to teaching the DNA code. (Contains 6 figures.)

  4. Crystallization and X-ray diffraction analysis of the DNA-remodelling protein DnaD from Bacillus subtilis

    SciTech Connect

    Schneider, Sabine; Carneiro, Maria J. V. M.; Ioannou, Charikleia; Soultanas, Panos; Paoli, Max

    2007-02-01

    Crystallization and preliminary X-ray diffraction analysis of the two domains of DnaD from B. subtilis is reported. The DnaD protein is an essential component of the chromosome-replication machinery of the Gram-positive bacterium Bacillus subtilis and is part of the primosomal cascade that ultimately loads the replicative ring helicase DnaC onto DNA. Moreover, DnaD is a global regulator of DNA architecture, as it forms higher order nucleoprotein structures in order to open supercoiled DNA. Here, the crystallization and preliminary X-ray diffraction analysis of the two domains of DnaD from B. subtilis are reported. Crystals of the N-terminal domain are trigonal, with either P3{sub 1}21 or P3{sub 2}21 space-group symmetry, and diffracted X-rays to 2.0 Å resolution; crystals of the C-terminal domain are hexagonal, with space group P6{sub 1} or P6{sub 5}, and diffracted X-rays to 2.9 Å resolution in-house. Determination of the structure of the DnaD domains will provide insight into how remodelling of the nucleoid is associated with priming of replication in the model Gram-positive organism B. subtilis.

  5. Characterisation of DNA forms associated with cassava latent virus infection.

    PubMed Central

    Stanley, J; Townsend, R

    1985-01-01

    In addition to the major encapsidated DNA species found in preparations of cassava latent virus (genomic DNAs 1 and 2) there are minor DNA populations of twice (dimeric) and approximately half genome length. Both minor species resemble the genomic DNAs in that they are composed of predominantly circular single-stranded DNA. All of these size groups have a corresponding covalently-closed circular double-stranded DNA form in infected tissue. Infectivity studies using cloned DNAs 1 and 2 show that dimeric DNA routinely appears, suggesting it to be an intermediate in the DNA replicative cycle that can be encapsidated at low efficiency. In contrast, half unit length DNA has not yet been detected after multiple passaging of virus derived from the cloned DNA inoculum. Half unit length DNAs appear to be derived exclusively from DNA 2 and consist of a population of molecules exhibiting a relatively specific deletion. As they have an inhibitory effect on virus multiplication, their encapsidated forms are analogous to defective interfering particles associated with other eukaryotic DNA containing viruses. Small primer molecules associated with the genomic single-stranded DNAs, as reported for another geminivirus, have not been detected in CLV. Images PMID:4000956

  6. Formation of phosphonester bonds catalyzed by DNA polymerase.

    PubMed Central

    Victorova, L S; Dyatkina, N B; Mozzherin DJu; Atrazhev, A M; Krayevsky, A A; Kukhanova, M K

    1992-01-01

    3'-Fluoro-2',3'-dideoxythymidine 5'-(alpha-methylphosphonyl)-beta,gamma- diphosphate and 2'-deoxythymidine-5'-(alpha-methylphosphonyl)-beta, gamma- diphosphate have been synthesized. Both compounds are incorporated into DNA chains during catalysis by reverse transcriptases of human immunodeficiency (HIV) and avian myeloblastosis (AMV) viruses, DNA polymerase beta from rat liver, terminal deoxynucleotidyl transferase from calf thymus and (at a very low rate) is by E. coli DNA polymerase I, Klenow fragment. The first compound is a termination substrate while the second is capable of multiple incorporation into the DNA chains. For instance, reverse transcriptase catalysis resulted in the appearance of 8 residues of second compound. DNA polymerases alpha and epsilon from human placenta incorporated none of the above compounds into DNA chains, although an inhibition of DNA synthesis by both compounds was observed with all enzymes mentioned. The 3'----5'-exonuclease activity of DNA polymerase I, Klenow fragment, hydrolyzed DNA fragments containing phosphonomethyl internucleoside groups, while such DNA fragments were resistant to the E. coli exonuclease III. Images PMID:1371865

  7. Influence of killing method on Lepidoptera DNA barcode recovery.

    PubMed

    Willows-Munro, Sandi; Schoeman, M Corrie

    2015-05-01

    The global DNA barcoding initiative has revolutionized the field of biodiversity research. Such large-scale sequencing projects require the collection of large numbers of specimens, which need to be killed and preserved in a way that is both DNA-friendly and which will keep voucher specimens in good condition for later study. Factors such as time since collection, correct storage (exposure to free water and heat) and DNA extraction protocol are known to play a role in the success of downstream molecular applications. Limited data are available on the most efficient, DNA-friendly protocol for killing. In this study, we evaluate the quality of DNA barcode (cytochrome oxidase I) sequences amplified from DNA extracted from specimens collected using three different killing methods (ethyl acetate, cyanide and freezing). Previous studies have suggested that chemicals, such as ethyl acetate and formaldehyde, degraded DNA and as such may not be appropriate for the collection of insects for DNA-based research. All Lepidoptera collected produced DNA barcodes of good quality, and our study found no clear difference in nucleotide signal strength, probability of incorrect base calling and phylogenetic utility among the three different treatment groups. Our findings suggest that ethyl acetate, cyanide and freezing can all be used to collect specimens for DNA analysis. PMID:25229871

  8. DNA from soil mirrors plant taxonomic and growth form diversity.

    PubMed

    Yoccoz, N G; Bråthen, K A; Gielly, L; Haile, J; Edwards, M E; Goslar, T; Von Stedingk, H; Brysting, A K; Coissac, E; Pompanon, F; Sønstebø, J H; Miquel, C; Valentini, A; De Bello, F; Chave, J; Thuiller, W; Wincker, P; Cruaud, C; Gavory, F; Rasmussen, M; Gilbert, M T P; Orlando, L; Brochmann, C; Willerslev, E; Taberlet, P

    2012-08-01

    Ecosystems across the globe are threatened by climate change and human activities. New rapid survey approaches for monitoring biodiversity would greatly advance assessment and understanding of these threats. Taking advantage of next-generation DNA sequencing, we tested an approach we call metabarcoding: high-throughput and simultaneous taxa identification based on a very short (usually <100 base pairs) but informative DNA fragment. Short DNA fragments allow the use of degraded DNA from environmental samples. All analyses included amplification using plant-specific versatile primers, sequencing and estimation of taxonomic diversity. We tested in three steps whether degraded DNA from dead material in soil has the potential of efficiently assessing biodiversity in different biomes. First, soil DNA from eight boreal plant communities located in two different vegetation types (meadow and heath) was amplified. Plant diversity detected from boreal soil was highly consistent with plant taxonomic and growth form diversity estimated from conventional above-ground surveys. Second, we assessed DNA persistence using samples from formerly cultivated soils in temperate environments. We found that the number of crop DNA sequences retrieved strongly varied with years since last cultivation, and crop sequences were absent from nearby, uncultivated plots. Third, we assessed the universal applicability of DNA metabarcoding using soil samples from tropical environments: a large proportion of species and families from the study site were efficiently recovered. The results open unprecedented opportunities for large-scale DNA-based biodiversity studies across a range of taxonomic groups using standardized metabarcoding approaches. PMID:22507540

  9. DNA Nanostructures on Membranes as Tools for Synthetic Biology.

    PubMed

    Czogalla, Aleksander; Franquelim, Henri G; Schwille, Petra

    2016-04-26

    Over the last decade, functionally designed DNA nanostructures applied to lipid membranes prompted important achievements in the fields of biophysics and synthetic biology. Taking advantage of the universal rules for self-assembly of complementary oligonucleotides, DNA has proven to be an extremely versatile biocompatible building material on the nanoscale. The possibility to chemically integrate functional groups into oligonucleotides, most notably with lipophilic anchors, enabled a widespread usage of DNA as a viable alternative to proteins with respect to functional activity on membranes. As described throughout this review, hybrid DNA-lipid nanostructures can mediate events such as vesicle docking and fusion, or selective partitioning of molecules into phase-separated membranes. Moreover, the major benefit of DNA structural constructs, such as DNA tiles and DNA origami, is the reproducibility and simplicity of their design. DNA nanotechnology can produce functional structures with subnanometer precision and allow for a tight control over their biochemical functionality, e.g., interaction partners. DNA-based membrane nanopores and origami structures able to assemble into two-dimensional networks on top of lipid bilayers are recent examples of the manifold of complex devices that can be achieved. In this review, we will shortly present some of the potentially most relevant avenues and accomplishments of membrane-anchored DNA nanostructures for investigating, engineering, and mimicking lipid membrane-related biophysical processes. PMID:27119630

  10. The DNA repair endonuclease Mus81 facilitates fast DNA replication in the absence of exogenous damage

    PubMed Central

    Fu, Haiqing; Martin, Melvenia M.; Regairaz, Marie; Huang, Liang; You, Yang; Lin, Chi-Mei; Ryan, Michael; Kim, RyangGuk; Shimura, Tsutomu; Pommier, Yves; Aladjem, Mirit I.

    2015-01-01

    The Mus81 endonuclease resolves recombination intermediates and mediates cellular responses to exogenous replicative stress. Here, we show that Mus81 also regulates the rate of DNA replication during normal growth by promoting replication fork progression while reducing the frequency of replication initiation events. In the absence of Mus81 endonuclease activity, DNA synthesis is slowed and replication initiation events are more frequent. In addition, Mus81 deficient cells fail to recover from exposure to low doses of replication inhibitors and cell viability is dependent on the XPF endonuclease. Despite an increase in replication initiation frequency, cells lacking Mus81 use the same pool of replication origins as Mus81-expressing cells. Therefore, decelerated DNA replication in Mus81 deficient cells does not initiate from cryptic or latent origins not used during normal growth. These results indicate that Mus81 plays a key role in determining the rate of DNA replication without activating a novel group of replication origins. PMID:25879486

  11. Small Group Research

    ERIC Educational Resources Information Center

    McGrath, Joseph E.

    1978-01-01

    Summarizes research on small group processes by giving a comprehensive account of the types of variables primarily studied in the laboratory. These include group structure, group composition, group size, and group relations. Considers effects of power, leadership, conformity to social norms, and role relationships. (Author/AV)

  12. Interaction of DNA and DNA-anti-DNA complexes to fibronectin

    SciTech Connect

    Gupta, R.C.; Simpson, W.A.; Raghow, R.; Hasty, K.

    1986-03-01

    Fibronectin (Fn) is a large multidomain glycoprotein found in the basement membrane, on cell surface and in plasma. The interactions of Fn with DNA may be significant in glomerular deposition of DNA-anti-DNA complexes in patients with systemic lupus erythematosus (SLE). The authors examined the binding of DNA and DNA-anti-DNA complexes to Fn by a solid phase assay in which Fn was coated to microtiter plates and reacted with (/sup 3/H)DNA or DNA complexes with a monoclonal anti-DNA antibody. The optimal interaction of DNA with Fn occurs at <0.1M NaCl suggesting that the binding is charge dependent; the specificity of this binding was shown by competitive inhibition and locking experiments using anti-Fn. The binding was maximum at pH 6.5 and in the absence of Ca/sup 2 +/. The addition of Clq enhanced the binding of DNA and DNA-anti-DNA complexes to Fn, whereas heparan sulfate inhibited such binding. The monomeric or aggregated IgC did not bind Fn but aggregated IgG bound to Fn in the presence of Clq. Furthermore, DNA-anti-DNA complexes in sera from active SLE patients bound Fn which was enhanced in the presence of Clq; DNase abolished this binding indicating that the interaction of these complexes was mediated by DNA. These observations may partially explain the molecular mechanism(s) of the deposition of DNA-anti-DNA complexes in basement membrane.

  13. Expansion of the DNA Alphabet beyond Natural DNA Recognition.

    PubMed

    Tateishi-Karimata, Hisae; Sugimoto, Naoki

    2016-07-15

    Simple and inexpensive DNA fibres: New, stable DNA structures are created by the binding of a small molecule to poly(A). Because these DNA fibres are formed from inexpensive materials by using very simple methods, DNA materials suitable for practical use such as information storage should be possible in the near future. PMID:27061868

  14. DNA computing in microreactors

    NASA Astrophysics Data System (ADS)

    Wagler, Patrick; van Noort, Danny; McCaskill, John S.

    2001-11-01

    The goal of this research is to improve the modular stability and programmability of DNA-based computers and in a second step towards optical programmable DNA computing. The main focus here is on hydrodynamic stability. Clockable microreactors can be connected in various ways to solve combinatorial optimisation problems, such as Maximum Clique or 3-SAT. This work demonstrates by construction how one micro-reactor design can be programmed to solve any instance of Maximum Clique up to its given maximum size (N). It reports on an implementation of the architecture proposed previously. This contrasts with conventional DNA computing where the individual sequence of biochemical operations depends on the specific problem. In this pilot study we are tackling a graph for the Maximum Clique problem with NDNA solution space will be presented, which is symbolized by a set of bit-strings (words).

  15. DNA sequencing conference, 2

    SciTech Connect

    Cook-Deegan, R.M.; Venter, J.C.; Gilbert, W.; Mulligan, J.; Mansfield, B.K.

    1991-06-19

    This conference focused on DNA sequencing, genetic linkage mapping, physical mapping, informatics and bioethics. Several were used to study this sequencing and mapping. This article also discusses computer hardware and software aiding in the mapping of genes.

  16. Close encounters with DNA

    PubMed Central

    Maffeo, C.; Yoo, J.; Comer, J.; Wells, D. B.; Luan, B.; Aksimentiev, A.

    2014-01-01

    Over the past ten years, the all-atom molecular dynamics method has grown in the scale of both systems and processes amenable to it and in its ability to make quantitative predictions about the behavior of experimental systems. The field of computational DNA research is no exception, witnessing a dramatic increase in the size of systems simulated with atomic resolution, the duration of individual simulations and the realism of the simulation outcomes. In this topical review, we describe the hallmark physical properties of DNA from the perspective of all-atom simulations. We demonstrate the amazing ability of such simulations to reveal the microscopic physical origins of experimentally observed phenomena and we review the frustrating limitations associated with imperfections of present atomic force fields and inadequate sampling. The review is focused on the following four physical properties of DNA: effective electric charge, response to an external mechanical force, interaction with other DNA molecules and behavior in an external electric field. PMID:25238560

  17. FBI's DNA analysis program

    NASA Astrophysics Data System (ADS)

    Brown, John R.

