SNP2RFLP: a computational tool to facilitate genetic mapping using benchtop analysis of SNPs

Genome-wide analysis of single nucleotide polymorphism (SNP) markers is an extremely efficient means for genetic mapping of mutations or traits in mice. However, this approach often defines a relatively large recombinant interval. To facilitate the refinement of this interval, we developed the program SNP2RFLP. This program can be used to identify region-specific SNPs in which the polymorphic nucleotide creates a restriction fragment length polymorphism (RFLP) that can be readily assayed at the benchtop using restriction enzyme digestion of SNP-containing PCR products. The program permits user-defined queries that maximize the informative markers for a particular application. This facilitates fine-mapping in a region containing a mutation of interest, which should prove valuable to the mouse genetics community. SNP2RFLP and further details are publicly available at http://genetics.bwh.harvard.edu/snp2rflp/.     

HDAPD: a web tool for searching the disease-associated protein structures

Background

PCR-restriction fragment length polymorphism (RFLP) assay is a cost-effective method for SNP genotyping and mutation detection, but the manual mining for restriction enzyme sites is challenging and cumbersome. Three years after we constructed SNP-RFLPing, a freely accessible database and analysis tool for restriction enzyme mining of SNPs, significant improvements over the 2006 version have been made and incorporated into the latest version, SNP-RFLPing 2.

Results

The primary aim of SNP-RFLPing 2 is to provide comprehensive PCR-RFLP information with multiple functionality about SNPs, such as SNP retrieval to multiple species, different polymorphism types (bi-allelic, tri-allelic, tetra-allelic or indels), gene-centric searching, HapMap tagSNPs, gene ontology-based searching, miRNAs, and SNP500Cancer. The RFLP restriction enzymes and the corresponding PCR primers for the natural and mutagenic types of each SNP are simultaneously analyzed. All the RFLP restriction enzyme prices are also provided to aid selection. Furthermore, the previously encountered updating problems for most SNP related databases are resolved by an on-line retrieval system.

Conclusions

The user interfaces for functional SNP analyses have been substantially improved and integrated. SNP-RFLPing 2 offers a new and user-friendly interface for RFLP genotyping that can be used in association studies and is freely available at http://bio.kuas.edu.tw/snp-rflping2.     

MonkeySNP: a web portal for non-human primate single nucleotide polymorphisms

MonkeySNP is a web-based resource created by the Genetic Resource and Informatics Program at the Oregon National Primate Research Center to facilitate access to non-human primate (NHP) single nucleotide polymorphisms (SNP) data. MonkeySNP is a mirror of the NCBI dbSNP database and contains additional NHP subpopulation genotype data and visual genotype displays to support SNP review and selection. AVAILABILITY: http://monkeysnp.ohsu.edu/snp/ SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.     

The SNP Consortium website: past,present and future

The SNP Consortium website (http://snp.cshl.org) has undergone many changes since its initial conception three years ago. The database back end has been changed from the venerable ACeDB to the more scalable MySQL engine. Users can access the data via gene or single nucleotide polymorphism (SNP) keyword searches and browse or dump SNP data to textfiles. A graphical genome browsing interface shows SNPs mapped onto the genome assembly in the context of externally available gene predictions and other features. SNP allele frequency and genotype data are available via FTP-download and on individual SNP report web pages. SNP linkage maps are available for download and for browsing in a comparative map viewer. All software components of the data coordinating center (DCC) website (http://snp.cshl.org) are open source.     

dChipSNP: significance curve and clustering of SNP-array-based loss-of-heterozygosity data

MOTIVATION: Oligonucleotide microarrays allow genotyping of thousands of single-nucleotide polymorphisms (SNPs) in parallel. Recently, this technology has been applied to loss-of-heterozygosity (LOH) analysis of paired normal and tumor samples. However, methods and software for analyzing such data are not fully developed. RESULT: Here, we report automated methods for pooling SNP array replicates to make LOH calls, visualizing SNP and LOH data along chromosomes in the context of genes and cytobands, making statistical inference to identify shared LOH regions, clustering samples based on LOH profiles and correlating the clustering results to clinical variables. Application of these methods to prostate and breast cancer datasets generates biologically important results. AVAILABILITY: The software module dChipSNP implementing these methods is available at http://biosun1.harvard.edu/complab/dchip/snp/ SUPPLEMENTARY INFORMATION: The breast cancer data are provided by Andrea L. Richardson, Zhigang C. Wang and James D. Iglehart.     

