转猪FSHR和KISS1基因小鼠繁殖性能分析

发布时间：2018-01-19 01:25

本文关键词： FSHR基因 KISS1基因转基因小鼠产仔数阳性率猪　出处：《西南大学》2015年硕士论文　论文类型：学位论文

【摘要】：繁殖性能是家畜的重要经济性状,繁殖力低是限制现代畜牧业发展的原因之一。卵泡刺激素受体(Follicle Stimulating Hormone Receptor, FSHR)和亲吻素(KISS1)基因对家畜的产仔数有显著影响。卵泡刺激素(Follicle stimulating hormone,FSH)在促进和维持性腺发育和生殖功能中发挥重要作用,但是其生理作用需通过分布于细胞膜表面的卵泡刺激素受体(Follicle stimulating hormone receptor, FSHR)介导。KISS1基因在下丘脑-垂体-性腺轴系统中起到中枢节点作用。KISS1基因的翻译产物kisspeptins刺激GnRH分泌,激活性腺轴,影响黄体生成素(Luteinizing hormone,LH)和FSH的分泌,进而对哺乳动物生殖内分泌起调控作用。KISS1及其受体G蛋白偶联受体(G protein-coupled receptor 54,GPR54)基因还是调节动物初情期的关键基因。实验室前期通过分别构建小鼠卵巢颗粒细胞特异表达的猪FSHR基因(pFSHR)真核载体AMH-pFSHR和小鼠脑部特异表达猪KISS1基因(pKISS1)的真核载体pCaMKIIa-KISS1,通过显微注射的方法制备了转猪FSHR和KISS1基因小鼠。本文针对F1-F5代小鼠阳性率、pFSHR和pKISS1基因在小鼠体内的表达情况、以及对雌性小鼠繁殖性能的影响进行研究,实验结果如下：1.转猪FSHR基因小鼠：F1-F5代转基因小鼠平均每窝产仔数均高于野生型小鼠,F1和F3代的窝产仔数达到显著水平(P0.05)在转基因小鼠卵巢中特异表达,心脏、肝脏、脾脏、肾脏、子宫及下丘脑中则未检测到其表达,而在转基因小鼠和野生型小鼠的卵巢都有mFSHR mRNA的表达。猪FSHR蛋白在转基因小鼠卵巢中有表达,其表达水平在阳性小鼠之间没有显著差异,与mRNA水平结果一致。FSH、LH、雌二醇、孕酮在间情期转基因小鼠和野生型小鼠中没有显著差异(P0.05)。产仔数曲线表明转基因小鼠和野生型小鼠的窝产仔数随着年龄增加先升高,再降低；在同一鼠龄,转基因小鼠的产仔数比野生型小鼠多,且都在220日龄左右具有最大的窝产仔数,且达到显著水平(P0.05)；转基因小鼠的生产间隔时间比野生型小鼠更短(P0.05)。2.转猪KISS1基因小鼠： F1-F5代转基因小鼠平均每窝产仔数均高于野生型小鼠,而且F1和F2代的窝产仔数达到显著水平(分别为P0.01和P0.05)。pKISS1 mRNA在转基因小鼠下丘脑中有表达,其他组织中则未检测到其表达；在转基因小鼠和野生型小鼠的卵巢和下丘脑中都能检测到mKISS1 mRNA的表达。猪kisspeptin蛋白在转基因小鼠下丘脑中有表达。F4代转基因小鼠血清中FSH和雌二醇激素浓度显著高于野生型小鼠(P0.05)。产仔数曲线表明：同一鼠龄转基因小鼠比野生型小鼠具有更多的产仔数,都在200日龄左右具有最大的窝产仔数,且生产间隔时间比野生型小鼠更短。F4代转猪KISS1基因小鼠后代提前进入初情期,卵巢重量显著增加(P0.05)。
[Abstract]:Reproductive performance is an important economic trait of livestock. Low fecundity is one of the reasons that restrict the development of modern animal husbandry. Follicle Stimulating Hormone Receptor. FSHRand KISS1) genes have a significant effect on litter size of domestic animals. Follicle stimulating hormone. FSHs play an important role in promoting and maintaining gonadal development and reproductive function. However, its physiological function is through Follicle stimulating hormone receptor, which is distributed on the surface of the cell membrane. FSHR). KISS1 gene plays a central node role in hypothalamus-pituitary-gonadal axis system. The translation product of KISS1 gene kisspeptins stimulates the secretion of GnRH. The activation of gonadal axis affects the secretion of luteinizing hormone (LHH) and FSH. KISS1 and its receptor G protein coupled receptor G protein-coupled receptor 54. GPR54) gene is also the key gene to regulate the early estrus of animal. The porcine FSHR gene expressed specifically in granulosa cells of mouse ovary was constructed by constructing porcine FSHR gene pFSHRs in the early stage of laboratory. Eukaryotic vector AMH-pFSHR and mouse brain specific eukaryotic vector pCaMKIIa-KISS1 expressing porcine KISS1 gene pKISS1). The transgenic FSHR and KISS1 gene mice were prepared by microinjection. The expression of pFSHR and pKISS1 genes in mice with positive rate of F1-F5 generation was studied. The results were as follows: 1. The average litter size of transgenic mice with FSHR gene was higher than that of wild-type mice. The litter size of F1 and F3 generation reached a significant level (P0.05), but no expression was found in the ovary, heart, liver, spleen, kidney, uterus and hypothalamus of transgenic mice. The expression of mFSHR mRNA was found in the ovaries of transgenic mice and wild-type mice, and porcine FSHR protein was expressed in the ovaries of transgenic mice. There was no significant difference between the positive mice and the positive mice, and the results were consistent with the results of mRNA. There was no significant difference in progesterone between transgenic mice and wild type mice. The litter size curve showed that the litter size of transgenic mice and wild type mice increased first and then decreased with age. The litter size of transgenic mice was more than that of wild-type mice at the same age, and it had the largest litter size at 220 days old, and reached the significant level of P0.05; The production interval of transgenic mice was shorter than that of wild-type mice. The average litter size of transgenic mice of F1-F5 generation was higher than that of wild-type mice. Moreover, the litter size of F1 and F2 generation reached a significant level (P0.01 and P0.05, respectively. PKISS1 mRNA was expressed in the hypothalamus of transgenic mice. No expression was detected in other tissues. MKISS1 was detected in ovary and hypothalamus of transgenic mice and wild-type mice. Expression of mRNA. The expression of porcine kisspeptin protein in the hypothalamus of transgenic mice. F4 generation transgenic mice serum FSH and estradiol hormone concentrations were significantly higher than wild-type mice (. The curve of litter size showed that transgenic mice of the same age had more offspring than wild-type mice. All of them had the largest litter size at the age of 200 days, and the interval of production was shorter than that of wild type mice. The progenies of transgenic pig KISS1 gene mice of passage F4 entered the early stage of estrus earlier than those of wild type mice. The weight of ovary increased significantly (P 0.05).
【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S828

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