Scale Single Nucleotide Polymorphism Research Articles

Evolutionary history of endangered and relict tree species Dipteronia sinensis in response to geological and climatic events in the Qinling Mountains and adjacent areas.

Geological and climatic events are considered to profoundly affect the evolution and lineage divergence of plant species. However, the evolutionary histories of tree species that have responded to past geological and climate oscillations in central China's mountainous areas remain mostly unknown. In this study, we assessed the evolutionary history of the endangered and relict tree species Dipteronia sinensis in the Qinling Mountains (QM) and adjacent areas in East Asia based on variations in the complete chloroplast genomes (cpDNA) and reduced‐genomic scale single nucleotide polymorphisms (SNPs). Population structure and phylogenetic analysis based on the cpDNA variations suggested that D. sinensis could be divided into two intraspecific genetic lineages in the eastern and western sides of the QM (EQM and WQM, respectively) in East Asia. Molecular dating suggested that the intraspecific divergence of D. sinensis occurred approximately 39.2 million years ago during the later Paleogene. It was significantly correlated with the orogeny of the QM, where the formation of this significant geographic barrier in the region may have led to the divergence of independent lineages. Bayesian clustering and demographic analysis showed that intraspecific gene flow was restricted between the EQM and WQM lineages. Isolation‐with‐migration analysis indicated that the two genetic lineages experienced significant demographic expansions after the Pleistocene ice ages. However, the genetic admixture was determined in some populations between the two lineages by the large scale of SNP variations due to DNA incompatibility, the large significant population size, and rapid gene flow of nuclear DNA markers. Our results suggest that two different conservation and management units should be constructed for D. sinensis in the EQM and WQM areas. These findings provide novel insights into the unprecedented effects of tectonic changes and climatic oscillations on lineage divergence and plant population evolution in the QM and adjacent areas in East Asia.

Stock composition of adult chum salmon Oncorhynchus keta caught in a setnet fishery estimated using genetic identification, scale patterns, and otolith thermal marking

Three different techniques for stock identification, namely scale patterns, otolith thermal marks, and single-nucleotide polymorphism (SNP) markers, were applied to the same mixture samples of chum salmon collected in a setnet fishery in the Sea of Okhotsk, off Hokkaido, Japan, to evaluate consistency and accuracy among the techniques. The scale pattern and SNP analyses provided similar estimates for two regional origins (Hokkaido and Honshu) with data for 2011–2013; for each year, the Hokkaido-origin stock was dominant in September and early October, but the Honshu-origin stock was dominant by late October. The SNP analysis specified five geographical origins of chum salmon (the Okhotsk coast, Hokkaido and Honshu coasts of the Sea of Japan, and Hokkaido and Honshu coasts of the Pacific), whereas the other techniques did not identify multi-regional origins. Recaptures of otolith-marked chum salmon were used for estimating the proportions of fish originating from the Hokkaido Sea of Japan region; these estimates differed from those based on the SNP analysis. This study demonstrated that scale pattern and SNP analyses have the potential to provide nearly consistent estimates for the two regional origins, and can possibly help us to understand the interception of migrating chum salmon in Japan.

Potential link between genetic polymorphisms of catechol-O-methyltransferase and dopamine receptors and treatment efficacy of risperidone on schizophrenia.

ObjectiveThe current study aimed to explore the association of single nucleotide polymorphisms (SNPs) within catechol-O-methyltransferase (COMT) and dopamine receptors with schizophrenia and genetic association with risperidone treatment response.MethodsA total of 690 schizophrenic patients (case group) were selected and 430 healthy people were included as the controls. All patients received risperidone treatment continuously for 8 weeks. Next, peripheral venous blood samples were collected and were subjected to polymerase chain reaction-restriction fragment length polymorphism to amplify and genotype the SNPs within COMT and dopamine receptors. Then, correlation analysis was conducted between Positive and Negative Syndrome Scale improvement rates and SNPs within COMT and the dopamine receptor gene.ResultsThe allele of DRD1 rs11749676 (A) emerged as a key element in reducing schizophrenia risk with statistical significance (P<0.001). Remarkably, alleles of COMT rs165774 (G), DRD2 rs6277 (T), and DRD3 rs6280 (C) were associated with raised predisposition to schizophrenia (all P<0.001). Regarding DRD1 rs11746641, DRD1 rs11749676, DRD2 rs6277, and DRD3 rs6280, the case group exhibited a lesser frequency of heterozygotes in comparison with wild homozygotes genotype (all P<0.001). SNPs (COMT rs4680, DRD2 rs6275, DRD2 rs1801028, and DRD2 rs6277) were remarkably associated with improvement rates of PANSS total scores (P<0.05). SNPs (COMT rs165599 and DRD2 rs1801028) were significantly associated with risperidone efficacy on negative symptoms (P<0.05).ConclusionCOMT SNPs and dopamine receptor SNPs were correlated with prevalence of schizophrenia and risperidone treatment efficacy of schizophrenia.

