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At least 1,567 records · Page 87Linked to original sources
PubMed2006

Single-nucleotide polymorphism genotyping by nanoparticle-enhanced surface plasmon resonance imaging measurements of surface ligation reactions.

A sensitive method for the analysis of single nucleotide polymorphisms (SNPs) in genomic DNA that utilizes nanoparticle-enhanced surface plasmon resonance imaging (SPRI) measurements of surface enzymatic ligation reactions on DNA microarrays is demonstrated. SNP identification was achieved by using sequence-specific surface reactions of the enzyme Taq DNA ligase, and the presence of ligation products on the DNA microarray elements was detected using SPRI through the hybridization adsorption of complementary oligonucleotides attached to gold nanoparticles. The use of gold nanoparticles increases the sensitivity of the SPRI so that single bases in oligonucleotides can be successfully identified at a concentration of 1 pM. This sensitivity is amply sufficient for performing multiplexed SNP genotyping by using multiple PCR amplicons and should also allow for the direct detection and identification of SNP sequences from 1 pM unamplified genomic DNA samples with this array-based and label-free SPRI methodology. As a first example of SNP genotyping, three different human genomic DNA samples were screened for a possible point mutation in the BRCA1 gene that is associated with breast cancer.

DNA↗
PubMed2005

Quantitative trait locus analysis reveals two intragenic sites that influence O6-alkylguanine-DNA alkyltransferase activity in peripheral blood mononuclear cells.

The repair of specific types of DNA alkylation damage by O6-alkylguanine-DNA alkyltransferase (MGMT) is a major mechanism of resistance to the carcinogenic and chemotherapeutic effects of certain alkylating agents. MGMT expression levels vary widely between individuals but the underlying causes of this variability are not known. To address this, we used an expressed single nucleotide polymorphism (SNP) and demonstrated that the MGMT alleles are frequently expressed at different levels in peripheral blood mononuclear cells (PBMC). This suggests that there is a genetic component of inter-allelic variation of MGMT levels that maps close to or within the MGMT locus. We then used quantitative trait locus (QTL) analysis using intragenic SNPs and found that there are at least two sites influencing inter-individual variation in PBMC MGMT activity. One is characterized by an SNP at the 3' end of the first intron and the second by two SNPs in the last exon. The latter are in perfect disequilibrium and both result in amino acid substitutions-one of them, Ile143Val, affecting an amino acid close to the Cys145 residue at the active site of MGMT. Using in vitro assays, we further showed that while the Val143 variant did not affect the activity of the protein on methylated DNA substrate, it was more resistant to inactivation by the MGMT pseudosubstrate, O6-(4-bromothenyl)guanine. These findings suggest that further investigations of the potential epidemiological and clinical significance of inherited differences in MGMT expression and activity are warranted.

Base Sequence↗
PubMed2025

Comprehensive analysis of DNA methylome and transcriptome reveals the epigenetic regulation of nitric oxide treatment in delaying apricot fruit senescence.

Apricot produces climacteric fruit, which are perishable after harvest. To elucidate the regulatory role of NO treatment through DNA methylation in post-harvest senescence, apricot fruits were treated with 0.2 mmol/L sodium nitroprusside (SNP) solution for 10 min, with distilled water treatment serving as the control. Treated fruits were then stored at 25°C and 80% relative humidity. Changes in appearance quality, physiological parameters, metabolome profiles, transcriptome dynamics, and DNA methylation patterns were analyzed before and after storage. Results showed that NO treatment delayed apricot softening, increased flavonoid metabolite accumulation, and reduced lipid and abscisic acid accumulation, with these effects correlated to the expression of specific genes and transcription factors. This work reveals the epigenetic regulatory mechanism underlying NO treatment delaying ripening and senescence. Further analysis revealed that the transcription levels of ACO, PAL, UFGT-like, NCED1, PP2C, MYB21, CCoAOMT-like, CYP707A, and ZNF7-like were all correlated with DNA methylation. This indicates that SNP treatment can lead to large changes in DNA methylation levels in apricot fruits, and that the differences in gene transcription levels are associated with the occurrence of hypomethylation and hypermethylation. Collectively, these findings establish an epigenetic framework for post-harvest regulation of apricot fruit, revealing DNA methylation-mediated freshness preservation mechanisms.

