Sequence Variation In Populations Research Articles

Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster

The sequencing of pooled non-barcoded individuals is an inexpensive and efficient means of assessing genome-wide population allele frequencies, yet its accuracy has not been thoroughly tested. We assessed the accuracy of this approach on whole, complex eukaryotic genomes by resequencing pools of largely isogenic, individually sequenced Drosophila melanogaster strains. We called SNPs in the pooled data and estimated false positive and false negative rates using the SNPs called in individual strain as a reference. We also estimated allele frequency of the SNPs using “pooled” data and compared them with “true” frequencies taken from the estimates in the individual strains. We demonstrate that pooled sequencing provides a faithful estimate of population allele frequency with the error well approximated by binomial sampling, and is a reliable means of novel SNP discovery with low false positive rates. However, a sufficient number of strains should be used in the pooling because variation in the amount of DNA derived from individual strains is a substantial source of noise when the number of pooled strains is low. Our results and analysis confirm that pooled sequencing is a very powerful and cost-effective technique for assessing of patterns of sequence variation in populations on genome-wide scales, and is applicable to any dataset where sequencing individuals or individual cells is impossible, difficult, time consuming, or expensive.

ChemInform Abstract: The Human Genome Project - An Overview

The human genome sequence will underpin human biology and medicine in the next century, providing a single, essential reference to all genetic information. The international program to determine the complete DNA sequence (3,000 million bases) is well underway. As of January 2000, 50% of the sequence is available in the public domain. A comprehensive working draft is expected this year, and the entire sequence is projected to be finished in 2003. DNA sequencing is carried out on mapped, overlapping bacterial clones of 150-200 kb. The working draft comprises assembled unfinished sequence and is released immediately in the public domain. The draft sequence of each clone is then completed, by closing any remaining gaps and resolving any ambiguities, before the entire sequence is checked, annotated, and submitted to the public databases. The sequence of each clone is finished to an accuracy of >99.99%. The availability of a reference sequence of the genome provides the basis for studying the nature of sequence variation, particularly single nucleotide polymorphisms (SNPs), in human populations. SNP typing is a powerful tool for genetic analysis, and will enable us to uncover the association of loci at specific sites in the genome with many disease traits. SNPs occur at a frequency of approximately 1 SNP/kb throughout the genome when the sequence of any two individuals is compared. Programs to detect and map SNPs in the human genome are underway with the aim of establishing a SNP map of the genome during the next two years. The human genome sequence will provide a complete description of all the genes. Annotation of the sequence with the gene structures is achieved by a combination of computational analysis (predictive and homology-based) and experimental confirmation by cDNA sequencing. Detecting homologies between newly defined gene products and proteins of known function helps to postulate biochemical functions for them, which can then be tested. Establishing the association of specific genes with disease phenotypes by mutation screening, particularly for monogenic disorders, provides further assistance in defining the functions of some gene products, as well as helping to establish the cause of the disease. As our knowledge of gene sequences and sequence variation in populations increases, we will pinpoint more and more of the genes and proteins that are important in common, complex diseases. A more detailed understanding of the function of the human genome will be achieved as we identify sequences that control gene expression. Given the availability of gene sequences, the expression status of genes in particular tissues can be monitored in parallel. By comparing corresponding genomic sequences in different species (for example: man, mouse, chicken, and zebrafish), regions that have been highly conserved during evolution can be identified, many of which reflect conserved functions such as gene regulation. These approaches promise to greatly accelerate our interpretation of the human genome sequence.

Sequence diversity, reproductive isolation and species concepts in Saccharomyces.

Using the biological species definition, yeasts of the genus Saccharomyces sensu stricto comprise six species and one natural hybrid. Previous work has shown that reproductive isolation between the species is due primarily to sequence divergence acted upon by the mismatch repair system and not due to major gene differences or chromosomal rearrangements. Sequence divergence through mismatch repair has also been shown to cause partial reproductive isolation among populations within a species. We have surveyed sequence variation in populations of Saccharomyces sensu stricto yeasts and measured meiotic sterility in hybrids. This allows us to determine the divergence necessary to produce the reproductive isolation seen among species. Rather than a sharp transition from fertility to sterility, which may have been expected, we find a smooth monotonic relationship between diversity and reproductive isolation, even as far as the well-accepted designations of S. paradoxus and S. cerevisiae as distinct species. Furthermore, we show that one species of Saccharomyces--S. cariocanus--differs from a population of S. paradoxus by four translocations, but not by sequence. There is molecular evidence of recent introgression from S. cerevisiae into the European population of S. paradoxus, supporting the idea that in nature the boundary between these species is fuzzy.

