SIMULTANEOUS AMPLIFICATION OF 4 DNA-REPAIR GENES AND BETA-ACTIN IN HUMAN-LYMPHOCYTES BY MULTIPLEX REVERSE TRANSCRIPTASE-PCR

We describe here the development, optimization, and use of a nonradioa ctive, quantitative, multiplex reverse transcriptase-PCR technique to measure, in a single reaction, the relative levels of the transcripts of four DNA repair genes (XPCC, hMSH2, XRCC1, and ERCC1) and the beta- actin gene in lymphoblastoid cell lines and frozen peripheral blood ly mphocytes. Expression of defective DNA repair genes was not detected i n DNA repair-deficient human cell lines, whereas the intact genes were detected in repair-proficient cell lines and in lymphocytes from a no rmal donor. The assay was reproducible, and repeated determinations of the same samples generated highly consistent results for each target gene. This approach should facilitate molecular epidemiological studie s that incorporate screening for germline alterations that may affect gene expression and for changes in the levels of gene expression.