半番鸭性腺发育关键基因17β-HSD3的克隆、序列特征及表达分析  

Cloning,Sequence Features and Expression Analysis of 17β-HSD3,A Key Gene Involved in Gonadal Development of Mulard Ducks(Cairina moschata)

作　　者：缪中纬[1,2] 李丽 朱志明[1,2] 章琳俐 郝晓娜[1,3] 黄勤楼 郑嫩珠[1,2] 辛清武 MIAO Zhong-Wei;LI Li;ZHU Zhi-Ming;ZHANG Lin-Li;HAO Xiao-Na;HUANG Qin-Lou;ZHENG Nen-Zhu;XIN Qing-Wu(Institute of Animal Husbandry and Veterinary Medicine,Fujian Academy of Agricultural Sciences,Fuzhou 350013,China;Fujian Key Laboratory of Animal Genetics and Breeding,Fuzhou 350013,China;College of Animal Science(College of Beeology),Fujian Agriculture and Forestry University,Fuzhou 350002,China)

机构地区：[1]福建省农业科学院畜牧兽医研究所,福州350013 [2]福建省畜禽遗传育种重点实验室,福州350013 [3]福建农林大学动物科学学院(蜂学学院),福州350002

出　　处：《农业生物技术学报》2023年第4期801-811,共11页Journal of Agricultural Biotechnology

基　　金：福建省自然科学基金(2021J01483;2020J01348);福建省省属公益类项目(2020R10260016;2022R1026008);福建省“5511”协同创新工程(XTCXGC2021008);福建省农科院科技创新团队(CXTD2021006-2);福建省现代家禽产业技术体系(2019-2022)。

摘　　要：17β-羟基类固醇脱氢酶(17β-hydroxysteroid dehydrogenase,17β-HSD)可将经胆固醇转化而成的雄烯二酮最终氧化为睾酮,对睾丸激素合成和雄性性状发育至关重要。本课题组前期研究发现17β-HSD3可能通过类固醇激素合成途径参与半番鸭(Cairina moschata)胚胎期性腺发育。本研究以半番鸭为研究对象,通过cDNA末端快速扩增技术(rapid amplification of cDNA ends,RACE)获得17β-HSD3基因全长cDNA序列,利用生物信息学对其序列特征和蛋白理化性质进行分析预测,并构建系统进化树;采用qPCR技术分析17β-HSD3基因在半番鸭胚胎期不同组织(脑,心,肝,肺,肌肉,性腺和肾)中的特异性表达以及在半番鸭、父本番鸭和母本北京鸭性腺中的差异表达。结果表明,半番鸭17β-HSD3基因全长cDNA序列全长为1332 bp,含有1个963 bp的开放阅读框、1个26 bp的5’非编码区和1个343 bp的3’非编码区。编码的蛋白质含有320个氨基酸,该编码蛋白为不稳定性蛋白。同源性和进化树系统分析表明,半番鸭17β-HSD3基因与天鹅(Cygnus olor)亲缘关系最近,同源性高达96.5%。组织特异性表达分析显示,17β-HSD3基因在半番鸭性腺中的表达量极显著高于其他组织(P<0.01),同时,随着胚胎发育17β-HSD3基因表达量在半番鸭性腺组织中逐渐升高,表明该基因可能参与半番鸭性腺发育;17β-HSD3基因在繁殖性状正常的番鸭和北京鸭性腺中表达量显著高于不育的半番鸭(P<0.05),表明其表达可能与半番鸭的不育相关。本研究为进一步研究17β-HSD3基因对半番鸭性腺发育和生殖细胞分化的调控机理提供理论依据。The 17β-hydroxysteroid dehydrogenase(17β-HSD)eventually oxidizes androstenedione converted from cholesterol to testosterone,which is essential for testosterone synthesis and male reproductive traits.17β-HSD3 was found to be involved in the pre-embryonic gonadal development of the Mulard duck(Cairina moschata)through the steroid hormone synthesis pathway.In this study,the full-length sequence of 17β-HSD3gene was obtained by rapid amplification of c DNA ends(RACE)cloning method,sequence characteristics and physicochemical properties of proteins were predicted using bioinformatics analysis,and phylogenetic trees were constructed.The qPCR was utilized to analyze the tissue-specific expression of 17β-HSD3 gene in Mulard duck(brain,heart,liver,lung,muscle,gonad,kidney)and differential expression in the gonads of Mulard ducks,male parent Muscovy and female parent Pekin ducks.The results showed that the full-length sequence of 17β-HSD3 gene was 1332 bp in Mulard duck,containing a 963 bp of open reading frame,and the length of 5’and 3’non-coding regions were 26 bp and 343 bp,respectively.The encoded protein contained 320amino acids,which was an unstable protein.Homology and phylogenetic tree phylogenetic analysis showed that the 17β-HSD3 gene in Mulard duck have the closest relationship with swans(Cygnus olor).Tissue-specific expression analysis revealed that the 17β-HSD3 gene was highly significantly more expressed in the gonads of Mulard ducks than in other tissues(P<0.01),suggesting that 17β-HSD3 gene might be involved in gonadal development in Mulard duck.The expression of the 17β-HSD3 gene was significantly higher in Muscovy ducks and Pekin ducks with normal reproductive traits than in Mulard ducks(P<0.01),revealing that 17β-HSD3 gene might be associated with sterility in Mulard ducks.The study provides a theoretical basis for the function of the 17β-HSD3 gene in gonadal development,germ cell differentiation and other reproductive processes in the Mulard duck.

关 键 词：半番鸭 RACE克隆 性腺 17β-羟基类固醇脱氢酶3(17β-HSD3) 

分 类 号：S813.22[农业科学—畜牧学]