Construction of eukaryotic expression vector of CCND1 gene in laying hens and its spatiotemporal expression pattern in different developmental stages of follicles
ZOU Xianqun
YU Jing
LIU Yongjie
CHEN Xiaoxia
Abstract：The experiment was to explore the spatiotemporal expression pattern of cyclin D1(CCND1)gene in laying hens follicles at different development stages and delve into its molecular biological functions.A eukaryotic expression vector has been constructed to predict CCND1's specific molecular roles during laying hens follicle development.Follicles of varying sizes from 150-day-old Hyline brown laying hens were selected as experimental materials.Semi-quantitative RT-PCR and immunohistochemistry were used for molecular-level spatiotemporal expression analysis.After verifying the ligation product's correctness via double enzyme digestion with BamH Ⅰ and XhoⅠ,sequencing was performed.The correctly sequenced eukaryotic expression vector was then transfected into chicken ovarian granulosa cells,and the transfection efficiency is confirmed using real-time fluorescent quantitative PCR.The results showed that CCND1 mRNA expression was significantly higher in follicles with diameters of 5～6 mm and 6～7 mm than in other follicles(P＜0.05).CCND1 protein localization was observed in oocytes and theca of pre-hierarchical follicles.A eukaryotic expression vector pYr-adshuttle-4-CCND1 had been successfully constructed through colony PCR amplification and sequencing analysis and effectively transfected into chicken follicle granulosa cells.The study indicates that CCND1 likely plays a role in laying hens follicle growth and development,offering a vital molecular basis and tool for future gene-function and application studies.
Keywords：laying hensCCND1follicleeukaryotic expression vectorsspatiotemporal expression
Publication Date：2025-05-16
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：6( 64-69 )