Nucleic Acids Research, 2001, Vol. 29, No. 22 e114
© 2001 Oxford University Press
Preparing a human membrane and secreted protein-enriched cDNA library using PCR primers derived from a genomic database
1Department of Life Science, 2Department of Chemistry and 3Department of Computer Sciences, National Tsing Hua University, Hsinchu 300, Taiwan, 4Development Center for Biotechnology, Taipei, Taiwan and 5Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan
We describe here a strategy for preparing a human membrane and secreted protein (MSP)-enriched cDNA library based on human MSP- and non-MSP-encoding cDNA sequences in the databases. The signal peptide parts of the MSP-encoding cDNA sequences, which currently comprise about half of the estimated total number in humans, were analyzed for common patterns. These patterns form a ‘minimal’ set of polymerase chain reaction primer candidates of length varying from 9 to 21 nt. The products stemming from each primer candidate were determined and the results allowed us to obtain an ‘optimal’ mixed-length primer set. Ninety-six percent of the primers in this set were predicted to yield 
10% undesired products, and the desired MSP-cDNA products could be easily separated by gel electrophoresis. The present analysis establishes a methodology for preparing a cDNA library that enables the analysis of individual MSPs. This methodology may also help identify new MSPs. As many cell regulatory processes are mediated by secreted proteins and their membrane-bound receptors, the preparation of a MSP-enriched cDNA library should benefit research on MSPs.
* To whom correspondence should be addressed at: Institute of Biomedical Sciences, Academia Sinica, Taipei 11529, Taiwan. Tel: +886 2 2652 3031; Fax: +886 2 2788 7641; Email: carmay{at}gate.sinica.edu.tw Correspondence may also be addressed to Tse Wen Chang at: Development Center for Biotechnology, 81 Chang Hsing Street, Taipei, Taiwan. Tel: +886 2 2735 6237; Fax: +886 2 2739 4260; Email: twchang@mail.dcb.org.tw The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors