mRNA differential display of acute-phase proteins in experimental Escherichia coli infection

We present a modification of mRNA differential display in which increased t hroughput results from the use of an automated fluorescent sequencer. The s equence analysis is performed directly on purified fragments without furthe r cloning. The amplified fragments carry a T7 RNA polymerase promoter seque nce tag for in vitro transcription of riboprobes for nonradioactive in situ hybridization. We compared changes in gene expression in the liver and col on of group II phospholipase A2 transgenic and group ii phospholipase A2 de ficient mice during the course of experimental Escherichia coli infection. Fluorescent mRNA differential display comprising a 7 x 24 set of primers wa s used to study a total of 31 257 amplified cDNA fragments. Sequence analys is of the displayed fragments associated with infection identified classica l acute-phase proteins in the liver and host defense proteins in the colon. The displayed mRNAs associated to transgenicity were the transgene itself, i.e., human group II phospholipase A2, and glutathione-S-transferase in th e liver. In the colon, the displayed mRNAs associated with transgenicity we re the pancreatitis-associated protein and mucin. The results show that flu orescent mRNA differential display is a reliable method to identify differe nces in the expression of the genes of acute-phase proteins.