Assessment of SNPs for linkage mapping in Eucalyptus: construction of a consensus SNP/microsatellite map from two unrelated pedigrees

Background High throughput genotyping of SNP (Single Nucleotide Polymorphisms) based markers has been developed for an increasing number of plant and animal species. In forest trees large scale SNP development has been approached mainly by amplicon resequencing targeting specific genes for association genetics studies. This approach, although successfully employed in conifers aided by the use of haploid tissue, is technically laborious in diploids because of the very high levels of nucleotide and indel diversity in highly heterozygous tree genomes. Direct SNP development from large in silico sequence resources developed by next-generation sequencing is now a very efficient approach for SNP development in forest trees. We have recently developed a first set of 768 SNPs assayed by the Golden Gate Genotyping Technology for the highly heterozygous genome of Eucalyptus from a mixed Sanger/454 database [1]. We saw that a careful sequence quality assessment and the application of stringent constraints on the SNP surrounding sequences have a significant impact on SNP genotyping performance and polymorphism. With the exception of 72 SNPs specifically selected in 20 candidate genes putatively associated with relevant wood phenotypes, all remaining validated SNPs were randomly picked based solely on in silico quality. In this study we wanted to position these SNPs relative to microsatellites and assess their information content for linkage map construction. To enhance our ability of mapping SNPs we employed two eucalyptus full-sib families involving four different Eucalyptus species.

Abstract B65: Targeted resequencing of the EphB2 gene in colorectal cancer for SNP discovery using next generation sequencing

Abstract Colorectal cancer (CRC) remains a leading cause of cancer morbidity and mortality in the United States. CRC is the third most common cause of deaths in the U. S. with the highest incidence observed in the African American (AA) population compared to other American populations. Our group was the first to implicate the EphB2 tyrosine kinase receptor as a tumor suppressor in prostate cancer (PC), where we reported somatic inactivating mutations occurring in 5%-10% of sporadic tumors. With its role in maintaining normal epithelial architecture and functional data suggestive of a tumor suppressor in both prostate and colon cancer. Recent advances in next-generation sequencing (NGS) afford new opportunities for large scale SNP discovery using a targeted resequencing approach. Several methodologies have been developed for targeted resequencing. Among these methods is the RainDance picoliter massive single-plex PCR assay, which provides the capability to assess up to 10,000 independent amplicons in a single reaction. We designed a RainDance PCR assay containing 700 amplicons, encompassing the entire 225,000bp genomic region of the EphB2 gene. These individual pools of amplicons from each sample were then used as template for NGS and barcoded to generate pools of 16 unique individuals using the Life Technologies SOLiD version 3-Plus system. Utilizing this method we screened 20 colorectal cancer cases and 27 sporadic prostate cancer cases. Thus far we have identified both known and novel variants within the EphB2 gene region among of which are several common coding variants including the 2055A&amp;gt;T nonsense mutation (K1019X) in three of 10 female CRC cases. We will provide data on the number of novel and known single nucleotide polymorphisms (SNPs) for all CRC and PC cases as well as quality assessment and overall performance of the RainDance method and performance metrics for SOliD NGS along with a comparison of utilizing several different analyses pipelines to identify the novel and known SNPs. These data will provide us with unprecedented details of the genetic variation of the EphB2 gene in African Americans. Citation Information: Cancer Epidemiol Biomarkers Prev 2010;19(10 Suppl):B65.

