绵羊PRLR,PRLH和PRLHR基因的克隆及其在下丘脑—垂体—性腺中表达的研究
发布时间：2018-11-19 14:54
【摘要】：催乳素(PRL)是由垂体前叶合成的多肽激素,通过与其受体(PRLR)结合而发挥众多的生理功能,研究也发现催乳素释放激素(PRLH)及其受体(PRLHR)与PRL的分泌有关。本研究以绵羊为实验材料,以下丘脑-垂体-性腺轴为研究对象,运用分子生物学、免疫组化、荧光定量的方法对PRLR及PRLHR的具体表达情况作初步研究,其结果如下:(1)成功克隆了PRLR,PRLH和PRLHR。对PRLR基因序列进行同源性比对和进化树建立显示,其与藏羚羊的基因序列最接近;预测PRLR基因编码581个氨基酸的蛋白序列,其分子量为65.23kDa,等电点为p I5.12,且PRLR蛋白不太稳定;预测PRLHR编码含有370个氨基酸的蛋白序列,其分子量为41.05kDa,等电点为p I9.41,且PRLHR蛋白不稳定。对PRLH和PRLHR基因序列进行同源性比对和进化树建立显示,绵羊的PRLH与山羊的同源性最高,而PRLHR则与藏羚羊的同源性最高。(2)三个基因在绵羊的四种组织中均有表达。PRLR以表达量最少的子宫为标准计算其他组织的表达量,结果卵巢的表达量显著高于其他组织(P0.05),其表达量由高到低依次是卵巢下丘脑垂体子宫。PRLH以表达最少的下丘脑为标准,子宫的表达量显著高于其他组织(P0.05),组织的表达量由高到低依次是子宫卵巢垂体下丘脑;PRLHR以表达量最少子宫为标准,发现子宫和垂体之间的表达差异显著(P0.05);组织的表达量由高到低依次是垂体卵巢下丘脑子宫。(3)PRLR和PRLHR在绵羊的垂体、下丘脑、子宫和卵巢组织中的不同细胞中均有一定量的表达。PRLR和PRLHR在垂体上远侧端的阳性反应着色深,阳性细胞多呈圆形或卵圆形;同时它们在下丘脑中均有阳性反应,但染色强度都较轻;PRLR在子宫腔、子宫内膜以及子宫腺等部位均有阳性产物,而PRLHR只在子宫内膜处阳性反应明显,其他部位很少反应;卵巢中PRLR阳性细胞主要分布于卵母细胞和颗粒细胞中,相反PRLHR的阳性产物较少;除此之外PRLR的阳性反应区域还包括上述组织的血管和红细胞。
[Abstract]:Prolactin (PRL) is a polypeptide hormone synthesized from the anterior pituitary gland and plays a large number of physiological functions by binding to its receptor (PRLR). It has also been found that the prolactin releasing hormone (PRLH) and its receptor (PRLHR) are related to the secretion of PRL. In this study, sheep were used as experimental materials and hypothalamus-pituitary-gonad axis was used to study the specific expression of PRLR and PRLHR by molecular biology, immunohistochemistry and fluorescence quantitative method. The results are as follows: (1) PRLR,PRLH and PRLHR. were cloned successfully. The homology alignment of PRLR gene sequence and the establishment of evolutionary tree showed that the sequence of PRLR gene was the closest to that of Tibetan antelope. The protein sequence of 581 amino acids encoded by PRLR gene was predicted. Its molecular weight was 65.23 kDa, the isoelectric point was Pi 5.12, and the PRLR protein was not stable. It is predicted that PRLHR encodes 370 amino acids with molecular weight of 41.05 kDa, isoelectric point of Pi 9.41, and instability of PRLHR protein. The homology alignment of PRLH and PRLHR gene sequence and the establishment of evolutionary tree showed that the homology of PRLH between sheep and goat was the highest. However, PRLHR had the highest homology with Tibetan antelope. (2) three genes were expressed in four tissues of sheep. PRLR was used to calculate the expression of other tissues according to the least expressed uterus. Results the expression of ovary was significantly higher than that of other tissues (P0.05), and the expression of ovarian hypothalamus-pituitary uterus was in turn from high to low. The expression of PRLH was based on the hypothalamus, and the expression of uterus was significantly higher than that of other tissues (P0.05). The order of tissue expression from high to low was the hypothalamus of uterus, ovary and pituitary. The expression of PRLHR was the lowest in uterus, and the difference between uterus and pituitary was significant (P0.05). The order of tissue expression from high to low is pituitary ovary hypothalamic uterus. (3) PRLR and PRLHR in the pituitary and hypothalamus of sheep. The positive reaction of PRLR and PRLHR in the distal end of pituitary gland was deep and the positive cells were round or oval. At the same time, they all had positive reaction in hypothalamus, but the intensity of staining was light, PRLR had positive products in uterine cavity, endometrium and uterus gland, but PRLHR only showed positive reaction in endometrium, but few in other parts. PRLR positive cells were mainly distributed in oocytes and granulosa cells, on the contrary, the positive products of PRLHR were less. In addition, the positive regions of PRLR also included the blood vessels and erythrocytes of the above tissues.