    1994-03-01

    Forensic DNA profiling technology is a significant law enforcement tool due to its superior discriminating power. Applying the principles of population genetics to the DNA profile obtained in violent crime investigations results in low frequency of occurrence estimates for the DNA profile. These estimates often range from a frequency of occurrence of 1 in 50 unrelated individuals to 1 in a million unrelated individuals or even smaller. It is this power to discriminate among individuals in the population that has propelled forensic DNA technology to the forefront of forensic testing in violent crime cases. Not only is the technology extremely powerful in including or excluding a criminal suspect as the perpetrator, but it also gives rise to the potential of identifying criminal suspects in cases where the investigators of unknown suspect cases have exhausted all other available leads.

  18. Making DNA Fingerprints.

    ERIC Educational Resources Information Center

    Nunley, Kathie F.

    1996-01-01

    Presents an activity to simulate electrophoresis using everyday items. Uses adding machine paper to construct a set of DNA fingerprints that can be used to solve crime cases designed by students in any biology class. (JRH)

  19. Harnessing DNA intercalation.

    PubMed

    Persil, Ozgül; Hud, Nicholas V

    2007-10-01

    Numerous small molecules are known to bind to DNA through base pair intercalation. Fluorescent dyes commonly used for nucleic acid staining, such as ethidium, are familiar examples. Biological and physical studies of DNA intercalation have historically been motivated by mutation and drug discovery research. However, this same mode of binding is now being harnessed for the creation of novel molecular assemblies. Recent studies have used DNA scaffolds and intercalators to construct supramolecular assemblies that function as fluorescent 'nanotags' for cell labeling. Other studies have demonstrated how intercalators can be used to promote the formation of otherwise unstable nucleic acid assemblies. These applications illustrate how intercalators can be used to facilitate and expand DNA-based nanotechnology. PMID:17825446

  20. DNA damage tolerance.

    PubMed

    Branzei, Dana; Psakhye, Ivan

    2016-06-01

    Accurate chromosomal DNA replication is fundamental for optimal cellular function and genome integrity. Replication perturbations activate DNA damage tolerance pathways, which are crucial to complete genome duplication as well as to prevent formation of deleterious double strand breaks. Cells use two general strategies to tolerate lesions: recombination to a homologous template, and trans-lesion synthesis with specialized polymerases. While key players of these processes have been outlined, much less is known on their choreography and regulation. Recent advances have uncovered principles by which DNA damage tolerance is regulated locally and temporally - in relation to replication timing and cell cycle stage -, and are beginning to elucidate the DNA dynamics that mediate lesion tolerance and influence chromosome structure during replication. PMID:27060551

  1. Multiplex analysis of DNA

    DOEpatents

    Church, George M.; Kieffer-Higgins, Stephen

    1992-01-01

    This invention features vectors and a method for sequencing DNA. The method includes the steps of: a) ligating the DNA into a vector comprising a tag sequence, the tag sequence includes at least 15 bases, wherein the tag sequence will not hybridize to the DNA under stringent hybridization conditions and is unique in the vector, to form a hybrid vector, b) treating the hybrid vector in a plurality of vessels to produce fragments comprising the tag sequence, wherein the fragments differ in length and terminate at a fixed known base or bases, wherein the fixed known base or bases differs in each vessel, c) separating the fragments from each vessel according to their size, d) hybridizing the fragments with an oligonucleotide able to hybridize specifically with the tag sequence, and e) detecting the pattern of hybridization of the tag sequence, wherein the pattern reflects the nucleotide sequence of the DNA.

  2. A compendium of human mitochondrial DNA control region: development of an international standard forensic database.

    PubMed

    Miller, K W; Budowle, B

    2001-06-01

    A compendium of human mitochondrial DNA (mtDNA) control region types has been constructed. This updated compilation indexes over 10,000 population-specific mtDNA nucleotide sequences in a standardized format. The sequences represent mtDNA types from the Scientific Working Group on DNA Analysis Methods (SWGDAM) mtDNA database and from the public literature. The SWGDAM data are considered to be of higher quality than the public data, particularly for counting the number of times a particular haplotype has been observed. PMID:11387646

  3. Das DNA-Puzzle

    NASA Astrophysics Data System (ADS)

    Kirchner, Stefan

    Im Jahre 1953 wurde von James Watson und Francis Crick erstmalig der strukturelle Aufbau der sogenannten DNA (Desoxyribonukleinsäure) beschrieben, welche das Erbgut jedes Lebewesens enthält. Der wesentliche Teil des Erbguts wird dabei durch eine sehr lange Folge der vier Basen Adenin (A), Cytosin (C), Guanin (G) und Thymin (T) codiert. Seit einigen Jahren ist es möglich, die Folge der vier Basen zu einer gegebenen DNA zu bestimmen. Biologen bezeichnen diesen Vorgang als Sequenzierung.

  4. DNA-Nanoparticle Tinkertoys.

    PubMed

    Chandrasekaran, Arun Richard

    2016-06-16

    Nanoparticle superlattices can be self-assembled by using DNA linkers, which gives control over their size, shape, and composition. Recently, such programmable atom equivalents have been tailored to respond to chemical stimuli and result in specific crystalline lattices. Moreover, the molecular recognition properties and the robustness of designed DNA nanostructures have been used in combination with metallic nanoparticles for the production of the elusive diamond superlattice. PMID:27080095

  5. Redefining Cohesiveness in Groups.

    ERIC Educational Resources Information Center

    Keyton, Joann; Springston, Jeff

    1990-01-01

    Attempted to replicate and extend research on work of Kelly and Duran in assessing relationship of group member perceptions of group interaction to group effectiveness. Concludes perceived similarity may not always align with perceptions of cohesiveness. (Author/ABL)

  6. Ribonucleotide triggered DNA damage and RNA-DNA damage responses

    PubMed Central

    Wallace, Bret D; Williams, R Scott

    2014-01-01

    Research indicates that the transient contamination of DNA with ribonucleotides exceeds all other known types of DNA damage combined. The consequences of ribose incorporation into DNA, and the identity of protein factors operating in this RNA-DNA realm to protect genomic integrity from RNA-triggered events are emerging. Left unrepaired, the presence of ribonucleotides in genomic DNA impacts cellular proliferation and is associated with chromosome instability, gross chromosomal rearrangements, mutagenesis, and production of previously unrecognized forms of ribonucleotide-triggered DNA damage. Here, we highlight recent findings on the nature and structure of DNA damage arising from ribonucleotides in DNA, and the identification of cellular factors acting in an RNA-DNA damage response (RDDR) to counter RNA-triggered DNA damage. PMID:25692233

  7. Probing Protein-DNA Interactions by Unzipping DNA

    NASA Astrophysics Data System (ADS)

    Wang, Michelle

    2003-03-01

    Protein-DNA interactions are essential to cellular processes. In replication, transcription, recombination, DNA repair, and DNA packaging, proteins bind to DNA as activators or repressors, to recruit other proteins, or to carry out various catalytic activities. I will present Unzipping Force Analysis of Protein Association (UFAPA) as a novel and versatile method for detection of the position and dynamic nature of protein-DNA interactions. A single DNA double helix was unzipped in the presence of DNA-binding proteins using a feedback-enhanced optical trap. When the unzipping fork in a DNA reached a bound protein molecule, we observed a dramatic increase in the tension in the DNA, followed by a sudden tension reduction. Analysis of the unzipping force throughout an unbinding "event" revealed information about the spatial location and dynamic nature of the protein-DNA complex.

  8. Multilocus sequence analysis supports the taxonomic position of Astragalus glycyphyllos symbionts based on DNA-DNA hybridization.

    PubMed

    Gnat, Sebastian; Małek, Wanda; Oleńska, Ewa; Wdowiak-Wróbel, Sylwia; Kalita, Michał; Rogalski, Jerzy; Wójcik, Magdalena

    2016-04-01

    In this study, the phylogenetic relationship and taxonomic status of six strains, representing different phenons and genomic groups of Astragalus glycyphyllos symbionts, originating from Poland, were established by comparative analysis of five concatenated housekeeping gene sequences (atpD, dnaK, glnA, recA and rpoB), DNA-DNA hybridization and total DNA G+C content. Maximum-likelihood phylogenetic analysis of combined atpD, dnaK, glnA, recA and rpoB sequence data placed the studied bacteria into the clade comprising the genus Mesorhizobium. In the core gene phylograms, four A. glycyphyllos nodule isolates (AG1, AG7, AG15 and AG27) formed a cluster common with Mesorhizobium ciceri, whereas the two other A. glycyphyllos symbionts (AG17 and AG22) were grouped together with Mesorhizobium amorphae and M. septentrionale. The species position of the studied bacteria was clarified by DNA-DNA hybridization. The DNA-DNA relatedness between isolates AG1, AG7, AG15 and AG27 and reference strain M. ciceri USDA 3383T was 76.4-84.2 %, and all these A. glycyphyllos nodulators were defined as members of the genomospecies M. ciceri. DNA-DNA relatedness for isolates AG17 and AG22 and the reference strain M. amorphae ICMP 15022T was 77.5 and 80.1 %, respectively. We propose that the nodule isolates AG17 and AG22 belong to the genomic species M. amorphae. Additionally, it was found that the total DNA G+C content of the six test A. glycyphyllos symbionts was 59.4-62.1 mol%, within the range for species of the genus Mesorhizobium. PMID:26704062

  9. DNA biosensors that reason.

    PubMed

    Sainz de Murieta, Iñaki; Rodríguez-Patón, Alfonso

    2012-08-01

    Despite the many designs of devices operating with the DNA strand displacement, surprisingly none is explicitly devoted to the implementation of logical deductions. The present article introduces a new model of biosensor device that uses nucleic acid strands to encode simple rules such as "IF DNA_strand(1) is present THEN disease(A)" or "IF DNA_strand(1) AND DNA_strand(2) are present THEN disease(B)". Taking advantage of the strand displacement operation, our model makes these simple rules interact with input signals (either DNA or any type of RNA) to generate an output signal (in the form of nucleotide strands). This output signal represents a diagnosis, which either can be measured using FRET techniques, cascaded as the input of another logical deduction with different rules, or even be a drug that is administered in response to a set of symptoms. The encoding introduces an implicit error cancellation mechanism, which increases the system scalability enabling longer inference cascades with a bounded and controllable signal-noise relation. It also allows the same rule to be used in forward inference or backward inference, providing the option of validly outputting negated propositions (e.g. "diagnosis A excluded"). The models presented in this paper can be used to implement smart logical DNA devices that perform genetic diagnosis in vitro. PMID:22406690

  10. Ancient dirt DNA

    NASA Astrophysics Data System (ADS)

    Willerslev, E.

    2007-12-01

    In the past two decades, ancient DNA research has progressed from the retrieval of small fragments of mitochondrial DNA from a few late Holocene specimens, to large-scale studies of ancient populations, phenotypically important nuclear loci, and even whole genomic studies of extinct species. However, the field is still regularly marred by erroneous reports, which underestimate the extent of contamination within laboratories and samples themselves. An improved understanding of these processes and the effects of damage on ancient DNA templates has started to provide a more robust basis for research. Recent methodological advances have included the discoveries of DNA preserved in ancient sediments, coprolites, and fossil ice (Ancient Dirt DNA). These findings promise to make possible the reconstructions of entire ecosystems through time and allow for studies of past population genetics in cases where fossils are rare. The advantages and pitfalls connected to the Ancient Dirt DNA approach will be discussed as will recently obtained data relating to Greenland environmental history, long-term bacterial survival and the first human migration into the Americas.

  11. Variations in brain DNA

    PubMed Central

    Avila, Jesús; Gómez-Ramos, Alberto; Soriano, Eduardo

    2014-01-01

    It is assumed that DNA sequences are conserved in the diverse cell types present in a multicellular organism like the human being. Thus, in order to compare the sequences in the genome of DNA from different individuals, nucleic acid is commonly isolated from a single tissue. In this regard, blood cells are widely used for this purpose because of their availability. Thus blood DNA has been used to study genetic familiar diseases that affect other tissues and organs, such as the liver, heart, and brain. While this approach is valid for the identification of familial diseases in which mutations are present in parental germinal cells and, therefore, in all the cells of a given organism, it is not suitable to identify sporadic diseases in which mutations might occur in specific somatic cells. This review addresses somatic DNA variations in different tissues or cells (mainly in the brain) of single individuals and discusses whether the dogma of DNA invariance between cell types is indeed correct. We will also discuss how single nucleotide somatic variations arise, focusing on the presence of specific DNA mutations in the brain. PMID:25505410

  12. Vibrational structure of DNA

    NASA Astrophysics Data System (ADS)

    Gómez C, S.; Rey-González, R. R.

    2003-10-01

    DNA has been object of more extensive research in last years. Human genome may be the main work. On the other hand, DNA has been used as physical system in opposite to its biological character. Examples of this are electronic, thermally and Ramman spectroscopy studies, in others. However, some DNA physical features are unclear, they deserve more work in a effort to understand them. In this work we are interesting on the vibrational properties of DNA. We model it as a lineal chain constituted by three different mass. We use two different constant forces into the Dynamical Matrix formalism. Two masses represent the real mass of DNA bases plus the glucose mass and the third represents the phosphate mass. In this model, DNA unit cell is composed by four masses The dispersion relation shows one acoustical and three optical branches. Also, there is a wide gap between the first and second optical branches. These features are confirmed by the density of states. Also we consider disorder effects in the proposal to do a more realistic model. In this case our results suggest a behavior as diatomic chain where the central and wide gap is preserved.

  13. Raman spectroscopy of topotecan, an inhibitor of DNA topoisomerase I

    NASA Astrophysics Data System (ADS)

    Mochalov, K. E.; Ustinova, O. A.; Strel'Tsov, S. A.; Grokhovskii, S. L.; Zhuze, A. L.; Nabiev, I. R.; Sukhanova, A. V.; Oleinikov, V. A.

