Identification of rare genetic variants
Next-generation sequencing is used for the identification of rare allelic variants in pooled DNAs.
IGA has borrowed this approach from cancer research and optimized the procedure for the quest for rare variants in plant populations. DNA targets are amplified from several individuals, and DNA barcoding retains the individual information in pooled sequencing. The following article guides you for designing your own experiment through all steps: pooling strategy, target enrichment, multiplexing, sequencing depth and appropriate bioinformatics.
Marroni et al (2012) The quest for rare variants: pooled multiplexed next generation sequencing in plants. Frontiers in Plant Science doi: 10.3389/fpls.2012.00133. eCollection 2012.
Variants that control complex traits are used in plant breeding for crop improvement. An illustrative example comes from black poplar, an important feedstock for the production of biofuels. Allelic variants of lignin biosynthetic genes in natural populations of poplar revealed trees with altered lignin composition, thus removing a major limiting factor in the conversion of plant biomass to biofuels.
Marroni et al (2011) Large-scale detection of rare variants via pooled multiplexed next-generation sequencing: towards next-generation Ecotilling. Plant Journal 67:736-745
Pooled NGS is also applied to study demography and selection in natural populations of Norway spruce. IGA has compared the European populations of Norway spruce that evolved northerly in the Fennoscandian region and southerly in the Alpine mountain range. We investigated DNA sequence variation in 88 genes associated with phenology and photoperiodism.
Chen et al (2016) Identifying genetic signatures of natural selection using pooled population sequencing in Picea abies. G3: Genes, Genomes, Genetics (Bethesda) doi:10.1534/g3.116.028753