In Silico Analysis of the Restriction Fragment Length Distribution in the Human Genome

The Restriction On Computer (ROC) program (freely available at http://www.mcb.harvard.edu/ gilbert/ROC) was developed and used to analyze the restriction fragment length distribution in the human genome. In contrast to other programs searching for restriction sites, ROC simultaneously analyzes several long nucleotide sequences, such as the entire genomes, and in essence simulates electrophoretic analysis of DNA restriction fragments. In addition, this program extracts and analyzes DNA repeats that account for peaks in the restriction fragment length distribution. The ROC analysis data are consistent with the experimental data obtained via in vitro restriction enzyme analysis (DNA taxonoprint). A difference between the in vitro and in silico results is explained by underrepresentation of tandem DNA repeats in genomic databases. The ROC analysis of individual genome fragments elucidated the nature of several DNA markers, which were earlier revealed by DNA taxonoprint, and showed that L1 and Alurepeats are nonrandomly distributed in various chromosomes. Another advantage is that the ROC procedure makes it possible to analyze the nonrandom character of a genomic distribution of short DNA sequences. The ROC analysis showed that a low poly(G) frequency is characteristic of the entire human genome, rather than of only coding sequences. The method was proposed for a more complex in silico analysis of the genome. For instance, it is possible to simulate DNA restriction together with blot hybridization and then to analyze the nature of markers revealed.     

WHAP: haplotype-based association analysis

We describe a software tool to perform haplotype-based association analysis, for quantitative and qualitative traits, in population and family samples, using single nucleotide polymorphism or multiallelic marker data. A range of tests is offered: omnibus and haplotype-specific tests; prospective and retrospective likelihoods; covariates and moderators; sliding window analyses; permutation P-values. We focus on the ability to flexibly impose constraints on haplotype effects, which allows for a range of conditional haplotype-based likelihood ratio tests: for example, whether an allele has an effect independent of its haplotypic background, or whether a single variant can explain the overall association at a locus. We illustrate using these tests to dissect a multi-locus association. AVAILABILITY: WHAP is a C/C++ program, freely available from the author's website: http://pngu.mgh.harvard.edu/purcell/whap/     

Mutagenic primer design for mismatch PCR-RFLP SNP genotyping using a genetic algorithm

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) is useful in small-scale basic research studies of complex genetic diseases that are associated with single nucleotide polymorphism (SNP). Designing a feasible primer pair is an important work before performing PCR-RFLP for SNP genotyping. However, in many cases, restriction enzymes to discriminate the target SNP resulting in the primer design is not applicable. A mutagenic primer is introduced to solve this problem. GA-based Mismatch PCR-RFLP Primers Design (GAMPD) provides a method that uses a genetic algorithm to search for optimal mutagenic primers and available restriction enzymes from REBASE. In order to improve the efficiency of the proposed method, a mutagenic matrix is employed to judge whether a hypothetical mutagenic primer can discriminate the target SNP by digestion with available restriction enzymes. The available restriction enzymes for the target SNP are mined by the updated core of SNP-RFLPing. GAMPD has been used to simulate the SNPs in the human SLC6A4 gene under different parameter settings and compared with SNP Cutter for mismatch PCR-RFLP primer design. The in silico simulation of the proposed GAMPD program showed that it designs mismatch PCR-RFLP primers. The GAMPD program is implemented in JAVA and is freely available at http://bio.kuas.edu.tw/gampd/.     

SNP Discovery and Linkage Map Construction in Cultivated Tomato

Few intraspecific genetic linkage maps have been reported for cultivated tomato, mainly because genetic diversity within Solanum lycopersicum is much less than that between tomato species. Single nucleotide polymorphisms (SNPs), the most abundant source of genomic variation, are the most promising source of polymorphisms for the construction of linkage maps for closely related intraspecific lines. In this study, we developed SNP markers based on expressed sequence tags for the construction of intraspecific linkage maps in tomato. Out of the 5607 SNP positions detected through in silico analysis, 1536 were selected for high-throughput genotyping of two mapping populations derived from crosses between ‘Micro-Tom’ and either ‘Ailsa Craig’ or ‘M82’. A total of 1137 markers, including 793 out of the 1338 successfully genotyped SNPs, along with 344 simple sequence repeat and intronic polymorphism markers, were mapped onto two linkage maps, which covered 1467.8 and 1422.7 cM, respectively. The SNP markers developed were then screened against cultivated tomato lines in order to estimate the transferability of these SNPs to other breeding materials. The molecular markers and linkage maps represent a milestone in the genomics and genetics, and are the first step toward molecular breeding of cultivated tomato. Information on the DNA markers, linkage maps, and SNP genotypes for these tomato lines is available at http://www.kazusa.or.jp/tomato/.     

Using AFLP markers and the Geneland program for the inference of population genetic structure

The use of dominant markers such as amplified fragment length polymorphism (AFLP) for population genetics analyses is often impeded by the lack of appropriate computer programs and rarely motivated by objective considerations. The point of the present note is twofold: (i) we describe how the computer program Geneland designed to infer population structure has been adapted to deal with dominant markers; and (ii) we use Geneland for numerical comparison of dominant and codominant markers to perform clustering. AFLP markers lead to less accurate results than bi-allelic codominant markers such as single nucleotide polymorphisms (SNP) markers but this difference becomes negligible for data sets of common size (number of individuals n≥100, number of markers L≥200). The latest Geneland version (3.2.1) handling dominant markers is freely available as an R package with a fully clickable graphical interface. Installation instructions and documentation can be found on http://www2.imm.dtu.dk/~gigu/Geneland.     

Genetics web sites

Human Genome Project Information http://www.ornl.gov/TechResources/Human_Genome/home.html

Access Excellence http://www.accessexcellence.org/

Genetics Science Learning Centre at the Eccles Institute of Human Genetics, University of Utah http://gslc.genetics.utah.edu/

Blazing a Genetic Trial http://www.hhmi.org/GeneticTrail/

A Hypermedia Glossary of Genetic Terms prepared and presented by Birgid Schlindwein http://www.weihenstephan.de/~schlind/genglos.html

Virtual Flylab http://vcourseware5.calstatela.edu/VirtualFlyLab/IntroVflyLab.html

Web watch topics coming up…     

The Synergizer service for translating gene, protein and other biological identifiers

The Synergizer is a database and web service that provides translations of biological database identifiers. It is accessible both programmatically and interactively. AVAILABILITY: The Synergizer is freely available to all users inter-actively via a web application (http://llama.med.harvard.edu/synergizer/translate) and programmatically via a web service. Clients implementing the Synergizer application programming interface (API) are also freely available. Please visit http://llama.med.harvard.edu/synergizer/doc for details.     

HaploBuild: an algorithm to construct non-contiguous associated haplotypes in family based genetic studies

We have created a program that searches densely genotyped regions for associated non-contiguous haplotypes using a standard family based haplotype association test. This program was designed to expand upon the 'sliding window' methodologies commonly used for haplotype construction by allowing the association of subsets of single nucleotide polymorphisms (SNPs) to drive the construction of the haplotype. This strategy permits HaploBuild to construct more biologically relevant haplotypes that are not constrained by arbitrary length and contiguous orientation. Availability: http://snp.bumc.bu.edu.     

In silico analysis of the restriction fragments length distribution in the human genome.]

The Restriction On Computer (ROC) program (freely available at http://www.mcb.harvard.edu/gilbert/ROC) was developed and used to analyze the restriction fragment length distribution in the human genome. In contrast to other programs searching for restriction sites, ROC simultaneously analyzes several long nucleotide sequences, such as the entire genomes, and in essence simulates electrophoretic analysis of DNA restriction fragments. In addition, this program extracts and analyzes DNA repeats that account for peaks in the restriction fragment length distribution. The ROC analysis data are consistent with the experimental data obtained via in vitro restriction enzyme analysis (taxonomic printing). A difference between the in vitro and in silico results is explained by underrepresentation of tandem DNA repeats in genomic databases. The ROC analysis of individual genome fragments elucidated the nature of several DNA markers, which were earlier revealed by taxonomic printing, and showed that L1 and Alu repeats are nonrandomly distributed in various chromosomes. Another advantage is that the ROC procedure makes it possible to analyze the nonrandom character of a genomic distribution of short DNA sequences. The ROC analysis showed that a low poly(G) frequency is characteristic of the entire human genome, rather than of only coding sequences. The method was proposed for a more complex in silico analysis of the genome. For instance, it is possible to simulate DNA restriction together with blot hybridization and then to analyze the nature of markers revealed.     

SNPP: automating large-scale SNP genotype data management

To manage high-throughput single nucleotide polymorphism (SNP) genotyping data efficiently, we developed a dynamic general database management system-SNPP (SNP Processor). It provides several functions, including data importing with comparison, Mendelian inheritance check within pedigrees, data compiling and exporting. Furthermore, SNPP may generate files for repeat genotyping and transform them into files that can be executed by a liquid handling system. AVAILABILITY: http://orclinux.creighton.edu/snpp/ CONTACT: lanjuanzhao@creighton.edu     

单核苷酸多态性技术在鸡遗传变异中的研究及应用

单核苷酸多态性(SNP)是继限制性片段长度多态性、微卫星标记之后的第三代分子标记,通常呈双等位基因多态。本文从SNP的特点、SNP的发现与检测、SNP数据库、SNP的频率及其与表型的关系等方面简述了SNP在鸡遗传变异中的研究及应用进展。     

snp.plotter: an R-based SNP/haplotype association and linkage disequilibrium plotting package

snp.plotter is a newly developed R package which produces high-quality plots of results from genetic association studies. The main features of the package include options to display a linkage disequilibrium (LD) plot below the P-value plot using either the r2 or D' LD metric, to set the X-axis to equal spacing or to use the physical map of markers, and to specify plot labels, colors, symbols and LD heatmap color scheme. snp.plotter can plot single SNP and/or haplotype data and simultaneously plot multiple sets of results. R is a free software environment for statistical computing and graphics available for most platforms. The proposed package provides a simple way to convey both association and LD information in a single appealing graphic for genetic association studies. AVAILABILITY: Downloadable R package and example datasets are available at http://cbdb.nimh.nih.gov/~kristin/snp.plotter.html and http://www.r-project.org.