Sequencing-Based Association Study of c.677C> T in MTHFR Gene with Azoospermia in Iranian Male

Background and Objective: Infertility is defined as inability to conceive after 1 year of unprotected intercourse. Infertility involve about 15% of couples (20-25% in Iran) with the man being responsible for nearly half of the cases. Both genetics and environmental factors play role in male infertility. Environmental factors include smoking, high temperature, pollutions, nurture and other life behaviors. Association of some genetic polymorphisms are one of the most important genetic factors which cause infertility. MTHFR is a gene maps to human chromosome 1p36.22 which encodes the protein MTHFR that is essential to DNA methylation and spermatogenesis and the mutation c.677C> T in this gene has been proposed to be related with male infertility. In this study our propose was to investigate the association of c.677C> T in MTHFR gene in Iranian male by sequencing. Materials and Methods:Peripheral blood samples of 100 azoospermia patients were collected from Jihad University of Qom. DNA was extracted by salting-out method. Then PCR was performed and 25 samples (from turk and fars ethnics) with the reverse primer were sequenced by Sanger, s method. Then the results from sequencing were compared with previously performed RFLP analysis of the rs1801133. Furthermore, networking analysis was performed for genetical variation between sequenced samples by PopArt software. Findings: The sequences obtained covered 109 SNPs within MTHFR Gene in this region. The analysis of sequences showed no polymorphism within the studied samples and therefore revealed no association between these SNPs and azoospermia. This is in agreement with RFLP and χ2-squared analysis of the same samples performed for rs1801133. The results of networking showed no variation between samples. Therefore, gene flow between individuals of these two ethnics is proved. Conclusion: In our study the c.677C> T SNP with rs1801133 was not related to Iranian male azoospermia, but further studies with more samples are demanded for association study using case-control method. Furthermore, the results of association of a special polymorphism vary between different populations, showing the role of the geographic factors or ethnic and diet in association studies.

کلیدواژه ها:

infertility ، azoospermia ، MTHFR Gene ، Single Nucleotide Polymorphism (SNP) ، sequencing