小鼠H1foo基因四环素诱导表达系统的构建与表达

Construction of Tetracycline-inducible Expression System of H1foo Gene in Mouse

卵母细胞特异性连接组蛋白H1foo(Linker histone H1foo)是连接组蛋白H1(Histone 1)在卵母细胞中特异性表达的1个亚型,H1foo在卵母细胞减数分裂和成熟发育过程中起着不可替代的作用,而且H1foo也参与了重编程过程。为了在体细胞中进一步研究H1foo的功能,本试验通过采用四环素诱导表达载体系统,用PCR的方法扩增了小鼠的H1foo基因和四环素诱导表达系统(Tet-on)的启动子区。用双酶切和连接等方法将上述元件连接到真核表达载体pVenus上,构建了重组质粒pVenus-tet-CMV-H1foo。双酶切和测序结果显示,片段连接正确,且全长测序正确。本试验所构建的四环素诱导的小鼠H1foo真核表达系统有助于人们在体细胞中更好地探讨H1foo在特定时间的作用和功能。

The oocyte-specific linker histone H1foo is a subtype of linker histone H1,H1foo plays an essential role during meiosis and maturation of the oocytes,moreover,it also participates in epigenetic reprogramming.In order to investigate the function of H1foofurther in somatic cells,we adopted the Tet gene expression system,The H1foo gene and the promoter of Tet-On regulatory region were amplified by PCR and inserted into pVenus to generate the recombinant vector pVenus-Tet-CMV-H1foo through double digestion and ligation.Restriction enzyme analysis and DNA sequencing showed that the sequence of the recombinant plasmid was correct and the fragments were properly connected.This Tetracycline-inducible mouse H1foo eukaryotic expression system will allow us to examine the role of H1foo at specific time points better during the somatic cell cycle.