【学位授予单位】：甘肃农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S826
本文编号：2342678
[Abstract]:Prolactin (PRL) is a polypeptide hormone synthesized from the anterior pituitary gland and plays a large number of physiological functions by binding to its receptor (PRLR). It has also been found that the prolactin releasing hormone (PRLH) and its receptor (PRLHR) are related to the secretion of PRL. In this study, sheep were used as experimental materials and hypothalamus-pituitary-gonad axis was used to study the specific expression of PRLR and PRLHR by molecular biology, immunohistochemistry and fluorescence quantitative method. The results are as follows: (1) PRLR,PRLH and PRLHR. were cloned successfully. The homology alignment of PRLR gene sequence and the establishment of evolutionary tree showed that the sequence of PRLR gene was the closest to that of Tibetan antelope. The protein sequence of 581 amino acids encoded by PRLR gene was predicted. Its molecular weight was 65.23 kDa, the isoelectric point was Pi 5.12, and the PRLR protein was not stable. It is predicted that PRLHR encodes 370 amino acids with molecular weight of 41.05 kDa, isoelectric point of Pi 9.41, and instability of PRLHR protein. The homology alignment of PRLH and PRLHR gene sequence and the establishment of evolutionary tree showed that the homology of PRLH between sheep and goat was the highest. However, PRLHR had the highest homology with Tibetan antelope. (2) three genes were expressed in four tissues of sheep. PRLR was used to calculate the expression of other tissues according to the least expressed uterus. Results the expression of ovary was significantly higher than that of other tissues (P0.05), and the expression of ovarian hypothalamus-pituitary uterus was in turn from high to low. The expression of PRLH was based on the hypothalamus, and the expression of uterus was significantly higher than that of other tissues (P0.05). The order of tissue expression from high to low was the hypothalamus of uterus, ovary and pituitary. The expression of PRLHR was the lowest in uterus, and the difference between uterus and pituitary was significant (P0.05). The order of tissue expression from high to low is pituitary ovary hypothalamic uterus. (3) PRLR and PRLHR in the pituitary and hypothalamus of sheep. The positive reaction of PRLR and PRLHR in the distal end of pituitary gland was deep and the positive cells were round or oval. At the same time, they all had positive reaction in hypothalamus, but the intensity of staining was light, PRLR had positive products in uterine cavity, endometrium and uterus gland, but PRLHR only showed positive reaction in endometrium, but few in other parts. PRLR positive cells were mainly distributed in oocytes and granulosa cells, on the contrary, the positive products of PRLHR were less. In addition, the positive regions of PRLR also included the blood vessels and erythrocytes of the above tissues.
【学位授予单位】：甘肃农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S826
【共引文献】
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2 陈骏;黎开燕;李艳;李文胜;;养血生乳口服液促进产后泌乳和产褥复旧的临床观察[J];现代中西医结合杂志;2014年14期
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2 秦姣;卡麦角林对黄毛鼠的不育效果及其作用机理[D];中国农业大学;2015年
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