Associations of TGFA and TGFB3 polymorphisms and NLCLP ,CPO

Objective: using PCP-SSCP and sliver-staining to analyze TGFA, TGFB3polymorphisms in nonsyndromic cleft lip with or without palate(NSCLP) and cleft palate only(CPO) and to evaluate the associations. Methods: TGFA and TGFB3 were amplified from peripheral leukocyte of 56 NSCLP (CLP 71.4%, CL28.6%),26 CPO patients and 28 unrelated controls. The primers were designed according to the 3'untranslated region of TGFA, 5th exon of TGFB3 published in the Internet. The PCR products were analyzed by single-stranded conformation polymorphism(SSCP). The aim fragments were further conformed by DNA sequencing after being cloned into pGEM-T vector. Results: The 345bp fragments of TGFA, 193bp of TGFB3 were amplified from NSCLP, CPO and control samples. In SSCP analysis, we found three alleles of TGFA, one of that of TGFB3. Sequencing results show three DNA polymorphic sites in TGFA and one in TGFB3 which are all base shifts. Conclusion:There are no direct associations between NSCLP ,CPO and controls. The PCR-SSCP with sliver-staining is an accurate, easy and unradiative method to identify gene polymorphism.