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Detection of Rare Genomic Variants from Pooled Sequencing Using SPLINTER
Published on: June 23, 2012
Detection of rare genomic variants from pooled sequencing using SPLINTER.
Francesco Vallania1, Enrique Ramos, Sharon Cresci
1Center for Genome Sciences and Systems Biology, Department of Genetics, Washington University School of Medicine.
Journal of Visualized Experiments : Jove
|July 5, 2012
Summary
Researchers developed a pooled sequencing method and SPLINTER software for accurate rare DNA variant detection. This approach offers cost and time savings for analyzing genetic variation in large populations, aiding common disease research.
Area of Science:
- Genetics
- Bioinformatics
Background:
- Advancements in DNA sequencing reveal greater genetic variation between individuals than previously understood.
- Array-based genotyping has limitations in identifying common sequence variants contributing to complex disease phenotypes.
- The Common Disease / Rare Variant hypothesis posits that rare DNA variants explain significant "missing heritability" in complex traits.
Purpose of the Study:
- To develop a cost-effective and efficient method for analyzing rare genetic variants in large populations.
- To introduce a novel software package for highly accurate rare variant detection from pooled sequencing data.
- To enable population-based surveys of genomic loci for rare variant analysis, overcoming previous limitations.
Main Methods:
- Developed a pooled sequencing approach to analyze genomes from entire populations in a single sequencing library.
- Created the SPLINTER software package for accurate detection of insertions, deletions, and substitutions (up to 4 base pairs).
- Utilized an internal variant calling control strategy with mean sequencing coverage of 25-fold per allele for pooled samples.
Main Results:
- Demonstrated high sensitivity and specificity for detecting rare variants (down to 1 in 500 individuals) using SPLINTER.
- Showcased excellent concordance between pooled sequencing and genome-wide array genotyping for over 20kb of sequencing per person in 947 individuals.
- Validated the scalability of the pooled sequencing method for any number of loci and individuals.
Conclusions:
- The pooled sequencing approach and SPLINTER software provide a powerful, cost-effective solution for rare variant detection.
- This methodology facilitates the investigation of the Common Disease / Rare Variant hypothesis by enabling large-scale genetic variation analysis.
- The strategy is adaptable for various sequencing applications, including hybridization capture and analysis of heterogeneous samples like tumor DNA.
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