Low linkage disequilibrium in wild Anopheles gambiae s.l. populations

BackgroundIn the malaria vector Anopheles gambiae, understanding diversity in natural populations and genetic components of important phenotypes such as resistance to malaria infection is crucial for developing new malaria transmission blocking strategies. The design and interpretation of many studies here depends critically on Linkage disequilibrium (LD). For example in association studies, LD determines the density of Single Nucleotide Polymorphisms (SNPs) to be genotyped to represent the majority of the genomic information. Here, we aim to determine LD in wild An. gambiae s.l. populations in 4 genes potentially involved in mosquito immune responses against pathogens (Gambicin, NOS, REL2 and FBN9) using previously published and newly generated sequences.ResultsThe level of LD between SNP pairs in cloned sequences of each gene was determined for 7 species (or incipient species) of the An. gambiae complex. In all tested genes and species, LD between SNPs was low: even at short distances (< 200 bp), most SNP pairs gave an r2 < 0.3. Mean r2 ranged from 0.073 to 0.766. In most genes and species LD decayed very rapidly with increasing inter-marker distance.ConclusionsThese results are of great interest for the development of large scale polymorphism studies, as LD generally falls below any useful limit. It indicates that very fine scale SNP detection will be required to give an overall view of genome-wide polymorphism. Perhaps a more feasible approach to genome wide association studies is to use targeted approaches using candidate gene selection to detect association to phenotypes of interest.

Pigeonpea genomics initiative (PGI): an international effort to improve crop productivity of pigeonpea (Cajanus cajan L.)

Pigeonpea (Cajanus cajan), an important food legume crop in the semi-arid regions of the world and the second most important pulse crop in India, has an average crop productivity of 780 kg/ha. The relatively low crop yields may be attributed to non-availability of improved cultivars, poor crop husbandry and exposure to a number of biotic and abiotic stresses in pigeonpea growing regions. Narrow genetic diversity in cultivated germplasm has further hampered the effective utilization of conventional breeding as well as development and utilization of genomic tools, resulting in pigeonpea being often referred to as an ‘orphan crop legume’. To enable genomics-assisted breeding in this crop, the pigeonpea genomics initiative (PGI) was initiated in late 2006 with funding from Indian Council of Agricultural Research under the umbrella of Indo-US agricultural knowledge initiative, which was further expanded with financial support from the US National Science Foundation’s Plant Genome Research Program and the Generation Challenge Program. As a result of the PGI, the last 3 years have witnessed significant progress in development of both genetic as well as genomic resources in this crop through effective collaborations and coordination of genomics activities across several institutes and countries. For instance, 25 mapping populations segregating for a number of biotic and abiotic stresses have been developed or are under development. An 11X-genome coverage bacterial artificial chromosome (BAC) library comprising of 69,120 clones have been developed of which 50,000 clones were end sequenced to generate 87,590 BAC-end sequences (BESs). About 10,000 expressed sequence tags (ESTs) from Sanger sequencing and ca. 2 million short ESTs by 454/FLX sequencing have been generated. A variety of molecular markers have been developed from BESs, microsatellite or simple sequence repeat (SSR)-enriched libraries and mining of ESTs and genomic amplicon sequencing. Of about 21,000 SSRs identified, 6,698 SSRs are under analysis along with 670 orthologous genes using a GoldenGate SNP (single nucleotide polymorphism) genotyping platform, with large scale SNP discovery using Solexa, a next generation sequencing technology, is in progress. Similarly a diversity array technology array comprising of ca. 15,000 features has been developed. In addition, >600 unique nucleotide binding site (NBS) domain containing members of the NBS-leucine rich repeat disease resistance homologs were cloned in pigeonpea; 960 BACs containing these sequences were identified by filter hybridization, BES physical maps developed using high information content fingerprinting. To enrich the genomic resources further, sequenced soybean genome is being analyzed to establish the anchor points between pigeonpea and soybean genomes. In addition, Solexa sequencing is being used to explore the feasibility of generating whole genome sequence. In summary, the collaborative efforts of several research groups under the umbrella of PGI are making significant progress in improving molecular tools in pigeonpea and should significantly benefit pigeonpea genetics and breeding. As these efforts come to fruition, and expanded (depending on funding), pigeonpea would move from an ‘orphan legume crop’ to one where genomics-assisted breeding approaches for a sustainable crop improvement are routine.

