title: De-novo transcriptome analysis of intestinal fluke Fasciolopsis buski
design: •2 ug micrograms of genomic DNA for each sample was made up to 100 ul with nuclease free water (Ambion) and sonicated using a Bioruptor (Diagenode) to obtain desired fragment length ranging between 150 to 600 bp for short insert library and 200 to 600 bp for long insert library.  •Fragmentation conditions (Bioruptor)  •FD2 Short insert library – (30 pulses at high for 30s ON and 30s OFF). Sonication was repeated with the same conditions for another aliquot of 2 ug.  •FD2 Long insert library – (20 pulses at high for 30s ON and 30s OFF)  •The size distribution was checked by running an aliquot of the sample on Agilent 7500 Bioanalyzer Chip. The fragmented DNA was cleaned up using 0.7X Agencourt Ampure XP SPRI beads (Beckman Coulter) for long insert libraries and 1.8X Agencourt for short insert libraries.  Yield of Recovered DNA after fragmentation    FD2 Short insert library: – (464 ng for 1st aliquot, 1472 ng for 2nd aliquot). The recovered DNA from both aliquots were pooled and 1 ug was taken for library preparation  FD2 Long insert library: – (1281 ng). 1 ug was taken for library preparation  Subsequently libraries for whole genome sequencing were constructed according to the TruSeq DNA library protocol outlined in “TruSeq DNA Sample preparation guide”. DNA was subjected to a series of enzymatic reactions that repair frayed ends, phosphorylate the fragments, and add a single nucleotide ‘A’ overhang and ligate adaptors (Illumina’s TruSeq DNA sample preparation kit). Sample cleanup was done using Ampure XP SPRI beads. After ligation, ~300-350 bp fragment for short insert libraries and ~500 – 550 bp fragment for long insert libraries was size selected by gel electrophoresis, gel extracted and purified using Minelute columns (QIAGEN). The libraries were amplified using 10 cycles of PCR for enrichment of adapter-ligated fragments. The prepared libraries were quantified using Nanodrop and validated for quality by running an aliquot on High Sensitivity Bioanalyzer Chip (Agilent).