The Human Genome Project?An Overview

The human genome sequence will underpin human biology and medicine in the next century, providing a single, essential reference to all genetic information. The international program to determine the complete DNA sequence (3,000 million bases) is well underway. As of January 2000, 50% of the sequence is available in the public domain. A comprehensive working draft is expected this year, and the entire sequence is projected to be finished in 2003. DNA sequencing is carried out on mapped, overlapping bacterial clones of 150-200 kb. The working draft comprises assembled unfinished sequence and is released immediately in the public domain. The draft sequence of each clone is then completed, by closing any remaining gaps and resolving any ambiguities, before the entire sequence is checked, annotated, and submitted to the public databases. The sequence of each clone is finished to an accuracy of >99.99%. The availability of a reference sequence of the genome provides the basis for studying the nature of sequence variation, particularly single nucleotide polymorphisms (SNPs), in human populations. SNP typing is a powerful tool for genetic analysis, and will enable us to uncover the association of loci at specific sites in the genome with many disease traits. SNPs occur at a frequency of approximately 1 SNP/kb throughout the genome when the sequence of any two individuals is compared. Programs to detect and map SNPs in the human genome are underway with the aim of establishing a SNP map of the genome during the next two years. The human genome sequence will provide a complete description of all the genes. Annotation of the sequence with the gene structures is achieved by a combination of computational analysis (predictive and homology-based) and experimental confirmation by cDNA sequencing. Detecting homologies between newly defined gene products and proteins of known function helps to postulate biochemical functions for them, which can then be tested. Establishing the association of specific genes with disease phenotypes by mutation screening, particularly for monogenic disorders, provides further assistance in defining the functions of some gene products, as well as helping to establish the cause of the disease. As our knowledge of gene sequences and sequence variation in populations increases, we will pinpoint more and more of the genes and proteins that are important in common, complex diseases. A more detailed understanding of the function of the human genome will be achieved as we identify sequences that control gene expression. Given the availability of gene sequences, the expression status of genes in particular tissues can be monitored in parallel. By comparing corresponding genomic sequences in different species (for example: man, mouse, chicken, and zebrafish), regions that have been highly conserved during evolution can be identified, many of which reflect conserved functions such as gene regulation. These approaches promise to greatly accelerate our interpretation of the human genome sequence.

The Human Genome Project--an overview.

The human genome sequence will underpin human biology and medicine in the next century, providing a single, essential reference to all genetic information. The international program to determine the complete DNA sequence (3,000 million bases) is well underway. As of January 2000, 50% of the sequence is available in the public domain. A comprehensive working draft is expected this year, and the entire sequence is projected to be finished in 2003. DNA sequencing is carried out on mapped, overlapping bacterial clones of 150-200 kb. The working draft comprises assembled unfinished sequence and is released immediately in the public domain. The draft sequence of each clone is then completed, by closing any remaining gaps and resolving any ambiguities, before the entire sequence is checked, annotated, and submitted to the public databases. The sequence of each clone is finished to an accuracy of >99.99%. The availability of a reference sequence of the genome provides the basis for studying the nature of sequence variation, particularly single nucleotide polymorphisms (SNPs), in human populations. SNP typing is a powerful tool for genetic analysis, and will enable us to uncover the association of loci at specific sites in the genome with many disease traits. SNPs occur at a frequency of approximately 1 SNP/kb throughout the genome when the sequence of any two individuals is compared. Programs to detect and map SNPs in the human genome are underway with the aim of establishing a SNP map of the genome during the next two years. The human genome sequence will provide a complete description of all the genes. Annotation of the sequence with the gene structures is achieved by a combination of computational analysis (predictive and homology-based) and experimental confirmation by cDNA sequencing. Detecting homologies between newly defined gene products and proteins of known function helps to postulate biochemical functions for them, which can then be tested. Establishing the association of specific genes with disease phenotypes by mutation screening, particularly for monogenic disorders, provides further assistance in defining the functions of some gene products, as well as helping to establish the cause of the disease. As our knowledge of gene sequences and sequence variation in populations increases, we will pinpoint more and more of the genes and proteins that are important in common, complex diseases. A more detailed understanding of the function of the human genome will be achieved as we identify sequences that control gene expression. Given the availability of gene sequences, the expression status of genes in particular tissues can be monitored in parallel. By comparing corresponding genomic sequences in different species (for example: man, mouse, chicken, and zebrafish), regions that have been highly conserved during evolution can be identified, many of which reflect conserved functions such as gene regulation. These approaches promise to greatly accelerate our interpretation of the human genome sequence.

Denaturing gradient gel electrophoresis (DGGE): a rapid and sensitive technique to screen nucleotide sequence variation in populations.

We describe a rapid and sensitive method for the detection of nucleotide sequence variation that can be used for large-scale screening of population markers. Denaturing gradient gel electrophoresis (DGGE) detects sequence variants of amplified fragments by the differences in their melting behavior. DGGE detects most single-base substitutions when carried out on products amplified with a primer to which a GC clamp has been added. Although DGGE has been primarily used for the detection of limited numbers of single-base mutations in disease studies, it offers great potential for use in population analysis of genetic markers with greater levels of sequence variation. The methodology described was developed to identify the number and distribution of MHC class I alpha 1 alleles among chinook salmon (Oncorhynchus tshawytscha) populations. DGGE detects 28 of 31 identified alpha 1 sequences, which differ by between 1 and 16 nucleotides and a two-codon indel. By creating a network of control alleles, 22-23 of the MHC alleles can be resolved rapidly and accurately by a single gel run condition, and 27 alleles can be resolved by two gel run conditions. This techniques has been used in surveys scoring alleles from two MHC markers (class I alpha 1 and alpha 2) in 20,000 individuals of chinook and coho (O. kisutch) salmon. A single person in our laboratory now analyzes 160 salmon from one MHC locus per day with DGGE.