Searching expressed sequence tag databases: Discovery and confirmation of a common polymorphism in the Thymidylate synthase gene

Databases of expressed sequence tags (EST) can be used to screen rapidly fo r potential polymorphisms in candidate proteins. As part of this study, we screened the gene for the enzyme thymidylate synthase (TS), TS is important physiologically because it is essential for the synthesis of deoxythymidyl ate, a nucleotide required for DNA synthesis and repair. TS is also a major target for cancer chemotherapeutic drugs, especially the widely used 5-flu orouracil. Using sequence alignment of ESTs, we identified a candidate 6-bp variation at bp 1494 in the 3'-untranslated region of the TS mRNA, This se quence variation occurred in 21 of 34 aligned ESTs at this location, includ ing ESTs from various tissue sources. The presence of this polymorphism was confirmed in a Caucasian population (n = 95) by polymerase chain restricti on amplification/RFLP analysis, The allele frequency of the 6-bp deletion w as found to be 0.29 (wild-type +6 bp/+6 bp, 48%; +6 bp/-6 bp, 44%; -6 bp/-6 bp, 7%), Although the function of this polymorphism has not yet been inves tigated, the 3'-untranslated region of a gene can play a role in mRNA stabi lity and translation. This study illustrates an approach to polymorphism di scovery in candidate enzymes of physiological interest by searches of publi cly available sequence data, a rapid and inexpensive method, The potential functional relevance of the common 6-bp deletion in the TS gene needs to be investigated, because this enzyme is plausibly of major importance not onl y in cancer treatment but also in cancer prevention.