The Study of MTHFR Gene C677T Polymorphism in Human Embryonic Blastomeres by Fluorescence PCR Method

Objectives: The aim of the present study was to evaluate methylene tetrahydrofolate reductase (MTHFR) gene C677T polymorphism in human pre-implantation embryos. Methods: Fifty embryos after biopsy at cleavage stage (which were inappropriate for assisted reproductive techniques) were analyzed in order to study MTHFR C677T polymorphism and sex determination. After cell lysis, in the first step, whole genome amplification (WGA) was performed by primer extension pre-amplification (PEP) PCR method. In the next step, fluorescence PCR with labeled primers was used to examine this polymorphism in the blastomeres. Finally, the Hinfl enzyme was employed to cut the fragments and the capillary electrophoresis was used to determine the size of PCR products. Results: only 37 blastomeres among 50 samples were successfully amplified; 23 male and 14 female embryos were observed. The frequencies of CC, CT and TT genotypes in embryos were 67.56%, 24.32%, 8.10%, respectively. Additionally, the frequencies of 677C and 677T alleles were 80% and 20%, respectively. Conclusions: In this study, the frequency of MTHFR c.677C˃T genotype was evaluated in human embryonic blastomeres before implantation. Therefore, this polymorphism may provide a new and valuable biomarker for evaluating the embryo usefulness and improving the In vitro fertilisation (IVF) process by choosing the best embryo. Our findings revealed that the detection rate of C677T polymorphism was 74% in the samples.