RNA-seq for Anisodus tanguticus: Old stems 2
here are two main ways to obtain mRNA: one is to enrich mRNA with polyA tails by Oligo(dT) magnetic beads using the structural feature that most eukaryotic mRNAs have polyA tails. two is to remove ribosomal RNA from total RNA to obtain mRNA. mRNAs are subsequently constructed in NEB Fragmentation Buffer with divalent The obtained mRNA was then randomly interrupted with divalent cations in NEB Fragmentation Buffer and library construction was performed according to the NEB normal library construction method or strand-specific library construction method. After library construction, initial quantification was performed using Qubit 2.0, diluting the library to 1.5 ng/ul, followed by detection of the insert size of the library using Agilent 2100, and after the insert size met expectations, qRT-PCR was performed to accurately quantify the effective concentration of the library (effective library concentration above 2 nM) to ensure library quality.