Study On Serum AMH Concentration In Sows And The Mutation Detection And Tissues Expression Analysis Of AMH Gene

Anti-Mullerian hormone(AMH)is one of the transforming growth factor-P superfamily members,it can be suppressed to raise AMH primordial follicles and reduced the sensitivity of FSH,but its genetical mechanisms are unclear until now.In recent years,AMH has been extensively studied as an indicator of female ovarian reserve function,especially in female ovarian disease study.At the same time,AMH can also serve as a predictor of ruminant ovulation,and played a good role in ruminant breeding.However,the pig as an ideal experimental model animal for human disease,there are still a little about AMH in pig.This study provides some theoretical basis and scientific reference for AMH application in sow production management by related research on serum AMH concentration and AMH gene.In experiment 1,we collect blood samples of 401 sow in haining experimental field in Zhejiang province,the serum AMH concentrations were detected by porcine AMH ELISA kits,and the results showed that serum AMH concentrations in the normal sows were 2.8ng/mL?5.0ng/mL,infertile sows and sows with ovarian disease were 5.561ng/mL and 6.506ng/mL respectively,the values were more than normal sows.Our results showed that the determination of AMH concentrations in sow serum can indirectly be used as an important indicator for evaluation of the ovarian function and ovarian disease in sows.In experiment 2,we collected the ear tissue samples of 370 sows from haining experimental field.Firstly,we extracted the genomic DNA,and then PCR amplifiled the fragment of porcine AMH gene,using direct sequencing method for PCR products and Chromas software,we obtain the part sequences of porcine AMH gene.In addition,we identified a mutation of insert C at 400bp position by blast through DNAMAN software.Furthermore,we also found the insert mutation change the protein sequence of AMH gene.In order to further statistical analysis the effect of the insert mutation,we used driect sequencing method for the PCR products to genotyping 370 sows in our animal population.The results showed 17 sows had the insert C mutation.And then we analyzed their effect on the litter size of sows,the number of weaned piglets and serum AMH levels,the results showed that litter sizes of sows which have the insertion C mutation was significantly higher than that of sows without the insertion C mutation(p<0.05).In experiment 3,we used real-time fluorescence quantitative PCR method to detecting the expression of AMH gene mRNA in ten tissues from three 7 months old Jinhua sows,including heart,liver,spleen,lungs,kidneys,stomach,abdominal muscles,uterus and ovarian tissue.The expression analysis results showed that the expression of AMH mRNA in ovary was the highest,followed by lower expression in lung tissue,and no expression were detected in other tissues.