    2002-10-01

    Topotecan (TPT), a water-soluble derivative of camptothecin (inhibitor of human DNA topoiomerase I), has found wide application in cancer chemotherapy. The central problem in using topotecan is the presence of lactone rings in its molecules, which undergo hydrolysis at a physiological pH yielding an inactive and even toxic form of the drug. The analysis of Raman spectra of TPT in H2O and D2O solutions made it possible to assign the spectral bands to the vibrations of particular molecular groups. Spectral features indicative of the opening of the lactone rings of the TPT molecules, deprotonation of the hydroxyl groups in their quinoline fragments, and of possible participation of the hydroxyl and carbonyl groups in H bonding are found. The data obtained are necessary to study the molecular mechanisms of TPT-DNA interaction and the formation of ternary complexes between TPT, DNA, and DNA topoisomerase I.

  14. Self-methylation of BspRI DNA-methyltransferase.

    PubMed Central

    Szilák, L; Finta, C; Patthy, A; Venetianer, P; Kiss, A

    1994-01-01

    The DNA (cytosine-5)-methyltransferase (m5C-MTase) M.BspRI is able to accept the methyl group from the methyl donor S-adenosyl-L-methionine (AdoMet) in the absence of DNA. Transfer of the methyl group to the enzyme is a slow reaction relative to DNA methylation. Self-methylation is dependent on the native conformation of the enzyme and is inhibited by S-adenosyl-L-homocysteine, DNA and sulfhydryl reagents. Amino acid sequencing of proteolytic peptides obtained from M.BspRI, which had been methylated with [methyl-3H]AdoMet, and thin layer chromatography of the modified amino acid identified two cysteines, Cys156 and Cys181 that bind the methyl group in form of S-methylcysteine. One of the acceptor residues, Cys156 is the highly conserved cysteine which plays the role of the catalytic nucleophile of m5C-MTases. Images PMID:8065896

  15. Preparation of covalently linked DNA-RNA hybrids and arabinocytidine containing DNA fragments.

    PubMed Central

    de Vroom, E; Roelen, H C; Saris, C P; Budding, T N; van der Marel, G A; van Boom, J H

    1988-01-01

    It will be demonstrated that 5'-O-DMT-N-acyl-deoxyribonucleosides, 5'-O-Lev-2'-O-MTHP-N-acyl-ribonucleosides and, also, 2'-O-MTHP-N-acyl-ara-cytidine can be coupled, via the hydroxybenzotriazole phosphotriester approach, to afford two types of DNA-RNA hybrids as well as ara-C containing DNA-fragments. The final removal of acid-labile DMT and MTHP groups could be effected by 1 h treatment with 80% acetic acid of the otherwise unprotected DNA-RNA hybrids. The same acidic hydrolysis did not result in complete removal of the 2'-O-MTHP group from the ara-C unit. Complete deblocking was accomplished after an additional 2 h aqueous HC1 (0.01 M; pH 2.00) treatment. PMID:2453027

  16. Group normalization for genomic data.

    PubMed

    Ghandi, Mahmoud; Beer, Michael A

    2012-01-01

    Data normalization is a crucial preliminary step in analyzing genomic datasets. The goal of normalization is to remove global variation to make readings across different experiments comparable. In addition, most genomic loci have non-uniform sensitivity to any given assay because of variation in local sequence properties. In microarray experiments, this non-uniform sensitivity is due to different DNA hybridization and cross-hybridization efficiencies, known as the probe effect. In this paper we introduce a new scheme, called Group Normalization (GN), to remove both global and local biases in one integrated step, whereby we determine the normalized probe signal by finding a set of reference probes with similar responses. Compared to conventional normalization methods such as Quantile normalization and physically motivated probe effect models, our proposed method is general in the sense that it does not require the assumption that the underlying signal distribution be identical for the treatment and control, and is flexible enough to correct for nonlinear and higher order probe effects. The Group Normalization algorithm is computationally efficient and easy to implement. We also describe a variant of the Group Normalization algorithm, called Cross Normalization, which efficiently amplifies biologically relevant differences between any two genomic datasets. PMID:22912661

  17. Influence of Mg 2+ and Cd 2+ on the interaction between sparfloxacin and calf thymus DNA

    NASA Astrophysics Data System (ADS)

    Yuan, Xiao-Ying; Guo, Dong-Sheng; Wang, Lin-Li

    2008-04-01

    Mg 2+ and Cd 2+ have different binding capacity to sparfloxacin, and have different combination modes with calf thymus DNA. Selecting these two different metal ions, the influence of them on the binding constants between SPFX and calf thymus DNA, as well as the related mechanism have been studied by using absorption and fluorescence spectroscopy. The result shows that Cd 2+ has weak binding capacity to SPFX in the SPFX-Cd 2+ binary system, but can decrease the binding between SPFX and DNA obviously in SPFX-DNA-Cd 2+ ternary system. Mg 2+ has strong binding capacity to SPFX. It can increase the binding between SPFX and DNA at concentrations <0.01 mM, and decrease the binding between them at concentrations >0.01 mM. Referring to the different modes of Mg 2+ and Cd 2+ binding to DNA, the mechanism of the influence of metal ions on the binding between SPFX and DNA has been proposed. SPFX can directly bind to DNA by chelating DNA base sites. If a metal ion at certain concentration mainly binds to DNA bases, it can decrease the binding constants between SPFX and DNA through competing with SPFX. While if a metal ion at certain concentration mainly binds to phosphate groups of DNA, it can increase the binding constants by building a bridge between SPFX and DNA. The influence direction of metal ions on the binding between quinolone and DNA relays on their binding ratio of affinity for bases to phosphate groups on DNA. Our result supports Palumbo's conclusion that the binding between SPFX and the phosphate groups is the precondition for the combination between SPFX and DNA, which is stabilized through stacking interactions between the condensed rings of SPFX and DNA bases.

  18. Supramolecular Complexes of DNA

    NASA Astrophysics Data System (ADS)

    Zuber, G.; Scherman, D.

    Deoxyribose nucleic acid or DNA is a linear polymer in the form of a double strand, synthesised by sequential polymerisation of a large number of units chosen from among the nucleic bases called purines (adenosine A and guanosine G) and pyrimidines (cytosine C and thymidine T). DNA contains all the genetic information required for life. It exists in the form of a limited number (a few dozen) of very big molecules, called chromosomes. This genetic information is first of all transcribed. In this process, a restricted fragment of the DNA called a gene is copied in the form of ribonucleic acid, or RNA. This RNA is itself a polymer, but with a single strand in which the sequence of nucleic acids is schematically analogous to the sequence on one of the two strands of the transcribed DNA. Finally, this RNA is translated into a protein, yet another linear polymer. The proteins make up the main part of the active constituents ensuring the survival of the cell. Any loss of information, either by mutation or by deletion of the DNA, will cause an imbalance in the cell's metabolism that may in turn lead to incurable pathologies. Several strategies have been developed to reduce the consequences of such genetic deficiencies or, more generally, to act, by amplifying or suppressing them, on the mechanisms leading from the reading of the genetic information to the production of proteins: Strategies aiming to introduce synthetic DNA or RNA, which selectively block the expression of certain genes, are now being studied by an increasing number of research scientists and pharmacologists. They use antisense oligodeoxyribonucleotides or interfering oligoribonucleotides and they already have clinical applications. This kind of therapy is often called gene pharmacology. Other, more ambitious strategies aim to repair in situ mutated or incomplete DNA within the chromosomes themselves, by introducing short sequences of DNA or RNA which recognise and take the place of mutations. This is the

  19. DNA templates silver clusters with magic sizes and colors for multi-cluster fluorescent assemblies

    NASA Astrophysics Data System (ADS)

    Copp, Stacy

    2015-03-01

    The natural inclusion of information in DNA, a vital part of life's rich complexity, can also be exploited to create diverse structures with multiple scales of complexity. Now emerging in novel photonic applications, DNA-stabilized silver clusters (AgN-DNA) are compelling examples of multi-scale DNA-directed assembly: individual fluorescent clusters, each templated by specific DNA base motifs, can then be arranged together in DNA-mediated multi-cluster assemblies with nanoscale precision. We discuss how DNA imbues AgN-DNA with unique features. Our optical data on pure AgN-DNA show that DNA base-cationic silver ligands impose rod-like shapes for neutral silver clusters, whose length primarily determines fluorescence color. This shape anisotropy leads to the aspherical AgN-DNA magic number cluster sizes and ``magic color'' groupings. We exploit DNA's sequence properties to extract multi-base motifs that select certain magic cluster sizes, using machine learning algorithms applied to large data sets. With these base motifs, we design DNA scaffolds to arrange multiple atomically precise AgN together in nanoscale proximity. We demonstrate that clusters are stable when held at separations below 10 nm, both in bicolor, dual cluster DNA clamp assemblies and in one-dimensional assemblies of atomically precise clusters arrayed on DNA nanotubes. Supported by NSF-CHE-1213895 and NSF-DMR-1309410. SMC acknowledges NSF-DGE-1144085, a NSF GRFP.

  20. DNA Knots: Theory and Experiments

    NASA Astrophysics Data System (ADS)

    Sumners, D. W.

    Cellular DNA is a long, thread-like molecule with remarkably complex topology. Enzymes that manipulate the geometry and topology of cellular DNA perform many vital cellular processes (including segregation of daughter chromosomes, gene regulation, DNA repair, and generation of antibody diversity). Some enzymes pass DNA through itself via enzyme-bridged transient breaks in the DNA; other enzymes break the DNA apart and reconnect it to different ends. In the topological approach to enzymology, circular DNA is incubated with an enzyme, producing an enzyme signature in the form of DNA knots and links. By observing the changes in DNA geometry (supercoiling) and topology (knotting and linking) due to enzyme action, the enzyme binding and mechanism can often be characterized. This paper will discuss some personal research history, and the tangle model for the analysis of site-specific recombination experiments on circular DNA.

  1. Genetic Organization of the Vibrio harveyi dnaA Gene Region and Analysis of the Function of the V. harveyi DnaA Protein in Escherichia coli

    PubMed Central

    Berenstein, Dvora; Olesen, Kirsten; Speck, Christian; Skovgaard, Ole

    2002-01-01

    The Vibrionaceae family is distantly related to Enterobacteriaceae within the group of bacteria possessing the Dam methylase system. We have cloned, sequenced, and analyzed the dnaA gene region of Vibrio harveyi and found that although the organization of the V. harveyi dnaA region differs from that of Escherichia coli, the expression of both genes is autoregulated and ATP-DnaA binds cooperatively to ATP-DnaA boxes in the dnaA promoter region. The DnaA proteins of V. harveyi and E. coli are interchangeable and function nearly identically in controlling dnaA transcription and the initiation of chromosomal DNA replication despite the evolutionary distance between these bacteria. PMID:11948168

  2. Energy and Technology Review: Unlocking the mysteries of DNA repair

    SciTech Connect

    Quirk, W.A.

    1993-04-01

    DNA, the genetic blueprint, has the remarkable property of encoding its own repair following diverse types of structural damage induced by external agents or normal metabolism. We are studying the interplay of DNA damaging agents, repair genes, and their protein products to decipher the complex biochemical pathways that mediate such repair. Our research focuses on repair processes that correct DNA damage produced by chemical mutagens and radiation, both ionizing and ultraviolet. The most important type of DNA repair in human cells is called excision repair. This multistep process removes damaged or inappropriate pieces of DNA -- often as a string of 29 nucleotides containing the damage -- and replaces them with intact ones. We have isolated, cloned, and mapped several human repair genes associated with the nucleotide excision repair pathway and involved in the repair of DNA damage after exposure to ultraviolet light or mutagens in cooked food. We have shown that a defect in one of these repair genes, ERCC2, is responsible for the repair deficiency in one of the groups of patients with the recessive genetic disorder xeroderma pigmentosum (XP group D). We are exploring ways to purify sufficient quantities (milligrams) of the protein products of these and other repair genes so that we can understand their functions. Our long-term goals are to link defective repair proteins to human DNA repair disorders that predispose to cancer, and to produce DNA-repair-deficient mice that can serve as models for the human disorders.

  3. Molecular dynamics simulations of DNA-polycation complexes

    NASA Astrophysics Data System (ADS)

    Ziebarth, Jesse; Wang, Yongmei

    2008-03-01

    A necessary step in the preparation of DNA for use in gene therapy is the packaging of DNA with a vector that can condense DNA and provide protection from degrading enzymes. Because of the immunoresponses caused by viral vectors, there has been interest in developing synthetic gene therapy vectors, with polycations emerging as promising candidates. Molecular dynamics simulations of the DNA duplex CGCGAATTCGCG in the presence of 20 monomer long sequences of the polycations, poly-L-lysine (PLL) and polyethyleneimine (PEI), with explicit counterions and TIP3P water, are performed to provide insight into the structure and formation of DNA polyplexes. After an initial separation of approximately 50 å, the DNA and polycation come together and form a stable complex within 10 ns. The DNA does not undergo any major structural changes upon complexation and remains in the B-form. In the formed complex, the charged amine groups of the polycation mainly interact with DNA phosphate groups, and rarely occupy electronegative sites in either the major or minor grooves. Differences between complexation with PEI and PLL will be discussed.

  4. Single Nucleotide Polymorphism Analysis of European Archaeological M. leprae DNA

    PubMed Central

    Watson, Claire L.; Lockwood, Diana N. J.

    2009-01-01

    Background Leprosy was common in Europe eight to twelve centuries ago but molecular confirmation of this has been lacking. We have extracted M. leprae ancient DNA (aDNA) from medieval bones and single nucleotide polymorphism (SNP) typed the DNA, this provides insight into the pattern of leprosy transmission in Europe and may assist in the understanding of M. leprae evolution. Methods and Findings Skeletons have been exhumed from 3 European countries (the United Kingdom, Denmark and Croatia) and are dated around the medieval period (476 to 1350 A.D.). we tested for the presence of 3 previously identified single nucleotide polymorphisms (SNPs) in 10 aDNA extractions. M. leprae aDNA was extracted from 6 of the 10 bone samples. SNP analysis of these 6 extractions were compared to previously analysed European SNP data using the same PCR assays and were found to be the same. Testing for the presence of SNPs in M. leprae DNA extracted from ancient bone samples is a novel approach to analysing European M. leprae DNA and the findings concur with the previously published data that European M. leprae strains fall in to one group (SNP group 3). Conclusions These findings support the suggestion that the M. leprae genome is extremely stable and show that archaeological M. leprae DNA can be analysed to gain detailed information about the genotypic make-up of European leprosy, which may assist in the understanding of leprosy transmission worldwide. PMID:19847306

  5. Forensic DNA Profiling and Database

    PubMed Central

    Panneerchelvam, S.; Norazmi, M.N.

