Study on the mRNA Expression Level and the Polymorphism of Follicle-Stimulating Hormone Receptor Gene in Taihu Pig’s Ovary
|School||Nanjing Agricultural College|
|Course||Animal Genetic Breeding and Reproduction|
|Keywords||Taihu pig FSHR 5’ regulatory region polymorphism gene expression litter size|
Taihu pig is a local variety of valuable resources, with a gentle temperament, fresh meat, bears crude feed, a significant heterosis, disease resistance and high reproductive performance. Depth study of the molecular mechanism of Taihu pig’s high breeding is very important for the cultivation of new varieties, improving reproductive performance. In order to further reveal the molecular mechanism of Taihu pig’s Reproductive performance, RT-PCR and real-time quantitative PCR were used to detect the expression level of the FSHR gene mRNA in ovarian tissue of Taihu pigs and commercial pigs, and to analyze the correlation of the expression level of the FSHR gene mRNA in ovarian tissue of Taihu pigs and the litter size of Taihu pig; by PCR amplification and cloning techniques obtained the 5’regulatory region of FSHR gene in Taihu pig, and analyzed the sequence features of the 5’regulatory region of FSHR gene in Taihu pig, through a variety of bioinformatics software; detected the polymorphism of FSHR gene 5’ regulatory region of Taihu pigs and commercial pigs by PCR-SSCP; analyzed the correlation of the expression of FSHR in ovarian tissue and the polymorphism of FSHR gene. The results were as follows:1, The expression level of FSHR gene mRNA in ovarian tissue of adult Erhualian was high, and was low in uterine tissue and fallopian tube, but did not express in other tissues. The expression of FSHR in the ovaries of adult Erhualian pigs was significantly higher than in the ovaries of commercial pigs (P=0.017). The expression of FSHR gene and the average number of born in later parities of Erhualian pigs was significantly positively correlated (P<0.05), the correlation coefficient was 0.8246. The expression of FSHR gene and the average number of born alive in later parities of Erhualian pigs was significantly positively correlated (P<0.05), the correlation coefficient was 0.8167. These indicate that there was a certain correlation between the mRNA expression of FSHR and the litter size of Erhualian pig.2, The length of the sequence of the FSHR 5’regulatory region of Mewashan pig was 1180bp by cloning and sequencing. Compared to the Genebank sequence, cattle, goat, mouse and human, the homology was 99.75%,67.17%,64.86%,36.49% and 33.56%.6 promoters were predicted by software.4 of them had the highest score, which were mainly distributed in the 25nt-369nt, and there were 15 transcriptional regulatory elements (score greater than or equal to 85), respectively, AML-la, CdxA, cap, SRY, S8, Oct-1, GATA-1, HSF2, TCF11, GATA-2, GATA-3, TATA and so on.3, Two mutations were detected by PCR-SSCP in FSHR gene 5’regulatory region of Meishan pig. In nt 1001, A mutated to T; in nt1001, A mutated to T. The mutation lead to enhance the prediction score of GATA-1 in the FSHR gene 5’ regulatory region of DD-Meishan pig. By comparative analysis of Meishan pigs and commercial pigs, the results showed that the A gene frequency and AA genotype frequency of Meishan pigs were 0.6223 and 0.4681, which were higher than in commercial pigs 0.3953 and 0.2093. The D gene frequency and DD genotype frequency of Meishan pigs were 0.3368 and 0.3053, which were higher than the commercial pig 0.2349 and 0.1807. The result showed that two mutations were moderate polymorphism by statistical analysis of the genotype homozygosity, effective number of alleles, heterozygosity and polymorphism information.4, The expression of FSHR mRNA in the ovarian tissue of the commercial pigs with different genotypes were comparativly analyzed by using RT-PCR and real-time fluorescence quantitative PCR. The result showed that the expression of AB-type FSHR was the highest, followed by AA-, and the expression of BB-type is the lowest. But there wasn’t significant difference between the three genotypes. The FSHR expression of DD-type was significantly higher than CC type and CD type, but there wasn’t significant difference between CC type and CD-type. The results showed that, the mutation in nt334 may affect the expression of FSHR.