SNP typing using molecular inversion probes

Single nucleotide polymorphism (SNP) typing will be useful in forensic analysis but is limited by the multiplexing capacity of multiple targets in the PCR. A strategy to circumvent this problem can be to use molecular inversion probes (MIPs). MIPs are linear oligonucleotides 70–100 nt in length, with end-segments that are complementary to a target sequence. In this study, 10 SNPs were selected for SNP detection performed by pyrosequencing. The high-multiplexing capacity of MIPs makes this technology very useful for large scale SNP genotyping, but may also be useful for DNA typing in forensic investigations.

경제동물 유전체학 연구의 최근 연구 동향

경제 동물의 유전체 연구는 최근에 급속하게 발전하여 초기의 유전체 지도로 부터 유전자의 발견에 필수적인 앙적/질적 형질 유전자를 확인 동정가능한 수준의 지도가 개발되었다. 이러한 발전은 경제동물의 전체 게놈 염기서열 결정과 대량 ESTs의 개발에 의해 가능해졌다. 특히 염시서열 결정은 경제형질과 연관된 대규모의 SNPs 개발에 의한 QTL 연구에 유용한 정보를 제공할 것으로 사료된다. 비교 유전체 연구를 통해 인간 및 설치류 모델동물에서 나온 유전체 정보를 이용하여 경제 동물의 유전체 연구에 있어 중요한 발견을 이루었다. 이러한 노력은 좀더 밀도 높은 QTL지도의 작성을 가능하게 하여 쉽게 측정하기 어려운 경제형질과 연관된 유전자의 확인 및 동정을 가능하게 하고 궁극적으로 산업체에서 이용 가능한 표지인자의 개발을 가능하게 할 것으로 사료된다. 이와 더불어, 경제동물 유전체 연구 성과는 인간의 생리현상의 유전체 측면의 이해를 더욱 증진시킬 것이다. Genome research in economic animals has progressed rapidly in recent years, transforming from primitive genome maps to quantitative/qualitative trait maps that are indispensable to gene discovery. These advances have been benefited from the result of animal genome sequencing projects and functional genomics that are being extensively applied in livestock animal research following the development of large expressed sequences tags (ESTs). Genome sequencing efforts will provide information to QTL study by larger scale single nucleotide polymorphisms (SNPs) association study. Comparative genomics which is applying the information from human genome research as well as rodents model has contributed to important discoveries in economic animal genome research. These efforts will speed up much denser QTL maps development for phenotypic traits which are not easy to measure and to be identified by quantitative genetics [20] and lead to development of convincing markers associated with economically important trait, which will be eventually applied to livestock industry. In addition to practical application, animal genome research will enrich the understanding of human physiology in terms of genome biology.

Advances in QTL Mapping in Pigs

Over the past 15 years advances in the porcine genetic linkage map and discovery of useful candidate genes have led to valuable gene and trait information being discovered. Early use of exotic breed crosses and now commercial breed crosses for quantitative trait loci (QTL) scans and candidate gene analyses have led to 110 publications which have identified 1,675 QTL. Additionally, these studies continue to identify genes associated with economically important traits such as growth rate, leanness, feed intake, meat quality, litter size, and disease resistance. A well developed QTL database called PigQTLdb is now as a valuable tool for summarizing and pinpointing in silico regions of interest to researchers. The commercial pig industry is actively incorporating these markers in marker-assisted selection along with traditional performance information to improve traits of economic performance. The long awaited sequencing efforts are also now beginning to provide sequence available for both comparative genomics and large scale single nucleotide polymorphism (SNP) association studies. While these advances are all positive, development of useful new trait families and measurement of new or underlying traits still limits future discoveries. A review of these developments is presented.

