核农学报2024，Vol.38Issue(11)：2106-2115,10.DOI:10.11869/j.issn.1000-8551.2024.11.2106
牦牛HMGN1多克隆抗体制备及应用
Preparation and Application of Polyclonal Antibody against Yak HMGN1 Protein
摘要
Abstract
The aim of this study was to clone the yak high mobility group nucleosome binding protein 1(HMGN1)gene and prepare a yak HMGN1 polyclonal antibody,so as to provide information for exploring the roles of HMGN1 in yak reproductive physiology.The HMGN1 gene sequence of yak was obtained by gene cloning and analyzed by bioinformatics.The relative expressions of HMGN1 in the yak uterus,ovaries and oviducts were detected by real-time fluorescence quantitative PCR(qRT-PCR).The recombinant protein of yak HMGN1 was purified by prokaryotic expression and affinity chromatography.10-month-old New Zealand white rabbits were immunized,and the polyclonal antibody against yak HMGN1 was purified by affinity chromatography.The expression and localization of HMGN1 in yak reproductive organs were detected by western blot(WB)and immunohistochemistry(IHC),respectively.The expression of the HMGN1 protein in granulosa cells was detected by immunofluorescence(IF).The HMGN1 gene of yaks was successfully cloned,and a polyclonal antibody against yak HMGN1 was prepared.The results show that the expressions of HMGN1 protein are higher in the uterus of the yak during the luteal phase and follicular phase than those in the gestation period(P<0.05).In ovarian tissues,the expressions of HMGN1 are the highest in the gestation period and lowest in the follicular phase(P<0.05).And in oviducts,the highest expression of HMGN1 was in the gestation period but the lowest in the luteal phase and follicular phase(P<0.05).The results of IHC show that HMGN1 is mainly located in the endometrium and uterine glands,theca and follicular granular layer,and mucosal epithelium of oviducts.And it has been found that HMGN1 is mainly expressed in the cytoplasm of granulosa cells by IF.It is stated that HMGN1 is involved in the regulation of the estrous cycle and pregnancy of the yak.The results of this study provide basic data for further study of the role of the HMGN1 protein in regulating the reproductive physiological mechanism of yaks.关键词
牦牛/HMGN1/原核表达/多克隆抗体Key words
yak/HMGN1/prokaryotic expression/polyclonal antibody引用本文复制引用
李玉平,王立斌,潘阳阳,王萌,张晖,张瑞,贾利毅,余四九..牦牛HMGN1多克隆抗体制备及应用[J].核农学报,2024,38(11):2106-2115,10.基金项目
国家自然科学基金(32273091),甘肃省自然科学基金(22JR5RA864) （32273091）