    2003-01-01

    The incredible power of DNA technology as an identification tool had brought a tremendous change in crimnal justice . DNA data base is an information resource for the forensic DNA typing community with details on commonly used short tandem repeat (STR) DNA markers. This article discusses the essential steps in compilation of COmbined DNA Index System (CODIS) on validated polymerase chain amplified STRs and their use in crime detection. PMID:23386793

  6. Interdependence and Group Effectiveness.

    ERIC Educational Resources Information Center

    Wageman, Ruth

    1995-01-01

    Investigated the differential effects of task design and reward system design on group functioning in a large U.S. corporation; the effectiveness of "hybrid" groups (having tasks and rewards with both individual and group elements); and how individuals' autonomy preferences moderate their responses to interdependence. Groups performed best when…

  7. Assertive Training in Groups

    ERIC Educational Resources Information Center

    Sansbury, David L.

    1974-01-01

    This article describes a group approach to helping the nonassertive client. After describing the group composition and goals, he presents a session by session description for conducting the assertive training group. In addition, he presents suggestions based on experiences in leading the group. (Author)

  8. Circadian Modulation of 8-Oxoguanine DNA Damage Repair

    PubMed Central

    Manzella, Nicola; Bracci, Massimo; Strafella, Elisabetta; Staffolani, Sara; Ciarapica, Veronica; Copertaro, Alfredo; Rapisarda, Venerando; Ledda, Caterina; Amati, Monica; Valentino, Matteo; Tomasetti, Marco; Stevens, Richard G.; Santarelli, Lory

    2015-01-01

    The DNA base excision repair pathway is the main system involved in the removal of oxidative damage to DNA such as 8-Oxoguanine (8-oxoG) primarily via the 8-Oxoguanine DNA glycosylase (OGG1). Our goal was to investigate whether the repair of 8-oxoG DNA damage follow a circadian rhythm. In a group of 15 healthy volunteers, we found a daily variation of Ogg1 expression and activity with higher levels in the morning compared to the evening hours. Consistent with this, we also found lower levels of 8-oxoG in morning hours compared to those in the evening hours. Lymphocytes exposed to oxidative damage to DNA at 8:00 AM display lower accumulation of 8-oxoG than lymphocytes exposed at 8:00 PM. Furthermore, altered levels of Ogg1 expression were also observed in a group of shift workers experiencing a deregulation of circadian clock genes compared to a control group. Moreover, BMAL1 knockdown fibroblasts with a deregulated molecular clock showed an abolishment of circadian variation of Ogg1 expression and an increase of OGG1 activity. Our results suggest that the circadian modulation of 8-oxoG DNA damage repair, according to a variation of Ogg1 expression, could render humans less susceptible to accumulate 8-oxoG DNA damage in the morning hours. PMID:26337123

  9. Circadian Modulation of 8-Oxoguanine DNA Damage Repair.

    PubMed

    Manzella, Nicola; Bracci, Massimo; Strafella, Elisabetta; Staffolani, Sara; Ciarapica, Veronica; Copertaro, Alfredo; Rapisarda, Venerando; Ledda, Caterina; Amati, Monica; Valentino, Matteo; Tomasetti, Marco; Stevens, Richard G; Santarelli, Lory

    2015-01-01

    The DNA base excision repair pathway is the main system involved in the removal of oxidative damage to DNA such as 8-Oxoguanine (8-oxoG) primarily via the 8-Oxoguanine DNA glycosylase (OGG1). Our goal was to investigate whether the repair of 8-oxoG DNA damage follow a circadian rhythm. In a group of 15 healthy volunteers, we found a daily variation of Ogg1 expression and activity with higher levels in the morning compared to the evening hours. Consistent with this, we also found lower levels of 8-oxoG in morning hours compared to those in the evening hours. Lymphocytes exposed to oxidative damage to DNA at 8:00 AM display lower accumulation of 8-oxoG than lymphocytes exposed at 8:00 PM. Furthermore, altered levels of Ogg1 expression were also observed in a group of shift workers experiencing a deregulation of circadian clock genes compared to a control group. Moreover, BMAL1 knockdown fibroblasts with a deregulated molecular clock showed an abolishment of circadian variation of Ogg1 expression and an increase of OGG1 activity. Our results suggest that the circadian modulation of 8-oxoG DNA damage repair, according to a variation of Ogg1 expression, could render humans less susceptible to accumulate 8-oxoG DNA damage in the morning hours. PMID:26337123

  10. Human DNA polymerase α in binary complex with a DNA:DNA template-primer

    PubMed Central

    Coloma, Javier; Johnson, Robert E.; Prakash, Louise; Prakash, Satya; Aggarwal, Aneel K.

    2016-01-01

    The Polα/primase complex assembles the short RNA-DNA fragments for priming of lagging and leading strand DNA replication in eukaryotes. As such, the Polα polymerase subunit encounters two types of substrates during primer synthesis: an RNA:DNA helix and a DNA:DNA helix. The engagement of the polymerase subunit with the DNA:DNA helix has been suggested as the of basis for primer termination in eukaryotes. However, there is no structural information on how the Polα polymerase subunit actually engages with a DNA:DNA helix during primer synthesis. We present here the first crystal structure of human Polα polymerase subunit in complex with a DNA:DNA helix. Unexpectedly, we find that portion of the DNA:DNA helix in contact with the polymerase is not in a B-form but in a hybrid A-B form. Almost all of the contacts observed previously with an RNA primer are preserved with a DNA primer – with the same set of polymerase residues tracking the sugar-phosphate backbone of the DNA or RNA primer. Thus, rather than loss of specific contacts, the free energy cost of distorting DNA from B- to hybrid A-B form may augur the termination of primer synthesis in eukaryotes. PMID:27032819

  11. An Electrochemical DNA Microbiosensor Based on Succinimide-Modified Acrylic Microspheres

    PubMed Central

    Ulianas, Alizar; Heng, Lee Yook; Hanifah, Sharina Abu; Ling, Tan Ling

    2012-01-01

    An electrochemical microbiosensor for DNA has been fabricated based on new acrylic microspheres modified with reactive N-acryloxysuccinimide (NAS) functional groups. Hydrophobic poly(n-butylacrylate-N-acryloxysuccinimide) microspheres were synthesized in an emulsion form with a simple one-step photopolymerization technique. Aminated DNA probe was attached to the succinimde functional group of the acrylic microspheres via covalent bonding. The hybridization of the immobilized DNA probe with the complementary DNA was studied by differential pulse voltametry using anthraquninone-2-sulfonic acid monohydrate sodium salt (AQMS) as the electroactive hybridization label. The influences of many factors such as duration of DNA probe immobilization and hybridization, pH, type of ions, buffer concentrations, ionic strength, operational temperature and non-complementary DNA on the biosensor performance were evaluated. Under optimized conditions, the DNA microbiosensor demonstrated a linear response range to target DNA over a wide concentration range of 1.0 × 10−16 and 1.0 × 10−8 M with a lower limit of detection (LOD) of 9.46 × 10−17 M (R2 = 0.97). This DNA microbiosensor showed good reproducibility with 2.84% RSD (relative standard deviation) (n = 3). Application of the NAS-modified acrylic microspheres in the construction of DNA microbiosensor had improved the overall analytical performance of the resultant DNA microbiosensor when compared with other reported DNA biosensors using other nano-materials for membranes and microspheres as DNA immobilization matrices. PMID:22778594

  12. Dynamic Modulation of DNA Hybridization Using Allosteric DNA Tetrahedral Nanostructures.

    PubMed

    Song, Ping; Li, Min; Shen, Juwen; Pei, Hao; Chao, Jie; Su, Shao; Aldalbahi, Ali; Wang, Lihua; Shi, Jiye; Song, Shiping; Wang, Lianhui; Fan, Chunhai; Zuo, Xiaolei

    2016-08-16

    The fixed dynamic range of traditional biosensors limits their utility in several real applications. For example, viral load monitoring requires the dynamic range spans several orders of magnitude; whereas, monitoring of drugs requires extremely narrow dynamic range. To overcome this limitation, here, we devised tunable biosensing interface using allosteric DNA tetrahedral bioprobes to tune the dynamic range of DNA biosensors. Our strategy takes the advantage of the readily and flexible structure design and predictable geometric reconfiguration of DNA nanotechnology. We reconfigured the DNA tetrahedral bioprobes by inserting the effector sequence into the DNA tetrahedron, through which, the binding affinity of DNA tetrahedral bioprobes can be tuned. As a result, the detection limit of DNA biosensors can be programmably regulated. The dynamic range of DNA biosensors can be tuned (narrowed or extended) for up to 100-fold. Using the regulation of binding affinity, we realized the capture and release of biomolecules by tuning the binding behavior of DNA tetrahedral bioprobes. PMID:27435955

  13. Strandwise translocation of a DNA glycosylase on undamaged DNA

    SciTech Connect

    Qi, Yan; Nam, Kwangho; Spong, Marie C.; Banerjee, Anirban; Sung, Rou-Jia; Zhang, Michael; Karplus, Martin; Verdine, Gregory L.

    2012-05-14

    Base excision repair of genotoxic nucleobase lesions in the genome is critically dependent upon the ability of DNA glycosylases to locate rare sites of damage embedded in a vast excess of undamaged DNA, using only thermal energy to fuel the search process. Considerable interest surrounds the question of how DNA glycosylases translocate efficiently along DNA while maintaining their vigilance for target damaged sites. Here, we report the observation of strandwise translocation of 8-oxoguanine DNA glycosylase, MutM, along undamaged DNA. In these complexes, the protein is observed to translocate by one nucleotide on one strand while remaining untranslocated on the complementary strand. We further report that alterations of single base-pairs or a single amino acid substitution (R112A) can induce strandwise translocation. Molecular dynamics simulations confirm that MutM can translocate along DNA in a strandwise fashion. These observations reveal a previously unobserved mode of movement for a DNA-binding protein along the surface of DNA.

  14. Active DNA demethylation by DNA repair: Facts and uncertainties.

    PubMed

    Schuermann, David; Weber, Alain R; Schär, Primo

    2016-08-01

    Pathways that control and modulate DNA methylation patterning in mammalian cells were poorly understood for a long time, although their importance in establishing and maintaining cell type-specific gene expression was well recognized. The discovery of proteins capable of converting 5-methylcytosine (5mC) to putative substrates for DNA repair introduced a novel and exciting conceptual framework for the investigation and ultimate discovery of molecular mechanisms of DNA demethylation. Against the prevailing notion that DNA methylation is a static epigenetic mark, it turned out to be dynamic and distinct mechanisms appear to have evolved to effect global and locus-specific DNA demethylation. There is compelling evidence that DNA repair, in particular base excision repair, contributes significantly to the turnover of 5mC in cells. By actively demethylating DNA, DNA repair supports the developmental establishment as well as the maintenance of DNA methylation landscapes and gene expression patterns. Yet, while the biochemical pathways are relatively well-established and reviewed, the biological context, function and regulation of DNA repair-mediated active DNA demethylation remains uncertain. In this review, we will thus summarize and critically discuss the evidence that associates active DNA demethylation by DNA repair with specific functional contexts including the DNA methylation erasure in the early embryo, the control of pluripotency and cellular differentiation, the maintenance of cell identity, and the nuclear reprogramming. PMID:27247237

  15. Programmable Quantitative DNA Nanothermometers.

    PubMed

    Gareau, David; Desrosiers, Arnaud; Vallée-Bélisle, Alexis

    2016-07-13

    Developing molecules, switches, probes or nanomaterials that are able to respond to specific temperature changes should prove of utility for several applications in nanotechnology. Here, we describe bioinspired strategies to design DNA thermoswitches with programmable linear response ranges that can provide either a precise ultrasensitive response over a desired, small temperature interval (±0.05 °C) or an extended linear response over a wide temperature range (e.g., from 25 to 90 °C). Using structural modifications or inexpensive DNA stabilizers, we show that we can tune the transition midpoints of DNA thermometers from 30 to 85 °C. Using multimeric switch architectures, we are able to create ultrasensitive thermometers that display large quantitative fluorescence gains within small temperature variation (e.g., > 700% over 10 °C). Using a combination of thermoswitches of different stabilities or a mix of stabilizers of various strengths, we can create extended thermometers that respond linearly up to 50 °C in temperature range. Here, we demonstrate the reversibility, robustness, and efficiency of these programmable DNA thermometers by monitoring temperature change inside individual wells during polymerase chain reactions. We discuss the potential applications of these programmable DNA thermoswitches in various nanotechnology fields including cell imaging, nanofluidics, nanomedecine, nanoelectronics, nanomaterial, and synthetic biology. PMID:27058370

  16. Electroeluting DNA fragments.

    PubMed

    Zarzosa-Alvarez, Ana L; Sandoval-Cabrera, Antonio; Torres-Huerta, Ana L; Bermudez-Cruz, Rosa M

    2010-01-01

    Purified DNA fragments are used for different purposes in Molecular Biology and they can be prepared by several procedures. Most of them require a previous electrophoresis of the DNA fragments in order to separate the band of interest. Then, this band is excised out from an agarose or acrylamide gel and purified by using either: binding and elution from glass or silica particles, DEAE-cellulose membranes, "crush and soak method", electroelution or very often expensive commercial purification kits. Thus, selecting a method will depend mostly of what is available in the laboratory. The electroelution procedure allows one to purify very clean DNA to be used in a large number of applications (sequencing, radiolabeling, enzymatic restriction, enzymatic modification, cloning etc). This procedure consists in placing DNA band-containing agarose or acrylamide slices into sample wells of the electroeluter, then applying current will make the DNA fragment to leave the agarose and thus be trapped in a cushion salt to be recovered later by ethanol precipitation. PMID:20834225

  17. Dynamical Behavior of DNA

    NASA Astrophysics Data System (ADS)

    Kumar, Shankar

    1990-01-01

    The crystal structure of the DNA-EcoRI complex (Kim et al., 1990) revealed the existence of a 'kink' (or a disruption of the helical symmetry) in the DNA. Part of this work was an investigation of whether or not the kinked structure is a physically meaningful metastable state that is intrinsic to DNA. By using the "All Atom" hamiltonian of Weiner et al (1986) it has been found that the kink is not a metastable feature of the DNA. Rapid scanning of conformational space is indispensable in statistical mechanical studies of proteins and DNA. The Quasi-Optimized-Monte-Carlo (or QOMC) method is more efficient than the conventional Metropolis Monte Carlo method in the simulated annealing calculations reported here. It is also shown here that using altered masses in Molecular Dynamics calculations enhances sampling efficiency. The Multiple Histogram technique (Ferrenberg, 1989) has been applied for the first time on complex biomolecular hamiltonians. This method is superior to the classical perturbation and multistage sampling techniques for calculating free energy differences and generating potential of mean force profiles for suitably chosen reaction coordinates. This was demonstrated by using the multiple histogram method to generate the potential of mean force for the pseudorotation phase angle of the sugar ring in adenosine.