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Identification of single nucleotide polymorphisms (SNPs) associated with adverse effects of gefinitib

3020 Background: Effectiveness of Gefinitib on non small cell lung carcinoma has been well demonstrated. However, some lethal adverse effects on Gefinitib treatment were observed, as well. Therefore, methods for prediction of adverse effects have to be established. In order to predict adverse effects of Gefinitib treatment, we conducted a large scale SNP typing analysis. Methods: 41 patients with non small cell lung carcinoma were enrolled in this study. All patients were orally administered one tablet of Gefinitib (250 mg) every day at least for 16 weeks. Adverse effects were evaluated every seven days and graded according to NCI-CTC grading. In addition, concentration of Gefinitib in peripheral blood was measured at 28th day of the treatment. DNA was isolated form peripheral blood. To identify SNPs associated with adverse effect, genotypes of 2,727 SNPs over 298 genes were determined by Invader assay. Association between SNPs and adverse effects was examined by Fisher's exact test. Results: During the course of treatment, 23 out of 41 patients exhibited diarrhea as an adverse effect. Genotyping analysis showed strong association between diarrhea and seven SNPs (p < 0.000067, Odds ratio: 20.8) located within a gene (GENE1). Among those, three SNPs were located putative regulatory region of GENE1 and rest were within intronic region. Haplotype analysis showed those SNPs belonged to one of three major haplotypes observed at the locus, suggesting a specific haplotype may harbor genetic alteration responsible for developing diarrhea. In addition, our analysis revealed that two SNPs on a gene (GENE2) were strongly associated with blood concentration of Gefinitib (p< 0.00015 Odds ratio: 20.2). Conclusions: By using those SNPs, appearance of diarrhea on Gefinitib treatment will be precisely predicted. As both genes belonged to same gene family, our result suggested this gene family may play roles in pharmacokinetics and/or pharmacodynamics of Gefinitib. No significant financial relationships to disclose.

Identification of single nucleotide polymorphisms (SNPs) associated with adverse effects of gefinitib

3020 Background: Effectiveness of Gefinitib on non small cell lung carcinoma has been well demonstrated. However, some lethal adverse effects on Gefinitib treatment were observed, as well. Therefore, methods for prediction of adverse effects have to be established. In order to predict adverse effects of Gefinitib treatment, we conducted a large scale SNP typing analysis. Methods: 41 patients with non small cell lung carcinoma were enrolled in this study. All patients were orally administered one tablet of Gefinitib (250 mg) every day at least for 16 weeks. Adverse effects were evaluated every seven days and graded according to NCI-CTC grading. In addition, concentration of Gefinitib in peripheral blood was measured at 28th day of the treatment. DNA was isolated form peripheral blood. To identify SNPs associated with adverse effect, genotypes of 2,727 SNPs over 298 genes were determined by Invader assay. Association between SNPs and adverse effects was examined by Fisher's exact test. Results: During the c...

High-throughput multiplex SNP genotyping with MALDI-TOF mass spectrometry: practice, problems and promise.

Single nucleotide polymorphisms (SNPs) are currently being identified and mapped at a remarkable pace, providing a rich genetic resource with vast potential for disease gene discovery, pharmacogenetics, and understanding the origins of modern humans. High-throughput, cost effective genotyping methods are essential in order to make the most advantageous and immediate use of these SNP data. We have incorporated the use of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) in our laboratory as a tool for differentiating genotypes based on the mass of the variant DNA sequence, and have utilized this method for production scale SNP genotyping. We have combined a 4 microl PCR amplification reaction using 3 ng of genomic DNA with a secondary enzymatic reaction (mini-sequencing) containing oligonucleotide primers that anneal immediately upstream of the polymorphic site, dideoxynucleotides, and a thermostable polymerase used to extend the PCR product by a single base pair. Mass spectrometry (MS) analysis of mini-sequencing reactions was performed using a MALDI-TOF instrument (Voyager-DE, Perseptive Biosystems, Framingham, MA). We performed both single and multiplex PCR and mini-sequencing reactions, and genotyped seven different variant sites in a random sample of 989 individuals. Genotypes generated with MS methods were compared with genotypes produced using a 5' exonuclease fluorescence-based assay (Taqman, Applied Biosystems, Foster City, CA) and a gel-based genotyping protocol. Because multiple polymorphisms can be detected in a single reaction, the MS technique provides a cost-effective and efficient method for high-throughput genotyping.