Plant sample
Library preparation and sequencing were performed at Genotypic Technologys Genomics facility using SureSelectXT Reagent Kit, ILM, 16 (Catalog # G9611A). 200 nanograms of genomic DNA from each sample was made up to 50 ul with Low TE and sonicated for ~300 sec on covaris to fragment DNA into size ranging between ~200 to 400 bp (Covaris). The size distribution was checked by running an aliquot of the sample on Agilent High Sensitivity Tape Station. Subsequently, genomic DNA libraries were constructed according to SureSelectXT Target Enrichment System for Illumina Paired-End Sequencing Library manual (Part # G7530-90000 VersionB.3, June 2015). DNA was subjected to a series of enzymatic reactions that repair frayed ends, phosphorylate the fragments, add a single nucleotide A overhang and ligate adaptors. After ligation, PCR amplification (10 cycles) was performed to enrich the adaptor-ligated fragments. The prepared library was then kept for hybridization as outlined in SureSelectXT Target Enrichment System for Illumina Paired-End Sequencing Library manual. Hybridized library fragments were isolated by magnetic capture using Magnetic Streptavidin coated beads(T1). The captured library beads were resuspended in 15 ul of NFW. PCR amplification (12 cycles) was carried out according to the protocol to amplify the captured library. Index was added at this stage. Clean up was done using HighPrep PCR clean up system. Captured Library was quantified using Qubit fluorometer. An aliquot of the captured library was run on a Agilent High Sensitivity Tape Station.