  18. Chromatin and DNA replication.

    PubMed

    MacAlpine, David M; Almouzni, Geneviève

    2013-08-01

    The size of a eukaryotic genome presents a unique challenge to the cell: package and organize the DNA to fit within the confines of the nucleus while at the same time ensuring sufficient dynamics to allow access to specific sequences and features such as genes and regulatory elements. This is achieved via the dynamic nucleoprotein organization of eukaryotic DNA into chromatin. The basic unit of chromatin, the nucleosome, comprises a core particle with 147 bp of DNA wrapped 1.7 times around an octamer of histones. The nucleosome is a highly versatile and modular structure, both in its composition, with the existence of various histone variants, and through the addition of a series of posttranslational modifications on the histones. This versatility allows for both short-term regulatory responses to external signaling, as well as the long-term and multigenerational definition of large functional chromosomal domains within the nucleus, such as the centromere. Chromatin organization and its dynamics participate in essentially all DNA-templated processes, including transcription, replication, recombination, and repair. Here we will focus mainly on nucleosomal organization and describe the pathways and mechanisms that contribute to assembly of this organization and the role of chromatin in regulating the DNA replication program. PMID:23751185

  19. Kinetoplast DNA minicircles: regions of extensive sequence divergence.

    PubMed Central

    Rogers, W O; Wirth, D F

    1987-01-01

    Previous work has shown that the kinetoplast minicircle DNA of Leishmania species exhibits species-specific sequence divergence and this observation has led to the development of a DNA probe-based diagnostic test for leishmaniasis. In the work reported here, we demonstrate that the minicircle is composed of three types of DNA sequences with differing specificities reflecting different rates of DNA sequence change. A library of cloned fragments of kinetoplast DNA (kDNA) from Leishmania mexicana amazonensis was prepared and the cloned subfragments were found to contain DNA sequences with different taxonomic specificities based on hybridization analysis with various species of Leishmania. Four groups of subfragments were found, those that hybridized with a large number of Leishmania sp. as well as sequences unique to the species, subspecies, or isolate. Analysis of nested deletions of a single, full-length minicircle demonstrates that these different taxonomic specificities are contained within a single minicircle. This implies that different regions of a single minicircle have DNA sequences that diverge at different rates. These sequences represent potentially valuable tools in diagnostic, epidemiologic, and ecological studies of leishmaniasis and provide the basis for a model of kDNA sequence evolution. Images PMID:3025880

  20. Structural analysis of DNA interaction with retinol and retinoic acid.

    PubMed

    Mandeville, J S; N'soukpoé-Kossi, C N; Neault, J F; Tajmir-Riahi, H A

    2010-06-01

    Dietary constituents of fresh fruits and vegetables may play a relevant role in DNA adduct formation by inhibiting enzymatic activities. Studies have shown the important role of antioxidant vitamins A, C, and E in the protection against cancer and cardiovascular diseases. The antioxidant activity of vitamin A and beta-carotene may consist of scavenging oxygen radicals and preventing DNA damage. This study was designed to examine the interaction of calf-thymus DNA with retinol and retinoic acid in aqueous solution at physiological conditions using a constant DNA concentration and various retinoid contents. Fourier transform infrared (FTIR), circular dichroism (CD), and fluorescence spectroscopic methods were used to determine retinoid binding mode, the binding constant, and the effects of retinol and retinoic acid complexation on DNA conformation and aggregation. Structural analysis showed that retinol and retinoic acid bind DNA via G-C and A-T base pairs and the backbone phosphate groups with overall binding constants of Kret = 3.0 (+/-0.50) x 10(3) (mol.L(-1))(-1) and Kretac = 1.0 (+/-0.20) x 10(4) (mol.L(-1))(-1). The number of bound retinoids per DNA were 0.84 for retinol and 1.3 for retinoic acid. Hydrophobic interactions were also observed at high retinol and retinoic acid contents. At a high retinoid concentration, major DNA aggregation occurred, while DNA remained in the B-family structure. PMID:20555389

  1. The effect of local melting of DNA on DNA loop formation

    NASA Astrophysics Data System (ADS)

    Jeong, Jiyoun; Kim, Harold

    Statistical mechanics of double-stranded DNA (dsDNA) is well described by the wormlike chain model (WLC) which assumes a harmonic bending potential. Such smooth bending potential may no longer be valid for large bending angles to form small loops (<100 bp). Instead, DNA may rely on rare structural transitions such as local melting (opening) of base pairs to lower the energetic cost. In theory, open base pairs called bubbles can increase the looping probability of short DNA molecules by a few orders of magnitude, but a robust experimental validation of this theoretical prediction is lacking. Here, we investigated the correlation between local melting probability and looping dynamics of dsDNA using single-molecule fluorescence resonance energy transfer (FRET). We designed two groups of short DNA molecules with low and high melting probabilities around their center and measured their looping and unlooping rates in equilibrium. Our data allow rigorous tests of meltable wormlike chain (MWLC) models at short length scales for setting ranges of acceptable free energy cost of bubble formation and flexibility values of a bubble.

  2. Wanderings of a DNA enzymologist: from DNA polymerase to viral latency.

    PubMed

    Lehman, I Robert

    2006-01-01

    I am a member of what has been called, perhaps too grandiosely, "The Greatest Generation." I grew up during the Great Depression and served in the U.S. Army during World War II. Because of my military service and the benefits of the GI Bill, I was able to attend college and, later, graduate school. Early in my graduate studies, I became fascinated with enzymes and the biochemical reactions that they catalyze. This fascination has never left me during the 50 years I have been a "DNA enzymologist." I was fortunate to have had as a mentor Arthur Kornberg, one of the great biochemists of the twentieth century, and a splendid group of postdocs and graduate students. I have studied DNA polymerases, DNA nucleases, DNA ligases, and DNA recombinases, enzymes that are critical to our understanding of DNA replication, repair, and recombination. Most recently, I have been studying herpes virus replication and inadvertently wandered into an entirely new area-viral latency. PMID:16756482

  3. DNA bending facilitates the error-free DNA damage tolerance pathway and upholds genome integrity

    PubMed Central

    Gonzalez-Huici, Victor; Szakal, Barnabas; Urulangodi, Madhusoodanan; Psakhye, Ivan; Castellucci, Federica; Menolfi, Demis; Rajakumara, Eerappa; Fumasoni, Marco; Bermejo, Rodrigo; Jentsch, Stefan; Branzei, Dana

    2014-01-01

    DNA replication is sensitive to damage in the template. To bypass lesions and complete replication, cells activate recombination-mediated (error-free) and translesion synthesis-mediated (error-prone) DNA damage tolerance pathways. Crucial for error-free DNA damage tolerance is template switching, which depends on the formation and resolution of damage-bypass intermediates consisting of sister chromatid junctions. Here we show that a chromatin architectural pathway involving the high mobility group box protein Hmo1 channels replication-associated lesions into the error-free DNA damage tolerance pathway mediated by Rad5 and PCNA polyubiquitylation, while preventing mutagenic bypass and toxic recombination. In the process of template switching, Hmo1 also promotes sister chromatid junction formation predominantly during replication. Its C-terminal tail, implicated in chromatin bending, facilitates the formation of catenations/hemicatenations and mediates the roles of Hmo1 in DNA damage tolerance pathway choice and sister chromatid junction formation. Together, the results suggest that replication-associated topological changes involving the molecular DNA bender, Hmo1, set the stage for dedicated repair reactions that limit errors during replication and impact on genome stability. PMID:24473148

  4. Structural heterogeneity of mitochondrial DNA molecules within the genus Drosophila.

    PubMed Central

    Fauron, C M; Wolstenholme, D R

    1976-01-01

    We have determined by electron microscopy the molecular weight of circular mitochondrial DNA (mtDNA) molecules from 39 species representing 13 groups of five subgenera of the genus Drosophila. mtDNA molecules of all species examined, other than members of the melanogaster group, had, with one exception, molecular weights in the rather narrow range 9.90 X 10(6). The one exception was D. robusta, which had a molecular weight of 10.61 X 10(6). In contrast, mtDNA molecules from species within the melanogaster group had molecular weights covering the considerably greater range 9.92 X 10(6) to 12.35 X 10(6). Applying the electron microscope denaturation mapping technique of Inman to mtDNA molecules of eight species of the melanogaster group, we found each of them to contain a region [presumably rich in adenine and thymine (A+T)] which denatured at a specific temperature (40 degrees) at which most of the remainder of the molecule remained undenatured. The size of the A+T-rich region was constant for mtDNA molecules of a species, but varied from 0.62 X 10(6) to 3.41 X 10(6) for mtDNA molecules of different species. It was demonstrated that the differences in molecular weights of the A+T-rich regions can almost completely account for the differences in total molecular weights of the mtDNA molecules from species of the melanogaster group. Images PMID:1068475

  5. Liposome-mediated DNA immunisation via the subcutaneous route.

    PubMed

    Perrie, Y; McNeil, S; Vangala, A

    2003-01-01

    liposome-mediated DNA immunisation employing two DRV(DNA) formulations as well as naked DNA revealed that humoural responses (immunoglobulin total IgG, and subclasses IgG1 and 1gG2a) engendered by the plasmid encoded NP were substantially higher after dosing twice, 28 days apart with 10 microg liposome-entrapped DNA compared to naked DNA. At all time points measured, mice immunised with naked DNA showed no greater immune response compared to the control, non-immunised group. In contrast, as early as day 49, responses were significantly higher in mice injected with DNA entrapped in DRV liposomes containing DOTAP compared to the control group and mice immunised with naked DNA. By day 56, all total IgG responses from mice immunised with both DRV formulations were significantly higher. Comparison between the DRV formulations revealed no significant difference in immune responses elicited except at day 114, where the humoural responses of the group injected with liposomal formulation containing DC-Chol dropped to significantly lower levels that those measured in mice which received the DOTAP formulation. Similar results were found when the IgG1 and IgG2a subclass responses were determined. These results suggest that, not only can DNA be effectively entrapped within liposomes using the DRV method but that such DRV liposomes containing DNA may be a useful system for subcutaneous delivery of DNA vaccines. PMID:15203925

  6. Transposon facilitated DNA sequencing

    SciTech Connect

    Berg, D.E.; Berg, C.M.; Huang, H.V.

    1990-01-01

    The purpose of this research is to investigate and develop methods that exploit the power of bacterial transposable elements for large scale DNA sequencing: Our premise is that the use of transposons to put primer binding sites randomly in target DNAs should provide access to all portions of large DNA fragments, without the inefficiencies of methods involving random subcloning and attendant repetitive sequencing, or of sequential synthesis of many oligonucleotide primers that are used to match systematically along a DNA molecule. Two unrelated bacterial transposons, Tn5 and {gamma}{delta}, are being used because they have both proven useful for molecular analyses, and because they differ sufficiently in mechanism and specificity of transposition to merit parallel development.

  7. DNA sequencing: chemical methods

    SciTech Connect

    Ambrose, B.J.B.; Pless, R.C.

    1987-01-01

    Limited base-specific or base-selective cleavage of a defined DNA fragment yields polynucleotide products, the length of which correlates with the positions of the particular base (or bases) in the original fragment. Sverdlov and co-workers recognized the possibility of using this principle for the determination of DNA sequences. In 1977 a fully elaborated method was introduced based on this principle, which allowed routine analysis of DNA sequences over distances greater than 100 nucleotide unite from a defined, radiolabeled terminus. Six procedures for partial cleavage were described. Simultaneous parallel resolution of an appropriate set of partial cleavage mixtures by polyacrylamide gel electrophoresis, followed by visualization of the radioactive bands by autoradiography, allows the deduction of nucleotide sequence.

  8. [DNA methylation in obesity].

    PubMed

    Pokrywka, Małgorzata; Kieć-Wilk, Beata; Polus, Anna; Wybrańska, Iwona

    2014-01-01

    The number of overweight and obese people is increasing at an alarming rate, especially in the developed and developing countries. Obesity is a major risk factor for diabetes, cardiovascular disease, and cancer, and in consequence for premature death. The development of obesity results from the interplay of both genetic and environmental factors, which include sedentary life style and abnormal eating habits. In the past few years a number of events accompanying obesity, affecting expression of genes which are not directly connected with the DNA base sequence (e.g. epigenetic changes), have been described. Epigenetic processes include DNA methylation, histone modifications such as acetylation, methylation, phosphorylation, ubiquitination, and sumoylation, as well as non-coding micro-RNA (miRNA) synthesis. In this review, the known changes in the profile of DNA methylation as a factor affecting obesity and its complications are described. PMID:25531701

  9. Electrochemical DNA sensor-based strategy for sensitive detection of DNA demethylation and DNA demethylase activity.

    PubMed

    Shen, Qingming; Fan, Mengxing; Yang, Yin; Zhang, Hui

    2016-08-31

    DNA demethylation and demethylase activity play important roles in DNA self-repair, and their detection is key to early diagnosis of fatal diseases. Herein, a facile electrochemical DNA (E-DNA) sensor was developed for the sensitive detection of DNA demethylation and demethylase activity based on an enzyme cleavage strategy. The thiol modified hemi-methylated hairpin probe DNA (pDNA) was self-assembled on a Au electrode surface through the formation of AuS bonds. The hemi-methylated pDNA served as the substrate of DNA demethylase (using methyl-CpG-binding domain protein 2 (MBD2) as an example). Following demethylation, the hairpin stem was then recognized and cleaved by BstUI endonuclease. The ferrocene carboxylic acid (FcA)-tagged pDNA strands were released into the buffer solution from the electrode surface, resulting in a significant decrease of electrochemical signal and providing a means to observe DNA demethylation. The activity of DNA demethylase was analyzed in the concentration ranging from 0.5 to 500 ng mL(-1) with a limit of detection as low as 0.17 ng mL(-1). With high specificity and sensitivity, rapid response, and low cost, this simple E-DNA sensor provides a unique platform for the sensitive detection of DNA demethylation, DNA demethylase activity, and related molecular diagnostics and drug screening. PMID:27506345

  10. DNA templated magnetic nanoparticles

    NASA Astrophysics Data System (ADS)

    Kinsella, Joseph M.

