RNA-Seq of blueberry leave
The poly-A containing mRNA molecules were purified using poly-T oligo-attached magnetic beads. Following purification, the mRNA was fragmented into small pieces and the cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and N6 random primers. This was followed by second strand cDNA synthesis using DNA Polymerase I and RNase H to generate dscDNA. The 5 end of cDNA fragments were phosphorylated resulted a single A base at 3 end and subsequently ligated to the adapter. The resultant product was purified and enriched with PCR amplification to yield the final cDNA library. The PCR yield was quantified and was subjected to single strand circularized DNA molecule (ssDNA circle) preparation for final library construction.