FOXL2基因在不同日龄蛋鸡卵巢中的表达研究

发布时间：2018-05-15 20:17

本文选题：海兰褐蛋鸡 + FOXL2基因　；参考：《河南科技学院》2017年硕士论文

【摘要】：FOXL2基因是在脊椎动物中最先发现的与卵巢分化有关的的性别二态性标记,同时也是人类常染色体基因中第一个被认定在促进人的眼睑的早期发育、卵巢发育和维持卵巢功能方面具有重要作用的因子。随着人们对于相关研究的不断加深,发现在人的眼睑、胚胎及成体卵巢颗粒细胞中FOXL2基因均呈现出特异性的表达,与人的卵巢早衰、卵巢肿瘤、不孕不育、性别决定以及小眼睑裂综合征等病症有关。FOXL2基因在抑制卵泡凋亡和促进卵巢储备功能方面发挥着重要作用,其正常表达对鸡产蛋性能高低和卵巢发育程度很重要。但FOXL2基因在不同时期的产蛋鸡体内各组织器官中的表达情况以及与蛋鸡卵巢发育不同时期的相关性仍未见报道。本研究首先通过HE染色法观察不同时期海兰褐蛋鸡卵巢的发育情况和组织学结构,并通过进行卵泡数量的统计学分析,研究蛋鸡卵巢卵泡的具体发育状况。根据GenBank上收录的红原鸡的FOXL2基因序列(登录号:JF_708868),通过生物信息学分析其保守序列及酶切位点,利用Primmer_5.0设计并合成3对引物。通过琼脂糖凝胶电泳试验,筛选出最优引物。采集30日龄,60日龄,90日龄,120日龄、160日龄、220日龄、330日龄、480日龄共8个时期的蛋鸡的下丘脑、眼睑、肌肉、心脏、胸腺、肝脏、脾脏、肾脏组织样品,进行qR T-PCR试验。对比不同时期卵巢中FOXL2基因相对表达量的差异,分析FOXL2基因与鸡卵巢发育及其与产蛋性能的相关性;对比FOXL2基因在不同时期和不同组织中相对表达量的变化,分析FOXL2基因与蛋鸡机体发育的相关性。HE染色和统计学分析结果表明,220日龄卵巢的体积和重量最大,之后卵巢逐渐萎缩,重量减少;随日龄增加,总卵泡数逐渐减少,初级卵泡数呈现先增加后减少趋势,60日龄的初级卵泡数最高,然后逐渐减少;120日龄的次级卵泡数最高,并且明显高于其他日龄产蛋鸡;从30日龄开始,随日龄增加,卵泡闭锁率逐渐增多。qRT-PCR试验结果表明,120日龄鸡卵巢中FOXL2基因的相对表达量高于其他日龄。FOXL2的表达与蛋鸡卵巢发育存在相关性,产蛋鸡不同组织中FOXL2基因相对表达量伴随着产蛋鸡组织器官的发育而变化,说明FOXL2基因与产蛋鸡其他组织的发育也存在一定的关系。本研究有助于进一步认识FOXL2基因在调控蛋鸡卵巢发育和产蛋性能方面的作用,为深入研究该基因的生物学特性和功能奠定了基础。
[Abstract]:The FOXL2 gene is the first sex-dimorphic marker associated with ovarian differentiation found in vertebrates and the first of the human autosomal genes to be thought to promote the early development of the human eyelid. An important factor in ovarian development and maintenance of ovarian function. With the deepening of related studies, it has been found that FOXL2 gene is expressed specifically in human eyelid, embryo and adult ovarian granulosa cells, and it is associated with premature ovarian failure, ovarian tumor, infertility. FOXL2 gene plays an important role in inhibiting follicular apoptosis and promoting ovarian reserve function. The normal expression of FOXL2 gene plays an important role in laying performance and ovarian development degree. However, the expression of FOXL2 gene in various tissues and organs of laying hens at different stages and the correlation with different stages of ovarian development of laying hens have not been reported. In this study, the development and histological structure of the ovary of Hailan brown laying hens at different stages were observed by HE staining method, and the specific development status of ovarian follicles was studied by statistical analysis of the number of follicles. Three pairs of primers were designed and synthesized by bioinformatics based on the sequence of FOXL2 gene (accession number: JFs 708868) collected on GenBank. Three pairs of primers were designed and synthesized by Primmer_5.0. The best primers were screened by agarose gel electrophoresis. The hypothalamus, eyelid, muscle, heart, thymus, liver, spleen and kidney of 8 laying hens were collected for QR T-PCR test in the hypothalamus, eyelid, muscle, heart, thymus, liver, spleen and kidney. To compare the relative expression of FOXL2 gene in different stages of ovary, to analyze the relationship between FOXL2 gene and ovarian development and egg laying performance, and to compare the changes of relative expression of FOXL2 gene in different stages and tissues. The correlation between FOXL2 gene and the development of laying hens was analyzed. The results of HE staining and statistical analysis showed that the size and weight of ovaries were the largest at the age of 220 days, then the ovaries gradually shrank and the weight decreased, and the total follicles decreased with the increase of age. The number of primary follicles increased first and then decreased. The number of primary follicles at 60 days of age was the highest, and the number of secondary follicles at 120 days of age was the highest, which was significantly higher than that of other laying hens, and increased with the age of 30 days. The results of follicular atresia. QRT-PCR test showed that the relative expression of FOXL2 gene in the ovaries of 120-day-old hens was higher than that of other day-old chickens, and there was a correlation between the expression of FOXL2 gene and the development of ovaries of laying hens. The relative expression of FOXL2 gene in different tissues of laying hens changed with the development of the tissues and organs of laying hens, which indicated that the FOXL2 gene had a certain relationship with the development of other tissues of laying hens. This study is helpful to further understand the role of FOXL2 gene in the regulation of ovarian development and egg laying performance of laying hens and lay a foundation for further study on the biological characteristics and functions of the gene.
【学位授予单位】：河南科技学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S831

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