MEI1基因可变剪切事件对蒙古马精子生成的调控作用

doi:10.11843/j.issn.0366-6964.2022.04.010

Abstract

Abstract: The purpose of this study was to explore the regulatory effect of MEI1 gene with alternative splicing event on spermatogenesis in Mongolian horses, improve sperm quantity and semen quality of horses by using molecular regulation techniques, effectively improve reproductive efficiency, reduce production costs, accelerate genetic progress, and promote population reproduction. The testicular sertoli cells(SC) of two-year-old Mongolian horses were used as the subjects, the MEI1-1-pIRES2-EGFP recombinant plasmids without exon skip (ES) events and MEI1-2-pIRES2-Dsred recombinant plasmids with ES events were constructed and transfected into testicular sertoli cells, respectively. There were 3 repeats in each group. The proliferation and activity of transfected cells were detected by CCK-8 at 0, 24, 48, 72 h of transfection, the transfected cells were screened by G418. The transfected cells were collected and total RNA was extracted, and the differential expression of meiosis related genes in the two cells was detected by qRT-PCR after reverse transcription. The MEI1-1-pIRES2-EGFP and MEI1-2-pIRES2-Dsred vectors were successfully constructed. Agarose gel electrophoresis and sequencing experiments showed that the vectors construction was successful, and the recombinant plasmid mass spectrum was drawn. CCK-8 assay showed that the cells transfected with two plasmids had the highest cell activity and proliferation level at 48 h after transfection, and cells transfected with MEI1-2-pIRES2-Dsred had the highest cell activity. The transfection efficiency of transfected cells increased by G418 screening; The test results showed that some genes in meiosis pathway (BUB1, CUL1, CCNB1, PPP2R5C, CCNB2, etc.) and some factors regulating sperm meiosis in sertoli cell signaling pathway (ARID4A, FSH, GDNF, Stra8, NRG1, etc.) had significant differences in expression in the two types of cells, and all had higher expressions in SC transfected MEI1-2-pIRES2-Dsred plasmids. The MEI1 gene with ES events occurence is more conducive to regulating the meiosis and spermatogenesis, which has laid a theoretical foundation for exploring the mechanism of alternative splicing events affecting spermatogenesis in Mongolian horses.

Key words: Mongolian horse, MEI1 gene, alternative splicing, spermatogenesis

CLC Number:

S821.3

CUI Yingying, DUGARJAVIIN Manglai, LI Bei, Terigele, ZHAO Yiping, REN Xiujuan, SONG Lianjie, SU Shaofeng. Regulatory Role of MEI1 with Alternative Splicing Event on Spermatogenesis in Mongolian Horse[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(4): 1096-1108.

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Regulatory Role of MEI1 with Alternative Splicing Event on Spermatogenesis in Mongolian Horse

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