DNA Methylation↗
PubMed2004

Single nucleotide polymorphism analysis on melanocortin receptor 1 (MC1R) of Chinese native pig.

Melanocortin receptor 1 (MC1R) gene, one of the important candidate genes for coat color trait, was used to analyze the single nucleotide polymorphism (SNP) in Chinese native pig breeds by PCR-single strand conformation polymorphism (PCR-SSCP). The study had also taken 3 imported pig breeds as control. The results showed that the three mutations G284A, T309C and T364C found in Chinese native pigs were consistent to the mutation found in the European Large Black individuals. However, 68CC or C492T and G728A were only found in the imported individuals, which were obviously different from the Chinese native pigs. Accordingly, we presumed that the coat colors of Chinese native pigs belonged to dominant black color system, which was completely distinct to that of imported pig breeds. Thus it was implied that MC1R gene was not the principal factor affecting the coat color differences of Chinese native pig breeds, but could be used to trace the molecular evolution of pig breeds.

Animals↗
PubMed2000

Analysis of single nucleotide polymorphisms by primer extension and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

A method for typing single nucleotide polymorphisms (SNPs) by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) is described, in which a mass-tagged dideoxynucleoside triphosphate is employed in a primer extension reaction in place of an unmodified dideoxynucleoside triphosphate (ddNTP). The increased mass difference due to the presence of the mass-tag greatly facilitates the accurate identification of the added nucleotide, and is particularly useful for typing heterozygous samples. Twenty commercially available mass-tagged dideoxynucleoside triphosphates were screened for amenability to incorporation by AmpliTaq FS and ThermoSequenase DNA polymerases in single nucleotide primer extension (SNuPE) reactions. Several sample preparation and purification methods were also examined and compared. Float dialysis was found to be a simple, versatile, and effective method for purification of the extension products. High specificity and sensitivity were obtained, and all six possible biallelic SNP heterozygotes were determined accurately using a 44-mer synthetic oligonucleotide target DNA as a model system. Further validation of the method was demonstrated in the analysis of five single-base mutations in exon IV of the human tyrosinase gene. Single nucleotide variations within 182-bp PCR amplicons amplified from three plasmid and three human genomic DNA samples were genotyped at five variable positions, with results in 100% concordance with conventional sequencing. Genotypes were determined accurately at five sequence-tagged sites (STSs).

Base Sequence↗
PubMed2004

Comparative informativeness for linkage of multiple SNPs and single microsatellites.

Single nucleotide polymorphisms (SNPs), or biallelic markers, are popular in genetic linkage studies due to their abundance in the genome, stability, and ease of scoring. We determined the 'information ratio' (IR) of closely spaced SNPs in simulated nuclear families and affected sib pairs (ASPs). (The IR is the ratio of actual average maximum lod score to the maximum lod score attainable if the marker were fully informative.) The nuclear families included parental information, whereas the ASPs did not. We analyzed these SNPs in two ways: (1) using multipoint analysis, and (2) treating the SNPs as 'composite markers' (i.e., haplotypes, as assigned by GENEHUNTER). (3) We also calculated the IR of a single microsatellite marker with multiple alleles and compared with the IR from the SNPs. For each set of input conditions, we simulated 1000 nuclear families, of 2, 3, 4, or 5 children each, as well as 1000 ASPs. We generated SNP marker data for strings of k = 1, 2, 3, 5, 7, and 10 SNP loci, with no recombination (theta = 0) and no linkage disequilibrium among the SNPs. The MAF (minor allele frequency) was either 0.5 or 0.25, and allele frequencies were the same for all k loci in any analysis. We also generated marker data for one single-locus microsatellite marker, with m = 3, 4, 5, 6, 7, and 9 equally frequent alleles. In all simulations, the disease was fully penetrant dominant, and there was no recombination or linkage disequilibrium among markers or between marker and disease. When multipoint analysis was used, we found that 5-7 closely spaced SNPs were usually enough to yield an IR of approximately 100%, for nuclear families of any size. However, for the ASPs, even 7-10 SNPs yielded an IR of only 70-80%. A microsatellite with 9 equally frequent alleles yielded about the same IR (86-88%) as a string of 4-5 SNPs, in nuclear families. SNPs analyzed as 'composite markers' analyses performed worse, due to the inherent ambiguity of SNP haplotyping.

