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Identification of genes with specific expression in pancreatic cancer by cDNA representational difference analysis
Author(s) -
Gress Thomas M.,
Wallrapp Christine,
Frohme Marcus,
MüllerPillasch Friederike,
Lacher Ulrike,
Friess Helmut,
Büchler Markus,
Adler Guido,
Hoheisel Jörg D.
Publication year - 1997
Publication title -
genes, chromosomes and cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.754
H-Index - 119
eISSN - 1098-2264
pISSN - 1045-2257
DOI - 10.1002/(sici)1098-2264(199706)19:2<97::aid-gcc5>3.0.co;2-v
Subject(s) - complementary dna , biology , representational difference analysis , pancreatic cancer , suppression subtractive hybridization , gene , microbiology and biotechnology , rapid amplification of cdna ends , expressed sequence tag , gene expression , carcinogenesis , northern blot , cdna library , cancer , genetics , molecular cloning
Abstract cDNA representational difference analysis (cDNA‐RDA) is a polymerase‐chain‐reaction‐coupled subtractive and kinetic enrichment procedure for the isolation of differentially expressed genes. In this study, the technique was used to isolate novel genes specifically expressed in pancreatic cancer. cDNA‐RDA was done on cDNA reverse transcribed from a poly(A) + mRNA pool made from 10 cancer tissues (tester) by using as a driver a cDNA from a poly(A) + mRNA pool made from a combination of 10 tissues of chronic pancreatitis and 10 healthy pancreatic tissues. The use of chronic pancreatitis in addition to healthy pancreas mRNA in the driver preparation eliminated the influence of stromal tissue components present as contamination in the cancer‐specific preparations. Such cDNA‐RDA led to the isolation of 16 distinct, cancer‐specific gene fragments. These were confirmed to be overexpressed in pancreatic cancer tissues by Northern blot analysis. Sequence analysis revealed homologies to five genes previously implicated in the carcinogenesis of the pancreas or other tissues. Eleven fragments had no significant homology to any known gene and thus represent novel candidate disease genes. The experiments demonstrate that cDNA‐RDA is a reproducible and highly efficient method for the identification of novel genes with cancer‐specific expression. Genes Chromosom. Cancer 19:97–103, 1997. © 1997 Wiley‐Liss, Inc.