摘要
【目的】克隆西农萨能山羊泌乳中期和后期乳腺组织差异表达基因Ubi-d4并进行生物信息学分析,为进一步研究该基因的功能以及乳腺退化的分子机制提供信息。【方法】以西农萨能羊乳腺组织为材料,利用抑制性消减杂交技术筛选泌乳中期和后期乳腺组织差异表达基因,用RT-PCR克隆Ubi-d4基因,用实时定量PCR分析Ubi-d4基因的表达水平变化,用生物信息学方法对Ubi-d4基因进行结构和功能域预测。【结果】山羊乳腺Ubi-d4基因开放读码框全长1176bp,编码391个氨基酸,GenBank登陆号为EF105410;与牛(XM_881516)、人(NM_006268.3)、鼠(NM_011262)、兔(XM_341998)对应基因的核苷酸和氨基酸同源性分别为98%、90%、93%、90%和99%、99%、99%、98%,泌乳后期乳腺组织中mRNA表达水平是泌乳中期的6.10倍;氨基酸序列中含有一个C2H2型锌指结构,一个PHD型锌指结构,没有信号肽和跨摸结构。【结论】筛选到西农萨能山羊乳腺泌乳后期高水平表达基因Ubi-d4,克隆了它的开放读码框(1176bp,GenBank No:EF105410);生物信息学分析推测它是乳腺退化过程中的重要功能基因。
【Objective】Screening,cloning and bioinformatics analysis of Ubi-d4 gene differently expressed in mammary gland between middle and late lactation stage. 【Method】Suppression subtractive hybridization （SSH） approach was used to isolate differently expressed genes in mammary gland of late lactation stage compared to middle stage,reverse transcriptase-polymerase chain reaction （RT-PCR） was implied to clone gene,real-time reverse transcriptase-polymerase chain reaction （RT-PCR） was used to analysis the change of Ubi-d4 mRNA level between middle and late lactation stage,and biotechnology were employed to predict gene specific structure and potential motif domain.【Result】Goat mammary gland zinc-finger protein Ubi-d4 had an open reading frame of 1176 nucleotides encoding a putative protein of 391 amino acids,the GenBank No was EF105410; the homologies of nucleotide and peptide sequence of this Ubi-d4 with bovine （XM_881516）,human （NM_006268.3）,mouse’s （NM_011262） and rabbit （XM_341998） were 98%,90%,93%,90% and 99%,99%,99% and 98%,respectively; the mRNA level of Ubi-d4 gene in mammary gland of late lactation stage was 6.10 folds higher than that in middle stage normalized by housekeeping gene Glyceraidehyde-3-phosphate dehydrogenase. The amino acid sequence of Ubi-d4 included one C2H2 type zinc-finger,one PHD-type zinc-finger,and had no signal peptide,no transmembrane structure. 【Conclusion】Ubi-d4 gene was identified which was highly expressed in mammary gland of late lactation stage,its open reading frame was cloned （1 176 bp,GenBank No： EF105410）. We predicted that goat zinc-finger protein Ubi-d4 gene is an important functional gene in mammary gland involution process by bioinformatics analyzing.
出处
《中国农业科学》
CAS
CSCD
北大核心
2008年第6期1804-1809,共6页
Scientia Agricultura Sinica
基金
教育部新世纪优秀人才支持计划项目(NCET-05-0857)
西北农林科技大学研究生教育创新计划项目(05YCH021)