Alleles↗
PubMed2006

Localization of candidate regions for a novel gene for Kartagener syndrome.

Asymmetric positioning of internal organs is a characteristics of vertebrates. The normal left-right anatomic positioning, situs solitus, sometimes does not occur normaly, leading to laterality defects. Studies in animal models have shown that laterality decisions are mediated by a cascade of genes that lead to the asymmetric expression of Nodal, LEFTA, LEFTB and PITX2 in the lateral plate mesoderm. A search for mutations in genes implicated in left-right patterning in animal models allowed genes associated with heterotaxia defects in humans to be identified. However, these genes explain only a small percentage of human situs defects, suggesting that other genes must play a role. In this study, we report a consanguineous family of Turkish origin, composed of two unaffected parents and three children, two of whom presented Kartagener syndrome. On the basis of their family history, we hypothesize autosomal recessive mode of inheritance. A genotype analysis with polymorphic markers did not show linkage with any known genes or loci causing laterality disorders. Array CGH did not detect a duplication or microdeletion greater than 1 Mb as a possible cause. Genome wide screening using 10 K Affymetrix SNP chips was performed, allowing the identification of two regions of autozygosity, one in chromosome 1 and the other on chromosome 7. In the chromosome 1 locus, a strong candidate gene, encoding the kinesin-associated protein 3 (KIF3AP) was not mutated, based on SSCP/heteroduplex analysis and direct sequencing. These data provide a basis for the identification of a novel gene implicated in Kartagener syndrome.

5' Untranslated Regions↗
PubMed2005

novoSNP, a novel computational tool for sequence variation discovery.

Technological improvements shifted sequencing from low-throughput, work-intensive, gel-based systems to high-throughput capillary systems. This resulted in a broad use of genomic resequencing to identify sequence variations in genes and regulatory, as well as extended genomic regions. We describe a software package, novoSNP, that conscientiously discovers single nucleotide polymorphisms (SNPs) and insertion-deletion polymorphisms (INDELs) in sequence trace files in a fast, reliable, and user-friendly way. We compared the performance of novoSNP with that of PolyPhred and PolyBayes on two data sets. The first data set comprised 1028 sequence trace files obtained from diagnostic mutation analyses of SCN1A (neuronal voltage-gated sodium channel alpha-subunit type I gene). The second data set comprised 9062 sequence trace files from a genomic resequencing project aiming at the construction of a high-density SNP map of MAPT (microtubule-associated protein tau gene). Visual inspection of these data sets had identified 38 sequence variations for SCN1A and 488 for MAPT. novoSNP automatically identified all 38 SCN1A variations including five INDELs, while for MAPT only 15 of the 488 variations were not correctly marked. PolyPhred detected far fewer SNPs as compared to novoSNP and missed nearly all INDELs. PolyBayes, designed for the sequence analysis of cloned templates, detected only a limited number of the variations present in the data set. Besides the significant improvement in the automated detection of sequence variations both in diagnostic mutation analyses and in SNP discovery projects, novoSNP also offers a user-friendly interface for inspecting possible genetic variations.

Animals↗
PubMed2003

DHPLC scoring of a SNP between promoter sequences of HMW glutenin x-type alleles at the Glu-D1 locus in wheat.

The promoter regions of HMW glutenin x-type genes at the Glu-D1 locus were surveyed for SNPs within a subpopulation of German bread wheat cultivars. On the basis of the promoter sequences of HMW glutenin subunit genes Glu-A1-x1, Glu-A1-x2, Glu-B1-x1, Glu-B1-x7, Glu-D1-x2, and Glu-D1-x5, an amplification refractory mutation system assay was designed to selectively amplify Dx-specific PCR fragments. Comparative sequence analysis among seven Glu-D1-x2 and seven Glu-D1-x5 wheat cultivars only confirmed a G-A transition in the promoter sequence to be a true polymorphism. SNP scoring by DHPLC of 95 German bread wheat cultivars, with the exception of cv. Anemos, showed that the transition completely agreed with the presence of HMW glutenin subunits 1Dx5 + 1Dy10 in SDS-PAGE. Therefore, the developed DHPLC assay is suitable for high-throughput genotyping to assist the selection of HMW glutenin genes in wheat quality breeding programs.

Alleles↗
PubMed2003

Molecular screening and association studies of retinoid-related orphan receptor gamma (RORC): a positional and functional candidate for type 2 diabetes.

The retinoid-related orphan receptor gamma (RORC) is a member of the nuclear hormone superfamily which maps to the 1q21-q23 region. Linkage of type 2 diabetes (T2DM) to this region is well replicated. Several factors argue that RORC is a strong candidate for T2DM susceptibility within this region. RORC may form heterodimers with peroxisome-proliferator activated receptor gamma, it is expressed at high levels in skeletal muscle, and expression is induced in adipocytes during differentiation. To test the hypothesis that sequence variation in RORC is a risk factor for T2DM, we screened approximately 21kb of DNA for sequence variation, including 11 exons of the RORC gene, a region 1-kb upstream (5' flanking region), intronic regions flanking the exons, and the entire 3' untranslated region (UTR). Screening was performed using single strand conformation polymorphism (SSCP) analysis in Caucasian individuals of northern European ancestry and in African American individuals. We detected 11 single nucleotide polymorphisms (SNPs), ranging from the promoter region to intron 10. We also confirmed 2 SNPs from public databases that were in regions not included in our screening. Only 1 SNP was nonsynonymous, resulting in Ala to Gly at residue 464 (exon 10). All other SNPs were noncoding. One SNP (intron 3) was unique to Caucasians, and three SNPs (Ala464Gly, intron 2, intron 6) were specific to African American subjects. We typed 7 SNPs spanning the gene from the promoter to 3' UTR in unrelated cases with T2DM and controls of Northern European ancestry. We also tested linkage of a microsatellite within the RORC gene. Modest evidence for linkage (LOD=1.47) was seen on two-point analysis, but no linkage to the RORC region was found on multipoint analysis. However, transmission of the microsatellite alleles from parents to affected offspring showed a trend to deviate from the expected 50% (p=0.078). No association of any other SNP with T2DM was found, but the Ala454Gly variant was 3-fold more common among African American patients with diabetes than in controls. SNPs 1, 2 and 4 were in strong linkage disequilibrium (D>0.85) and may constitute a haplotype block. Our data suggest that RORC cannot explain the linkage of T2DM in this region. The role of the unusual Ala454Gly variant will require a much larger study size to evaluate.

Black or African American↗
PubMed2006

Haplotype analysis revealed no association between the PTPN22 gene and RA in a Japanese population.

OBJECTIVE: The protein tyrosine phosphatase non-receptor type 22 (PTPN22) gene is a member of the PTPs that negatively regulate T-cell activation. A missense single nucleotide polymorphism (SNP) in the PTPN22 gene known as R620W was recently reported to be associated with several autoimmune diseases including rheumatoid arthritis (RA). The association was confirmed repeatedly in the populations of North European ancestry. However, the SNP was reported to be non-polymorphic in the Asian populations. Because the gene confers an impact on autoimmune diseases, we attempt to explore an association between PTPN22 gene and RA in a Japanese population without restricting to the SNP, R620W. METHODS: We studied 1128 RA patients and 455 controls. In addition to the SNP, R620W, we selected eight testing SNPs spanning 45 kb over the PTPN22 gene using the International HapMap Project. Genotyping was performed using the TaqMan fluorogenic 5' nuclease assay. Associations between RA and each of the SNPs were estimated by the Fisher's exact test. Haplotype was constructed using the expectation-maximization algorithm. RESULTS: R620W was not polymorphic enough in both the patients and the controls, and was therefore excluded from further analysis. Each allele frequency for the eight other SNPs in both groups was compared and no association was detected. Haplotype analysis also revealed that PTPN22 gene was not associated with RA in a Japanese population. CONCLUSION: We found no association between PTPN22 and RA in a Japanese population. The result suggests that the PTPN22 gene is associated with RA only in a specific ethnic group.

Arthritis, Rheumatoid↗
PubMed2002

Analysis of candidate genes on chromosome 19 in coeliac disease: an association study of the KIR and LILR gene clusters.

Coeliac disease is strongly heritable, with more than half of the genetic susceptibility estimated to come from genes outside the HLA region. Several candidate regions have been suggested from genome-wide linkage studies including chromosome 19q13.4 where linkage has been replicated between populations. The natural killer (NK) cell immunoglobulin-like receptors (KIRs) and leukocyte immunoglobulin-like receptor (LILR, also known as ILT and LIR) gene clusters lie within this region in the leukocyte receptor cluster (LRC). KIR molecules are involved in cytotoxic lymphocyte function and expressed by intraepithelial T and NK cells in the duodenum. We studied 132 unrelated UK Caucasian coeliac patients and their parents together with a control group of 171 UK Caucasians. PCR-SSP for KIR2DL1, KIR2DL2, KIR2DL3, KIR2DL5, LILRA3 (ILT6), LILRA3 deletion and an LILRA3 exon 3 single nucleotide polymorphism (SNP) allowed classification of KIR genotypes into five categories and determination of homozygosity or heterozygosity for the common A and B type KIR haplotypes (as defined in the text) and for the LILRA3 deletion. Case-control analysis found no association of the five KIR genotype categories, the A or B KIR haplotypes, the LILRA3 gene deletion or the LILRA3 exon 3 SNP with coeliac disease. A transmission disequilibrium test also found no association of the A and B KIR haplotypes or the LILRA3 gene deletion with coeliac disease.

Case-Control Studies↗
PubMed2005

A genome-wide linkage analysis of alcoholism on microsatellite and single-nucleotide polymorphism data, using alcohol dependence phenotypes and electroencephalogram measures.

The Collaborative Study on the Genetics of Alcoholism (COGA) is a large-scale family study designed to identify genes that affect the risk for alcoholism and alcohol-related phenotypes. We performed genome-wide linkage analyses on the COGA data made available to participants in the Genetic Analysis Workshop 14 (GAW 14). The dataset comprised 1,350 participants from 143 families. The samples were analyzed on three technologies: microsatellites spaced at 10 cM, Affymetrix GeneChip Human Mapping 10 K Array (HMA10K) and Illumina SNP-based Linkage III Panel. We used ALDX1 and ALDX2, the COGA definitions of alcohol dependence, as well as electrophysiological measures TTTH1 and ECB21 to detect alcoholism susceptibility loci. Many chromosomal regions were found to be significant for each of the phenotypes at a p-value of 0.05. The most significant region for ALDX1 is on chromosome 7, with a maximum LOD score of 2.25 for Affymetrix SNPs, 1.97 for Illumina SNPs, and 1.72 for microsatellites. The same regions on chromosome 7 (96-106 cM) and 10 (149-176 cM) were found to be significant for both ALDX1 and ALDX2. A region on chromosome 7 (112-153 cM) and a region on chromosome 6 (169-185 cM) were identified as the most significant regions for TTTH1 and ECB21, respectively. We also performed linkage analysis on denser maps of markers by combining the SNPs datasets from Affymetrix and Illumina. Adding the microsatellite data to the combined SNP dataset improved the results only marginally. The results indicated that SNPs outperform microsatellites with the densest marker sets performing the best.

Alcoholism↗
PubMed2005

Polymorphism study of seven SNPs at ADH genes in 15 Chinese populations.

