Combined laser capture microdissection and serial analysis of gene expression from human tissue samples

摘要: Cell-specific gene expression profiling from heterogeneous human tissues is confounded by cell purification limitations. Here, we describe a technique to generate profiles of pure populations prostate cancer cells obtained fresh-frozen prostatectomy specimens and small initial quantities RNA combining laser capture microdissection microserial analysis (LCM–microSAGE). Two microSAGE libraries were approximately 100 000 pulses, estimated contain fewer than 3 × 105 20–30 ng mRNA. sequenced depth 10 111 10 463 unique tags normal cells, representing 6453 6923 genes, respectively. Most transcripts expressed at similar levels, but compared with had increased 385 decreased 389 tags. A total 20 genes differentially (P<0.05); five these upregulated 15 downregulated in cells. Quantitative reverse transcriptase-polymerase chain reaction results three selected corroborated the existence cell-specific LCM–microSAGE-derived libraries. In conclusion, LCM–microSAGE approach demonstrates that large-scale known unknown can be generated target tissue samples comprised mixtures types.