RNA-seq P. phosphorea
After the total extraction of RNA and DNase I treatment, the following steps were applied; mRNA enrichment: Oligo dT Selection or rRNA depletion; RNA fragment and reverse transcription: Fragment the RNA and reverse transcription to double-strand cDNA (dscDNA) by N6 random primer; End repair, add A tailing and adaptor ligation: The synthesized cDNA was subjected to end-repair and then was 3 adenylated. Adaptors were ligated to the ends of these 3 adenylated cDNA fragments; PCR amplification: The ligation products were purified and many rounds of PCR amplification were performed to enrich the purified cDNA template using PCR primer; Denature and cyclization: Denature the PCR product by heat and the single strand DNA is cyclized by splint oligo and DNA ligase. During the QC steps, Agilent 2100 Bioanalyzer and ABIStepOnePlus Real-Time PCR System were used in quantification and qualification of the sample library. At last, the library was sequenced on BGISEQ-500 platform.
SUB14642123
Pph_rachis