Marine Maroalga Kappaphycus alvarezii transcriptome under low temperature
Oligo-dT magnetic beads used to purify the poly-A mRNA from the total RNA. Fragmentation buffer was added to cleave mRNA into short fragments (≈155 bp) and the cleaved RNA fragments were used for first stand cDNA synthesis using reverse transcriptase and random primers. Double-stranded cDNA was synthesized using the SuperScript Double-Stranded cDNA Synthesis kit (Invitrogen, Camarillo, CA) from Illumina. The synthesized cDNA was subjected to end repair, phosphorylation, 3′adenylation and adapter ligation in sequential. AMPure XP beads were used to purify amplified templates before enriching properly ligated template strands, to generate enough DNA. Quantification of dsDNA was carried out by Quantifluor-ST fluorometer (Promega, E6090) and Quant-iT PicoGreen dsDNA Assay Kit (Invitrogen, P7589). cDNA libraries were further verified using a 2100 Bioanalyzer RNA Nanochip (Agilent, SantaClara, CA).