Technical variations in low-input RNA-seq methodologies.

ABSTRACT: Recent advances in RNA-seq methodologies from limiting amounts of mRNA have facilitated the characterization of rare cell-types in various biological systems. So far, however, technical variations in these methods have not been adequately characterized, vis-à-vis sensitivity, starting with reduced levels of mRNA. Here, we generated sequencing libraries from limiting amounts of mRNA using three amplification-based methods, viz. Smart-seq, DP-seq and CEL-seq, and demonstrated significant technical variations in these libraries. Reduction in mRNA levels led to inefficient amplification of the majority of low to moderately expressed transcripts. Furthermore, noise in primer hybridization and/or enzyme incorporation was magnified during the amplification step resulting in significant distortion

SUBMITTER: Bhargava V 

PROVIDER: S-EPMC3890974 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature