Degradome sequencing in Swarnaprabha Rice under 75% shade-Test sample
Degradome sequencing libraries were prepared using a modified protocol from TruSeq Small RNA Sample Preparation. PolyA mRNA was isolated using 50g of total RNA by employing Dynabeads Oligo (dT)25 magnetic isolation kit. The isolated polyA mRNA was subjected to 3 and 5 adapter ligation. Adapter ligated fragment was reverse transcribed with Superscript III reverse transcriptase by priming with reverse transcription primers. cDNA formed was enriched and barcoded by PCR amplification. Amplified library was size selected using Polyacrylamide gel. Libraries were size selected in the range of 140 bp - 350 bp followed by overnight gel elution and ethanol precipitation in presence of glycogen and 3M sodium acetate.Illumina-compatible sequencing libraries were quantified by Qubit fluorometer and fragment size distribution of the libraries were analyzed on Agilent 2200 TapeStation.