RNA-Seq of vallisneria natans: leaf lamina
Libraries were prepared using Illumina TruSeqTM RNA sample preparation Kit (San Diego, CA). Poly(A) mRNA was purified from total RNA using oligo-dT-attached magnetic beads and then fragmented by fragmentation buffer. Taking these short fragments as templates, double-stranded cDNA was synthesized using a SuperScript double-stranded cDNA synthesis kit (Invitrogen, CA) with random hexamer primers (Illumina). Then the synthesized cDNA was subjected to end-repair, phosphorylation and A base addition according to Illuminas library construction protocol. Libraries were size selected for cDNA target fragments of 200300 bp on 2% Low Range Ultra Agarose followed by PCR amplified using Phusion DNA polymerase (New England Biolabs, Boston, MA) for 15 PCR cycles. After quantified by TBS380, two RNAseq libraries were sequenced in single lane on an Illumina NovaSeq 6000 sequencer for 2150bp paired-end reads.