Profiling the HeLa S3 transcriptome using randomly primed cDNA and massively parallel short-read sequencing

Biotechniques. 2008 Jul;45(1):81-94. doi: 10.2144/000112900.


Sequence-based methods for transcriptome characterization have typically relied on generation of either serial analysis of gene expression tags or expressed sequence tags. Although such approaches have the potential to enumerate transcripts by counting sequence tags derived from them, they typically do not robustly survey the majority of transcripts along their entire length. Here we show that massively parallel sequencing of randomly primed cDNAs, using a next-generation sequencing-by-synthesis technology, offers the potential to generate relative measures of mRNA and individual exon abundance while simultaneously profiling the prevalence of both annotated and novel exons and exon-splicing events. This technique identifies known single nucleotide polymorphisms (SNPs) as well as novel single-base variants. Analysis of these variants, and previously unannotated splicing events in the HeLa S3 cell line, reveals an overrepresentation of gene categories including those previously implicated in cancer.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA, Complementary / genetics*
  • Exons
  • Gene Expression Profiling*
  • HeLa Cells
  • Humans
  • Polymorphism, Single Nucleotide
  • RNA Splicing
  • RNA, Messenger / analysis
  • Transcription Initiation Site


  • DNA, Complementary
  • RNA, Messenger