    Recent discoveries in nanoscience are predicted to potentially revolutionize future technologies in an extensive number of fields. These developments are contingent upon discovering new and often unconventional methods to synthesize and control nanoscale components. Nature provides several examples of working nanotechnology such as the use of programmed self assembly to build and deconstruct complex molecular systems. We have adopted a method to control the one dimensional assembly of magnetic nanoparticles using DNA as a scaffold molecule. With this method we have demonstrated the ability to organize 5 nm particles into chains that stretch up to ˜20 mum in length. One advantage of using DNA compared is the ability of the molecule to interact with other biomolecules. After assembling particles onto DNA we have been able to cleave the molecule into smaller fragments using restriction enzymes. Using ligase enzymes we have re-connected these fragments, coated with either gold or iron oxide, to form long one-dimensional arrangements of the two different types of nanoparticles on a single molecular guide. We have also created a sensitive magnetic field sensor by incorporating magnetic nanoparticle coated DNA strands with microfabricated electrodes. The IV characteristics of the aligned nanoparticles are dependant on the magnitude of an externally applied magnetic field. This transport phenomenon known as tunneling magnetoresistance (TMR) shows room temperature resistance of our devices over 80% for cobalt ferrite coated DNA when a field of 20 kOe is applied. In comparison, studies using two dimensional nanoparticle films of irox oxides xii only exhibit a 35% MR effect. Confinement into one dimension using the DNA guide produces a TMR mechanism which produces significant increases in magnetoresistance. This property can be utilized for applications in magnetic field sensing, data storage, and logic elements.

  11. Comprehensive gene expression profiling following DNA vaccination of rainbow trout against infectious hematopoietic necrosis virus

    USGS Publications Warehouse

    Purcell, Maureen K.; Nichols, Krista M.; Winton, James R.; Kurath, Gael; Thorgaard, Gary H.; Wheeler, Paul; Hansen, John D.; Herwig, Russell P.; Park, Linda K.

    2006-01-01

    The DNA vaccine based on the glycoprotein gene of Infectious hematopoietic necrosis virus induces a non-specific anti-viral immune response and long-term specific immunity against IHNV. This study characterized gene expression responses associated with the early anti-viral response. Homozygous rainbow trout were injected intra-muscularly (I.M.) with vector DNA or the IHNV DNA vaccine. Gene expression in muscle tissue (I.M. site) was evaluated using a 16,008 feature salmon cDNA microarray. Eighty different genes were significantly modulated in the vector DNA group while 910 genes were modulated in the IHNV DNA vaccinate group relative to control group. Quantitative reverse-transcriptase PCR was used to examine expression of selected immune genes at the I.M. site and in other secondary tissues. In the localized response (I.M. site), the magnitudes of gene expression changes were much greater in the vaccinate group relative to the vector DNA group for the majority of genes analyzed. At secondary systemic sites (e.g. gill, kidney and spleen), type I IFN-related genes were up-regulated in only the IHNV DNA vaccinated group. The results presented here suggest that the IHNV DNA vaccine induces up-regulation of the type I IFN system across multiple tissues, which is the functional basis of early anti-viral immunity.

  12. Automated DNA sequencing.

    PubMed

    Wallis, Yvonne; Morrell, Natalie

    2011-01-01

    Fluorescent cycle sequencing of PCR products is a multistage process and several methodologies are available to perform each stage. This chapter will describe the more commonly utilised dye-terminator cycle sequencing approach using BigDye® terminator chemistry (Applied Biosystems) ready for analysis on a 3730 DNA genetic analyzer. Even though DNA sequencing is one of the most common and robust techniques performed in molecular laboratories it may not always produce desirable results. The causes of the most common problems will also be discussed in this chapter. PMID:20938839

  13. Ancient human DNA.

    PubMed

    Kirsanow, Karola; Burger, Joachim

    2012-01-20

    The contribution of palaeogenetic data to the study of various aspects of hominin biology and evolution has been significant, and has the potential to increase substantially with the widespread implementation of next generation sequencing techniques. Here we discuss the present state-of-the-art of ancient human DNA analysis and the characteristics of hominin aDNA that make sequence validation particularly complex. A brief overview of the development of anthropological palaeogenetic analysis is given to illustrate the technical challenges motivating recent technological advancements. PMID:22169595

  14. Rigidity of melting DNA.

    PubMed

    Pal, Tanmoy; Bhattacharjee, Somendra M

    2016-05-01

    The temperature dependence of DNA flexibility is studied in the presence of stretching and unzipping forces. Two classes of models are considered. In one case the origin of elasticity is entropic due to the polymeric correlations, and in the other the double-stranded DNA is taken to have an intrinsic rigidity for bending. In both cases single strands are completely flexible. The change in the elastic constant for the flexible case due to thermally generated bubbles is obtained exactly. For the case of intrinsic rigidity, the elastic constant is found to be proportional to the square root of the bubble number fluctuation. PMID:27300825

  15. Rigidity of melting DNA

    NASA Astrophysics Data System (ADS)

    Pal, Tanmoy; Bhattacharjee, Somendra M.

    2016-05-01

    The temperature dependence of DNA flexibility is studied in the presence of stretching and unzipping forces. Two classes of models are considered. In one case the origin of elasticity is entropic due to the polymeric correlations, and in the other the double-stranded DNA is taken to have an intrinsic rigidity for bending. In both cases single strands are completely flexible. The change in the elastic constant for the flexible case due to thermally generated bubbles is obtained exactly. For the case of intrinsic rigidity, the elastic constant is found to be proportional to the square root of the bubble number fluctuation.

  16. Extraktion von DNA

    NASA Astrophysics Data System (ADS)

    Pöpping, Bert; Unterberger, Claudia

    Eine DNA-gestützte Analytik spielt im Lebensmittelbereich eine große Rolle. So wird die PCR bzw. die Real Time PCR z. B. für den Nachweis von Krankheitserreger in Lebensmitteln, zur Tier- und Pflanzenartendifferenzierung und den Nachweis von gentechnologisch veränderten Organismen eingesetzt [1]. Grundvoraussetzung für die sehr sensitiven molekularbiologischen Methoden ist eine saubere und kontaminationsfreie Nukleinsäure [2]. Die Qualität der Nukleinsäure entscheidet über Erfolg und Misserfolg der anschließenden molekularbiologischen Analytik. Deshalb werden im Bereich der Lebensmittelanalytik hohe Anforderungen an das jeweilige DNA-Extraktionsprotokoll gestellt. Durch die Anwendung eines geeigneten Extraktionsverfahrens soll die nachzuweisende DNA möglichst in hochmolekularer Form und frei von die nachfolgende Analytik hemmenden Substanzen vorliegen [1]. Gerade hier stellt die Natur der Lebensmittelmatrix eine besondere Herausforderung dar. Matrixkomponenten wie Fette, Zucker, Proteine und sekundäre Inhaltsstoffe erschweren die DNA-Extraktion und können, wenn sie nicht durch die Extraktion vollständig entfernt werden, zu einer Inhibierung der PCR führen [3]. Des Weiteren müssen auf der Matrixoberfläche vorhandene DNA-abbauende Enzyme gehemmt werden [1]. Daneben spielt der Einfluss verschiedener chemischer und physikalischer Parameter (pH-Wert, Temperatur, Enzyme, Scherkräfte) bei der Lebensmittelproduktion eine große Rolle für die Qualität der extrahierten DNA. So führen z. B. hohe Temperaturen und saure pH-Werte während der Lebensmittelverarbeitung zu einer Fragmentierung der DNA. Auch die physikalischen und chemischen Bedingungen der verwendeten Extraktionsmethode beeinflussen die Qualität der DNA [2]. Bleiben nach der Extraktion organische Lösungsmittel (Phenol, Ethanol), Enzyme, Proteine oder Salze zurück, können diese ebenfalls eine nachfolgende PCR inhibieren. Um eine Inhibition der PCR auszuschließen, sollten in der

  17. Mystic Reflection Groups

    NASA Astrophysics Data System (ADS)

    Bazlov, Yuri; Berenstein, Arkady

    2014-04-01

    This paper aims to systematically study mystic reflection groups that emerged independently in the paper [Selecta Math. (N.S.) 14 (2009), 325-372] by the authors and in the paper [Algebr. Represent. Theory 13 (2010), 127-158] by Kirkman, Kuzmanovich and Zhang. A detailed analysis of this class of groups reveals that they are in a nontrivial correspondence with the complex reflection groups G(m,p,n). We also prove that the group algebras of corresponding groups are isomorphic and classify all such groups up to isomorphism.

  18. DNA and RNA technology in soil biodiversity

    NASA Astrophysics Data System (ADS)

    Pereg, Lily

    2016-04-01

    DNA technology has come a long way and state of the art techniques are currently used in the analysis of soil biodiversity. Current methods will be presented and their strengths and limitations discussed. RNA technology, for the study of gene expression and potential activity of functional groups in the soil, is lagging behind, mostly due to the difficulties of extracting stable RNA from the soil. The potentials and challenges of adopting RNA technology for soil analysis will be discussed.

  19. DNA banking and DNA databanking by academic and commercial laboratories

    SciTech Connect

    McEwen, J.E. |; Reilly, P.R.

    1994-09-01

    The advent of DNA-based testing is giving rise to DNA banking (the long-term storage of cells, transformed cell lines, or extracted DNA for subsequent retrieval and analysis) and DNA data banking (the indefinite storage of information derived from DNA analysis). Large scale acquisition and storage of DNA and DNA data has important implications for the privacy rights of individuals. A survey of 148 academically based and commercial DNA diagnostic laboratories was conducted to determine: (1) the extent of their DNA banking activities; (2) their policies and experiences regarding access to DNA samples and data; (3) the quality assurance measures they employ; and (4) whether they have written policies and/or depositor`s agreements addressing specific issues. These issues include: (1) who may have access to DNA samples and data; (2) whether scientists may have access to anonymous samples or data for research use; (3) whether they have plans to contact depositors or retest samples if improved tests for a disorder become available; (4) disposition of samples at the end of the contract period if the laboratory ceases operations, if storage fees are unpaid, or after a death or divorce; (5) the consequence of unauthorized release, loss, or accidental destruction of samples; and (6) whether depositors may share in profits from the commercialization of tests or treatments developed in part from studies of stored DNA. The results suggest that many laboratories are banking DNA, that many have already amassed a large number of samples, and that a significant number plan to further develop DNA banking as a laboratory service over the next two years. Few laboratories have developed written policies governing DNA banking, and fewer still have drafted documents that define the rights and obligations of the parties. There may be a need for increased regulation of DNA banking and DNA data banking and for better defined policies with respect to protecting individual privacy.

  20. Anthocyanidins modulate the activity of human DNA topoisomerases I and II and affect cellular DNA integrity.

    PubMed

    Habermeyer, Michael; Fritz, Jessica; Barthelmes, Hans U; Christensen, Morten O; Larsen, Morten K; Boege, Fritz; Marko, Doris

    2005-09-01

    In the present study, we investigated the effect of anthocyanidins on human topoisomerases I and II and its relevance for DNA integrity within human cells. Anthocyanidins bearing vicinal hydroxy groups at the B-ring (delphinidin, DEL; cyanidin, CY) were found to potently inhibit the catalytic activity of human topoisomerases I and II, without discriminating between the IIalpha and the IIbeta isoforms. However, in contrast to topoisomerase poisons, DEL and CY did not stabilize the covalent DNA-topoisomerase intermediates (cleavable complex) of topoisomerase I or II. Using recombinant topoisomerase I, the presence of CY or DEL (> or = 1 microM) effectively prohibited the stabilization of the cleavable complex by the topoisomerase I poison camptothecin. We furthermore investigated whether the potential protective effect vs topoisomerase I poisons is reflected also on the cellular level, affecting the DNA damaging properties of camptothecin. Indeed, in HT29 cells, low micromolar concentrations of DEL (1-10 microM) significantly diminished the DNA strand breaking effect of camptothecin (100 microM). However, at concentrations > or = 50 microM, all anthocyanidins tested (delphinidin, cyanidin, malvidin, pelargonidin, and paeonidin), including those not interfering with topoisomerases, were found to induce DNA strand breaks in the comet assay. All of these analogues were able to compete with ethidium bromide for the intercalation into calf thymus DNA and to replace the minor groove binder Hoechst 33258. These data indicate substantial affinity to double-stranded DNA, which might contribute at least to the DNA strand breaking effect of anthocyanidins at higher concentrations (> or = 50 microM). PMID:16167831

  1. Genomes, neurotoxins and biology of Clostridium botulinum Group I and Group II

    PubMed Central

    Carter, Andrew T.; Peck, Michael W.

    2015-01-01

    Recent developments in whole genome sequencing have made a substantial contribution to understanding the genomes, neurotoxins and biology of Clostridium botulinum Group I (proteolytic C. botulinum) and C. botulinum Group II (non-proteolytic C. botulinum). Two different approaches are used to study genomics in these bacteria; comparative whole genome microarrays and direct comparison of complete genome DNA sequences. The properties of the different types of neurotoxin formed, and different neurotoxin gene clusters found in C. botulinum Groups I and II are explored. Specific examples of botulinum neurotoxin genes are chosen for an in-depth discussion of neurotoxin gene evolution. The most recent cases of foodborne botulism are summarised. PMID:25445012

  2. DNA Structural Correlation in Short and Long Ranges.

    PubMed

    Gu, Chan; Zhang, Jun; Yang, Y Isaac; Chen, Xi; Ge, Hao; Sun, Yujie; Su, Xiaodong; Yang, Lijiang; Xie, Sunney; Gao, Yi Qin

    2015-11-01

    Recent single-molecule measurements have revealed the DNA allostery in protein/DNA binding. MD simulations showed that this allosteric effect is associated with the deformation properties of DNA. In this study, we used MD simulations to further investigate the mechanism of DNA structural correlation, its dependence on DNA sequence, and the chemical modification of the bases. Besides a random sequence, poly d(AT) and poly d(GC) are also used as simpler model systems, which show the different bending and twisting flexibilities. The base-stacking interactions and the methyl group on the 5-carbon site of thymine causes local structures and flexibility to be very different for the two model systems, which further lead to obviously different tendencies of the conformational deformations, including the long-range allosteric effects. PMID:26439165

  3. Reconstructing phylogeny from the multifractal spectrum of mitochondrial DNA

    NASA Astrophysics Data System (ADS)

    Glazier, James A.; Raghavachari, Sridhar; Berthelsen, Cheryl L.; Skolnick, Mark H.

    1995-03-01

    Conventional methods of phylogenetic reconstruction from DNA sequences require simplified models of evolutionary dynamics. We present a method based on fractal analysis to reconstruct the evolutionary history of organisms from mitochondrial DNA sequences. We map animal mtDNA into four-dimensional random walks and estimate their long range correlations using multifractal spectra. We see systematic changes in correlations in mtDNA sequences across taxonomic lines, which translate into changes in the scaling of the random walks. We use cluster analysis to group the multifractal spectra and obtain the phylogeny of the organisms. Though our method uses no a priori assumptions and is independent of gene order, it yields phylogenetic relationships broadly consistent with established results. Several recent papers have analyzed DNA using fractal analysis and have found long range correlations. However, no one has succeeded in using them to deduce biologically significant relationships.

  4. Cytotoxic activity and DNA-binding properties of isoeuxanthone derivatives.

    PubMed

    Wang, Hui Fang; Yan, Hong; Gao, Xianghua; Niu, Baolong; Guo, Ruijie; Wei, Liqiao; Xu, Bingshe; Tang, Ning

    2014-01-01

    In this study, the interactions of different groups substituted isoeuxanthone derivatives with calf thymus DNA (ct DNA) were investigated by spectrophotometric methods and viscosity measurements. Results indicated that the xanthone derivatives could intercalate into the DNA base pairs by the plane of xanthone ring and the various substituents may influence the binding affinity with DNA according to the calculated quenching constant values. Furthermore, two tumor cell lines including the human cervical cancer cell line (HeLa) and human hepatocellular liver carcinoma cell line (HepG2) were used to evaluate the cytotoxic activities of xanthone derivatives by acid phosphatase assay. Analyses showed that the oxiranylmethoxy substituted xanthone exhibited more effective cytotoxic activity against the cancer cells than the other substituted xanthones. The effects on the inhibition of tumor cells in vitro agreed with the studies of DNA-binding. PMID:24583780

  5. Computational Study of Phosphate Vibrations as Reporters of DNA Hydration.

    PubMed

    Floisand, D J; Corcelli, S A

    2015-10-15

    The sensitivity of the phosphate asymmetric stretch vibrational frequency to DNA hydration was investigated with molecular dynamics (MD) simulations and a spectroscopic map relating the vibrational frequency to the electrostatics of its environment. 95% of the phosphate vibrational frequency shift in fully hydrated DNA was due to water within two hydration layers. The phosphate vibration was relatively insensitive to water in the major and minor grooves and to the sodium counterions but was enormously sensitive to water interacting with the DNA backbone. Comparisons to experimental measurements on DNA as a function of relative humidity suggest that one water molecule per phosphate group likely persists at the lowest values of the relative humidity. Finally, the calculated spectral diffusion dynamics show that water in the vicinity of the DNA backbone is slowed by a factor of ∼5, in agreement with NMR and solvation dynamics experiments, as well as previous MD simulations. PMID:26722770

  6. Highly sensitive DNA sensor based on polypyrrole nanowire

    NASA Astrophysics Data System (ADS)

    Mai, Anh Tuan; Duc, Thanh Pham; Thi, Xuan Chu; Nguyen, Minh Hieu; Nguyen, Hoang Hai

    2014-08-01

    This paper describes the development of a DNA sensor based on polypyrrole nanowire. By using potentiostatic technique, in the presence of gelatin as the soft mold, the polypyrrole nanowires were synthesized on the surface of the micro-sensor. The surface enhanced Raman spectroscopy shows that the Nsbnd H ends of the polypyrrole nanowires orientate upward from the surface facilitating the DNA probe immobilization through the simple linkage with the phosphate groups of the probe DNA. The label-free signal readout was carried out by lock-in amplifier technique. The response time of the DNA sensor is 10 s and the measurement time was 5 min. The lowest detectable concentration of Escherichia coli DNA was 0.1 nM.

  7. DNATCO: assignment of DNA conformers at dnatco.org.

    PubMed

    Černý, Jiří; Božíková, Paulína; Schneider, Bohdan

    2016-07-01

    The web service DNATCO (dnatco.org) classifies local conformations of DNA molecules beyond their traditional sorting to A, B and Z DNA forms. DNATCO provides an interface to robust algorithms assigning conformation classes called NTC: to dinucleotides extracted from DNA-containing structures uploaded in PDB format version 3.1 or above. The assigned dinucleotide NTC: classes are further grouped into DNA structural alphabet NTA: , to the best of our knowledge the first DNA structural alphabet. The results are presented at two levels: in the form of user friendly visualization and analysis of the assignment, and in the form of a downloadable, more detailed table for further analysis offline. The website is free and open to all users and there is no login requirement. PMID:27150812

  8. [Polymorphism of LW blood group gene in Chinese population].

    PubMed

    Su, Yu-Qing; Yu, Qiong; Liu, Xu; Liang, Yan-Lian; Wei, Tian-Li

    2008-06-01

    In order to study the polymorphism of Landsteiner-Wiener (LW) blood group gene in Chinese population, peripheral blood samples anticoagulated with EDTA from 160 unrelated volunteer blood donors were randomly collected, and genomic DNA were extracted. 160 DNA samples were analyzed for exon 1 of LW gene by direct DNA sequencing, and detected for LWa/LWb allele by improved PCR-SSP genotyping. The results showed that all LW allele in 160 donors were LWa homozygous, and the LWa allele occurred commonly. In conclusion, LWa allele occurs with incidence of 100% of donors in this study, while LWb allele has not been found in Chinese population. PMID:18549656

  9. DNA Nanotechnology for Cancer Therapy

    PubMed Central

    Kumar, Vinit; Palazzolo, Stefano; Bayda, Samer; Corona, Giuseppe; Toffoli, Giuseppe; Rizzolio, Flavio

    2016-01-01

    DNA nanotechnology is an emerging and exciting field, and represents a forefront frontier for the biomedical field. The specificity of the interactions between complementary base pairs makes DNA an incredible building material for programmable and very versatile two- and three-dimensional nanostructures called DNA origami. Here, we analyze the DNA origami and DNA-based nanostructures as a drug delivery system. Besides their physical-chemical nature, we dissect the critical factors such as stability, loading capability, release and immunocompatibility, which mainly limit in vivo applications. Special attention was dedicated to highlighting the boundaries to be overcome to bring DNA nanostructures closer to the bedside of patients. PMID:27022418

  10. DNA Nanotechnology for Cancer Therapy.

    PubMed

    Kumar, Vinit; Palazzolo, Stefano; Bayda, Samer; Corona, Giuseppe; Toffoli, Giuseppe; Rizzolio, Flavio

    2016-01-01

    DNA nanotechnology is an emerging and exciting field, and represents a forefront frontier for the biomedical field. The specificity of the interactions between complementary base pairs makes DNA an incredible building material for programmable and very versatile two- and three-dimensional nanostructures called DNA origami. Here, we analyze the DNA origami and DNA-based nanostructures as a drug delivery system. Besides their physical-chemical nature, we dissect the critical factors such as stability, loading capability, release and immunocompatibility, which mainly limit in vivo applications. Special attention was dedicated to highlighting the boundaries to be overcome to bring DNA nanostructures closer to the bedside of patients. PMID:27022418

  11. What Makes Groups Tick.

    ERIC Educational Resources Information Center

    Allcorn, Seth

    1985-01-01

    By reviewing this analysis of the behavior of both groups and individuals in groups, human resources managers can learn to tell whether committees, task forces, and departments may be encouraging or inhibiting the work they set out to do. (Author)

  12. DNA polymerase having modified nucleotide binding site for DNA sequencing

    DOEpatents

    Tabor, S.; Richardson, C.

    1997-03-25

    A modified gene encoding a modified DNA polymerase is disclosed. The modified polymerase incorporates dideoxynucleotides at least 20-fold better compared to the corresponding deoxynucleotides as compared with the corresponding naturally-occurring DNA polymerase. 6 figs.

  13. Quantification of human mitochondrial DNA using synthesized DNA standards.

    PubMed

    Kavlick, Mark F; Lawrence, Helen S; Merritt, R Travis; Fisher, Constance; Isenberg, Alice; Robertson, James M; Budowle, Bruce

    2011-11-01

    Successful mitochondrial DNA (mtDNA) forensic analysis depends on sufficient quantity and quality of mtDNA. A real-time quantitative PCR assay was developed to assess such characteristics in a DNA sample, which utilizes a duplex, synthetic DNA to ensure optimal quality assurance and quality control. The assay's 105-base pair target sequence facilitates amplification of degraded DNA and is minimally homologous to nonhuman mtDNA. The primers and probe hybridize to a region that has relatively few sequence polymorphisms. The assay can also identify the presence of PCR inhibitors and thus indicate the need for sample repurification. The results show that the assay provides information down to 10 copies and provides a dynamic range spanning seven orders of magnitude. Additional experiments demonstrated that as few as 300 mtDNA copies resulted in successful hypervariable region amplification, information that permits sample conservation and optimized downstream PCR testing. The assay described is rapid, reliable, and robust. PMID:21883207

  14. DNA polymerase having modified nucleotide binding site for DNA sequencing

    DOEpatents

    Tabor, Stanley; Richardson, Charles

    1997-01-01

    Modified gene encoding a modified DNA polymerase wherein the modified polymerase incorporates dideoxynucleotides at least 20-fold better compared to the corresponding deoxynucleotides as compared with the corresponding naturally-occurring DNA polymerase.

  15. The GROOP Effect: Groups Mimic Group Actions

    ERIC Educational Resources Information Center

    Tsai, Jessica Chia-Chin; Sebanz, Natalie; Knoblich, Gunther

    2011-01-01

    Research on perception-action links has focused on an interpersonal level, demonstrating effects of observing individual actions on performance. The present study investigated perception-action matching at an inter-group level. Pairs of participants responded to hand movements that were performed by two individuals who used one hand each or they…

  16. GROUP ASPIRATIONS AND GROUP COPING BEHAVIOR.

    ERIC Educational Resources Information Center

    MEDOW, HERMAN; ZANDER, ALVIN

    THIS RESEARCH PROJECT WAS CONCERNED WITH THE EFFECTS OF CERTAIN INTERNAL AND EXTERNAL CONDITIONS UPON THE SELECTION OF A GROUP'S LEVEL OF ASPIRATION AND THE EFFECTS OF THESE CONDITIONS ON MEMBERS' COPING BEHAVIOR. SEVEN EXPERIMENTS WERE DESIGNED WHICH UTILIZED MALE HIGH SCHOOL STUDENTS OF SUBURBAN SCHOOLS AS SUBJECTS. RESULTS OBTAINED FROM THE…

  17. Impact of Alternative DNA Structures on DNA Damage, DNA Repair, and Genetic Instability

    PubMed Central

    Wang, Guliang; Vasquez, Karen M.

    2014-01-01

    Repetitive genomic sequences can adopt a number of alternative DNA structures that differ from the canonical B-form duplex (i.e. non-B DNA). These non-B DNA-forming sequences have been shown to have many important biological functions related to DNA metabolic processes; for example, they may have regulatory roles in DNA transcription and replication. In addition to these regulatory functions, non-B DNA can stimulate genetic instability in the presence or absence of DNA damage, via replication-dependent and/or replication-independent pathways. This review focuses on the interactions of non-B DNA conformations with DNA repair proteins and how these interactions impact genetic instability. PMID:24767258

  18. The Wisdom of Groups

    ERIC Educational Resources Information Center

    Herreid, Clyde Freeman

    2009-01-01

    What is it about small groups that make them so powerful? The answer is straightforward: Groups tend to solve problems better than even the brightest individuals because "many hands make light work," and "two heads are better than one." This is especially true when the groups are diverse and individuals act somewhat independently. In this month's…

  19. Working Group 7 Summary

    SciTech Connect

    Nagaitsev S.; Berg J.

    2012-06-10

    The primary subject of working group 7 at the 2012 Advanced Accelerator Concepts Workshop was muon accelerators for a muon collider or neutrino factory. Additionally, this working group included topics that did not fit well into other working groups. Two subjects were discussed by more than one speaker: lattices to create a perfectly integrable nonlinear lattice, and a Penning trap to create antihydrogen.

