cDNA derived from trophectoderm (TE) and embryonic disc (ED) of a single day 12 porcine embryo was subjected to next-generation sequencing using the Illumina platform. The short sequencing reads from triplicate sequencing runs were aligned to a custom database designed to represent the known porcine transcriptome. As expected, genes involved in epithelial cell function and steroid biosynthesis were more abundant in cells from the TE; genes involved in maintenance of pluripotency and chromatin remodeling were more highly expressed in cells from the ED. Quantitative real-time PCR was used to confirm the validity of the approach. We conclude that gene expression profiles of even extremely small samples (<or=1,000 cells) can be adequately described without RNA/cDNA preamplification. We also demonstrate the utility of pre-genome genomics resources-such as EST repositories-in the analysis and application of next-generation sequencing data in the absence of an appropriately annotated reference genome.
(c) 2010 Wiley-Liss, Inc.