It has been shown that the variants of alcohol dehydrogenase (ADH) genes exhibit great diversities among various populations and are associated with susceptibility to alcoholism. To investigate the distribution of SNPs at ADH genes in Chinese populations and the genetic relationship of these groups, we collected 467 individuals from 15 groups distributing widely from north to south in China and genotyped 7 SNPs at ADH genes respectively. The statistic analyses of allele frequencies, estimated haplotype frequencies, pairwise linkage disequilibrium, AMOVA (analysis of molecular variance), pairwise Fst', and cluster analysis indicated (1) that six of these seven SNPs showed great variations in the 15 Chinese populations, and three of them (RsaI, SspI, EcoRI), were confirmed to be informative SNPs. However, the causative SNP ADH1B Arg47His confirmed in case-control studies could not act as significant indicator to distinguish bibulous groups from non-bibulous groups in healthy individuals; (2) haplotypes constructed with ADH SNPs could be used as markers to discern different populations in China, and six-allele haplotype "221211" was the most common one defined in present study; (3) on the basis of SNPs analysis of ADH genes, the 15 populations were grouped into northern groups and southern groups. Moreover, the origin relationship among the populations was indicated according to the results of cluster analysis.

Alcohol Dehydrogenase↗
PubMed2005

Linkage mapping bovine EST-based SNP.

BACKGROUND: Existing linkage maps of the bovine genome primarily contain anonymous microsatellite markers. These maps have proved valuable for mapping quantitative trait loci (QTL) to broad regions of the genome, but more closely spaced markers are needed to fine-map QTL, and markers associated with genes and annotated sequence are needed to identify genes and sequence variation that may explain QTL. RESULTS: Bovine expressed sequence tag (EST) and bacterial artificial chromosome (BAC)sequence data were used to develop 918 single nucleotide polymorphism (SNP) markers to map genes on the bovine linkage map. DNA of sires from the MARC reference population was used to detect SNPs, and progeny and mates of heterozygous sires were genotyped. Chromosome assignments for 861 SNPs were determined by twopoint analysis, and positions for 735 SNPs were established by multipoint analyses. Linkage maps of bovine autosomes with these SNPs represent 4585 markers in 2475 positions spanning 3058 cM. Markers include 3612 microsatellites, 913 SNPs and 60 other markers. Mean separation between marker positions is 1.2 cM. New SNP markers appear in 511 positions, with mean separation of 4.7 cM. Multi-allelic markers, mostly microsatellites, had a mean (maximum) of 216 (366) informative meioses, and a mean 3-lod confidence interval of 3.6 cM Bi-allelic markers, including SNP and other marker types, had a mean (maximum) of 55 (191) informative meioses, and were placed within a mean 8.5 cM 3-lod confidence interval. Homologous human sequences were identified for 1159 markers, including 582 newly developed and mapped SNP. CONCLUSION: Addition of these EST- and BAC-based SNPs to the bovine linkage map not only increases marker density, but provides connections to gene-rich physical maps, including annotated human sequence. The map provides a resource for fine-mapping quantitative trait loci and identification of positional candidate genes, and can be integrated with other data to guide and refine assembly of bovine genome sequence. Even after the bovine genome is completely sequenced, the map will continue to be a useful tool to link observable phenotypes and animal genotypes to underlying genes and molecular mechanisms influencing economically important beef and dairy traits.

Alleles↗
PubMed2001

[Post-genome challenges against inflammatory bowel diseases].

Advancement of genome analysis might give great impact to the treatment of inflammatory bowel diseases(IBD). IBD patients are treated by sulfadrugs, steroids and anti-immune drugs. For difficult cases, leukocytapheresis, beclomethasone dipropionate, anti-TNF therapy, anti-LTB4 therapy and other new methods are applied. Developing epoch-making drugs will be achieved by finding new molecular targets. Histologic identification of dysplasia is important in the surveillance of long-standing ulcerative colitis. The molecular diagnosis is required for the distinction of dysplasia from the regenerative inflammatory changes. P53 immunostaining have been proved useful. Various molecular targets will be taken into discussion as additional procedures. Recent genome analysis have revealed some genetic factors contribute to pathogenesis of IBD, which are HLA, IL4, MUC3, IBD1 locus, IBD2 locus and so on. More information about genes concerning IBD will be provided by analyzing dense SNP map using DNA tip. They will open the way to the tailored therapy.

Biomarkers↗