  20. Internet minimal group paradigm.

    PubMed

    Amichai-Hamburger, Yair

    2005-04-01

    Over many years, social psychologists have sought to understand what causes individuals to form themselves into groups. Initially, it was believed that groups were formed when people bonded around a common goal. Later, it was found that, when individuals were divided into groups on a random basis, this in itself was sufficient for them to feel part of a group and show a preference for their own group over others. Since the environment in cyberspace is different from that of the offline world, for example, there is no physical proximity between participants; it may be assumed that it would be difficult to achieve feelings of affiliation among potential or actual group members. This pioneer study seeks to discover which components are requisite to the creation of a group identity among individuals surfing the Internet. For this experiment, 24 people were divided into two Internet chat groups according to their intuitive preference in a decision-making task. It was found that group members perceived their own group performance as superior on a cognitive task as compared with that of the other group. These results demonstrate that for surfers, the Internet experience is very real and even a trivial allocation of people to a group is likely to create a situation of ingroup favoritism. PMID:15938653

  1. Practice and Group Learning

    ERIC Educational Resources Information Center

    Hager, Paul

    2014-01-01

    Although learning has always been a central topic for philosophy of education, little attention has been paid to the notion of group learning. This article outlines and discusses some plausible examples of group learning. Drawing on these examples, various principles and issues that surround the notion of group learning are identified and…

  2. Internet Discussion Groups.

    ERIC Educational Resources Information Center

    Bull, Glen; Bull, Gina; Sigmon, Tim

    1997-01-01

    Discusses newsgroups, listservs, and Web-based discussion groups. Highlights include major categories of international USENET discussion groups; newsgroups versus mailing lists; newsreaders; news servers; newsgroup subscriptions; newsgroups versus Web discussion groups; linking newsgroups, mailing lists, and the Web; and setting up a news host. A…

  3. Change through Group Work.

    ERIC Educational Resources Information Center

    McAllan, Les; Friedman, Amy; Spears, Evans

    Perhaps the most well known treatment modalities in the field of prevention and treatment of addiction are groups. Group settings serve to bring individuals with addictions together at one time in one place to work on relevant issues together. Groups may serve as a safe environment for learning new social and relationship skills, gaining…

  4. Infant Group Care Risks.

    ERIC Educational Resources Information Center

    Kendall, Earline D.

    Children under 3 years of age who are in group care face special health risks. The U.S. Centers for Disease Control indicate the existence of a causal relationship between infant group day care and certain diseases that are spread through contact at day care centers. Children in group care who are still in diapers are especially vulnerable to…

  5. Integrated Play Groups

    ERIC Educational Resources Information Center

    Glovak, Sandra

    2007-01-01

    As an occupational therapist running social play groups with sensory integration for children on the autism spectrum, the author frequently doubted the wisdom of combining several children on the spectrum into a group. In fact, as the owner of a clinic she said, "No more!" The groups seemed like a waste of parents' time and money, and she refused…

  6. Parent Group Spotlight

    ERIC Educational Resources Information Center

    Parenting for High Potential, 2014

    2014-01-01

    This issue's "Parent Group Spotlight" features Deborah Simon, president of West Sound Gifted, Talented & Twice-Exceptional (WSGT2e), who started a parent group in Washington in 2013. In just one year, this small, but mighty group has held community forums, attended school board meetings, and helped influence local gifted programming.…

  7. Towards a DNA Nanoprocessor: Reusable Tile-Integrated DNA Circuits.

    PubMed

    Gerasimova, Yulia V; Kolpashchikov, Dmitry M

    2016-08-22

    Modern electronic microprocessors use semiconductor logic gates organized on a silicon chip to enable efficient inter-gate communication. Here, arrays of communicating DNA logic gates integrated on a single DNA tile were designed and used to process nucleic acid inputs in a reusable format. Our results lay the foundation for the development of a DNA nanoprocessor, a small and biocompatible device capable of performing complex analyses of DNA and RNA inputs. PMID:27430161

  8. DNA Methylation in Osteoarthritis

    PubMed Central

    den Hollander, Wouter; Meulenbelt, Ingrid

    2015-01-01

    Osteoarthritis (OA) is a prevalent disease of articular joints and primarily characterized by degradation and calcification of articular cartilage. Presently, no effective treatment other than pain relief exists and patients ultimately need to undergo replacement surgery of the affected joint. During disease progression articular chondrocytes, the single cell type present in articular cartilage, show altered transcriptional profiles and undergo phenotypic changes that resemble the terminal differentiation route apparent in growth plate chondrocytes. Hence, given its prominent function in both regulating gene expression and maintaining cellular phenotypes, DNA methylation of CpG dinucleotides is intensively studied in the context of OA. An increasing number of studies have been published that employed a targeted approach on genes known to play a role in OA pathophysiology. As of such, it has become clear that OA responsive DNA methylation changes seem to mediate disease associated aberrant gene expression. Furthermore, established OA susceptibility alleles such as GDF5 and DIO2 appear to confer OA risk via DNA methylation and respective pathophysiological expression changes. In more recent years, genome wide profiling of DNA methylation in OA affected articular cartilage has emerged as a powerful tool to address the epigenetic changes in their entirety, which has resulted in the identification of putative patient subgroups as well as generic OA associated pathways. PMID:27019616

  9. Enzymatic DNA molecules

    NASA Technical Reports Server (NTRS)

    Joyce, Gerald F. (Inventor); Breaker, Ronald R. (Inventor)

    1998-01-01

    The present invention discloses deoxyribonucleic acid enzymes--catalytic or enzymatic DNA molecules--capable of cleaving nucleic acid sequences or molecules, particularly RNA, in a site-specific manner, as well as compositions including same. Methods of making and using the disclosed enzymes and compositions are also disclosed.

  10. TRANSFECTION WITH BACULOVIRUS DNA

    EPA Science Inventory

    Purified DNA from the nuclear polyhedrosis viruses of Autographa californica (AcM NPV) and Rachiplusia ou (RoM NPV) were found to be infectious in TN-368 cells employing the calcium phosphate precipitation technique (F.L. Graham and A.J. van der Eb, Virology, 52, 456-467, 1973). ...

  11. DNA Replication Origins

    PubMed Central

    Leonard, Alan C.; Méchali, Marcel

    2013-01-01

    The onset of genomic DNA synthesis requires precise interactions of specialized initiator proteins with DNA at sites where the replication machinery can be loaded. These sites, defined as replication origins, are found at a few unique locations in all of the prokaryotic chromosomes examined so far. However, replication origins are dispersed among tens of thousands of loci in metazoan chromosomes, thereby raising questions regarding the role of specific nucleotide sequences and chromatin environment in origin selection and the mechanisms used by initiators to recognize replication origins. Close examination of bacterial and archaeal replication origins reveals an array of DNA sequence motifs that position individual initiator protein molecules and promote initiator oligomerization on origin DNA. Conversely, the need for specific recognition sequences in eukaryotic replication origins is relaxed. In fact, the primary rule for origin selection appears to be flexibility, a feature that is modulated either by structural elements or by epigenetic mechanisms at least partly linked to the organization of the genome for gene expression. PMID:23838439

  12. DNA Methylation in Osteoarthritis.

    PubMed

    den Hollander, Wouter; Meulenbelt, Ingrid

    2015-12-01

    Osteoarthritis (OA) is a prevalent disease of articular joints and primarily characterized by degradation and calcification of articular cartilage. Presently, no effective treatment other than pain relief exists and patients ultimately need to undergo replacement surgery of the affected joint. During disease progression articular chondrocytes, the single cell type present in articular cartilage, show altered transcriptional profiles and undergo phenotypic changes that resemble the terminal differentiation route apparent in growth plate chondrocytes. Hence, given its prominent function in both regulating gene expression and maintaining cellular phenotypes, DNA methylation of CpG dinucleotides is intensively studied in the context of OA. An increasing number of studies have been published that employed a targeted approach on genes known to play a role in OA pathophysiology. As of such, it has become clear that OA responsive DNA methylation changes seem to mediate disease associated aberrant gene expression. Furthermore, established OA susceptibility alleles such as GDF5 and DIO2 appear to confer OA risk via DNA methylation and respective pathophysiological expression changes. In more recent years, genome wide profiling of DNA methylation in OA affected articular cartilage has emerged as a powerful tool to address the epigenetic changes in their entirety, which has resulted in the identification of putative patient subgroups as well as generic OA associated pathways. PMID:27019616

  13. DNA-coated microcrystals.

    PubMed

    Kreiner, Michaela; Fuglevand, Geeta; Moore, Barry D; Parker, Marie-Claire

    2005-06-01

    Coprecipitation leads to self-assembly of bioactive DNA on the surface of salt, sugar or amino-acid crystals and provides a rapid inexpensive immobilization method suitable for preparing dry-powder formulations of nucleic acids, useful for storage, imaging and drug delivery. PMID:15917916

  14. Nutrients and DNA Methylation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Epigenetics is a new mechanism responsible for development, aging, and disease process such as cancer development. One major epigenetic phenomenon is DNA methylation, which attributes to gene expression and integrity. Deepening the knowledge on one-carbon metabolism is very important to understandin...

  15. Automated DNA Sequencing System

    SciTech Connect

    Armstrong, G.A.; Ekkebus, C.P.; Hauser, L.J.; Kress, R.L.; Mural, R.J.

    1999-04-25

    Oak Ridge National Laboratory (ORNL) is developing a core DNA sequencing facility to support biological research endeavors at ORNL and to conduct basic sequencing automation research. This facility is novel because its development is based on existing standard biology laboratory equipment; thus, the development process is of interest to the many small laboratories trying to use automation to control costs and increase throughput. Before automation, biology Laboratory personnel purified DNA, completed cycle sequencing, and prepared 96-well sample plates with commercially available hardware designed specifically for each step in the process. Following purification and thermal cycling, an automated sequencing machine was used for the sequencing. A technician handled all movement of the 96-well sample plates between machines. To automate the process, ORNL is adding a CRS Robotics A- 465 arm, ABI 377 sequencing machine, automated centrifuge, automated refrigerator, and possibly an automated SpeedVac. The entire system will be integrated with one central controller that will direct each machine and the robot. The goal of this system is to completely automate the sequencing procedure from bacterial cell samples through ready-to-be-sequenced DNA and ultimately to completed sequence. The system will be flexible and will accommodate different chemistries than existing automated sequencing lines. The system will be expanded in the future to include colony picking and/or actual sequencing. This discrete event, DNA sequencing system will demonstrate that smaller sequencing labs can achieve cost-effective the laboratory grow.

  16. Field Deployable DNA analyzer

    SciTech Connect

    Wheeler, E; Christian, A; Marion, J; Sorensen, K; Arroyo, E; Vrankovich, G; Hara, C; Nguyen, C

    2005-02-09

    This report details the feasibility of a field deployable DNA analyzer. Steps for swabbing cells from surfaces and extracting DNA in an automatable way are presented. Since enzymatic amplification reactions are highly sensitive to environmental contamination, sample preparation is a crucial step to make an autonomous deployable instrument. We perform sample clean up and concentration in a flow through packed bed. For small initial samples, whole genome amplification is performed in the packed bed resulting in enough product for subsequent PCR amplification. In addition to DNA, which can be used to identify a subject, protein is also left behind, the analysis of which can be used to determine exposure to certain substances, such as radionuclides. Our preparative step for DNA analysis left behind the protein complement as a waste stream; we determined to learn if the proteins themselves could be analyzed in a fieldable device. We successfully developed a two-step lateral flow assay for protein analysis and demonstrate a proof of principle assay.

  17. DNA tagged microparticles

    DOEpatents

    Farquar, George R.; Leif, Roald N.; Wheeler, Elizabeth

    2016-03-22

    In one embodiment, a product includes a plurality of particles, each particle including: a carrier that includes a non-toxic material; and at least one DNA barcode coupled to the carrier, where the particles each have a diameter in a range from about 1 nanometer to about 100 microns.

  18. Making environmental DNA count.

    PubMed

    Kelly, Ryan P

    2016-01-01

    The arc of reception for a new technology or method--like the reception of new information itself--can pass through predictable stages, with audiences' responses evolving from 'I don't believe it', through 'well, maybe' to 'yes, everyone knows that' to, finally, 'old news'. The idea that one can sample a volume of water, sequence DNA out of it, and report what species are living nearby has experienced roughly this series of responses among biologists, beginning with the microbial biologists who developed genetic techniques to reveal the unseen microbiome. 'Macrobial' biologists and ecologists--those accustomed to dealing with species they can see and count--have been slower to adopt such molecular survey techniques, in part because of the uncertain relationship between the number of recovered DNA sequences and the abundance of whole organisms in the sampled environment. In this issue of Molecular Ecology Resources, Evans et al. (2015) quantify this relationship for a suite of nine vertebrate species consisting of eight fish and one amphibian. Having detected all of the species present with a molecular toolbox of six primer sets, they consistently find DNA abundances are associated with species' biomasses. The strength and slope of this association vary for each species and each primer set--further evidence that there is no universal parameter linking recovered DNA to species abundance--but Evans and colleagues take a significant step towards being able to answer the next question audiences tend to ask: 'Yes, but how many are there?' PMID:26768195

  19. A new family of polymerases related to superfamily A DNA polymerases and T7-like DNA-dependent RNA polymerases.

    PubMed

    Iyer, Lakshminarayan M; Abhiman, Saraswathi; Aravind, L

    2008-01-01

    Using sequence profile methods and structural comparisons we characterize a previously unknown family of nucleic acid polymerases in a group of mobile elements from genomes of diverse bacteria, an algal plastid and certain DNA viruses, including the recently reported Sputnik virus. Using contextual information from domain architectures and gene-neighborhoods we present evidence that they are likely to possess both primase and DNA polymerase activity, comparable to the previously reported prim-pol proteins. These newly identified polymerases help in defining the minimal functional core of superfamily A DNA polymerases and related RNA polymerases. Thus, they provide a framework to understand the emergence of both DNA and RNA polymerization activity in this class of enzymes. They also provide evidence that enigmatic DNA viruses, such as Sputnik, might have emerged from mobile elements coding these polymerases. PMID:18834537

  20. A new family of polymerases related to superfamily A DNA polymerases and T7-like DNA-dependent RNA polymerases

    PubMed Central

    Iyer, Lakshminarayan M; Abhiman, Saraswathi; Aravind, L

    2008-01-01

    Using sequence profile methods and structural comparisons we characterize a previously unknown family of nucleic acid polymerases in a group of mobile elements from genomes of diverse bacteria, an algal plastid and certain DNA viruses, including the recently reported Sputnik virus. Using contextual information from domain architectures and gene-neighborhoods we present evidence that they are likely to possess both primase and DNA polymerase activity, comparable to the previously reported prim-pol proteins. These newly identified polymerases help in defining the minimal functional core of superfamily A DNA polymerases and related RNA polymerases. Thus, they provide a framework to understand the emergence of both DNA and RNA polymerization activity in this class of enzymes. They also provide evidence that enigmatic DNA viruses, such as Sputnik, might have emerged from mobile elements coding these polymerases. This article was reviewed by Eugene Koonin and Mark Ragan. PMID:18834537