西农萨能奶山羊泌乳中期和后期乳腺组织差异基因的筛选

畜牧兽医学报 ›› 2010, Vol. 41 ›› Issue (1): 16-21.doi:

西农萨能奶山羊泌乳中期和后期乳腺组织差异基因的筛选

武会娟^1,2，罗军¹*，赵旺生¹，王伟¹

1. 西北农林科技大学动物科技学院，杨凌 712100；2. 北京市理化分析测试中心，北京 100193

收稿日期:2008-12-24 修回日期:1900-01-01 出版日期:2010-01-24 发布日期:2010-01-24
通讯作者: 罗军

Screening of Differentially Expressed Genes in Mammary Gland of Xinong Saanen Dairy Goat at Middle and Late Lactation Stages

WU Huijuan^1,2, LUO Jun ¹*, ZHAO Wangsheng ¹, WANG Wei ¹

1. College of Animal Science and Technology, Northwest A&F University, Yangling 712100,China; 2. Beijing Center for Physical and Chemical Analysis, Beijing 100193, China

Received:2008-12-24 Revised:1900-01-01 Online:2010-01-24 Published:2010-01-24
Contact: LUO Jun

摘要/Abstract

摘要： 旨在利用抑制性削减杂交（SSH）技术筛选西农萨能奶山羊泌乳中期和后期的差异表达基因，并用实时定量PCR（QPCR）分析差异基因的表达丰度，探讨奶山羊不同泌乳阶段乳腺组织的基因表达规律。本研究以西农萨能奶山羊泌乳中期和后期乳腺组织的mRNA互为检测子（Tester）和驱动子(Driver)构建cDNA消减文库（ML和LM），随机挑选克隆测序，进行序列比对分析，并检测部分差异基因在乳腺组织中的表达丰度。结果，成功构建了泌乳中期和后期的cDNA文库，随机挑选30个克隆测序，得到ML和LM文库中与细胞凋亡、抗氧化、脂类代谢、能量代谢等生理过程相关的差异基因，对其中的6个基因进行实时定量分析，发现5个均为阳性克隆，表达水平增加了1.3～5.5倍不等。结果表明，利用SSH技术成功构建了泌乳中期和后期乳腺组织正反向消减cDNA文库，筛选出24个差异基因，为进一步研究奶山羊乳腺组织基因调控机理奠定了基础。

Abstract: The objective of this study was to use suppression subtractive hybridization (SSH) to screen the differentially expressed genes in mammary gland of Xinong Saanen Dairy goats at the middle and late lactation stages, and the expression abundance of several genes were analyzed by realtime quantitative PCR (QPCR) to explore the gene expression mechanism of mammary gland in the different lactation stages of dairy goats. The subtracted cDNA libraries (ML and LM) were constructed by using mRNA of mammary gland of Xinong Saanen Dairy goat at the middle and late lactation stages as the Tester and/or Driver, clones were randomly selected and sequenced for sequence alignment, the expression abundance of several genes in mammary gland was analyzed. 30 randomly selected clones from the constructed cDNA subtraction libraries of mammary gland at middle and late lactation stages were sequenced, and the differentially expressed genes related with physiological process including cell apoptosis, antioxidation, lipid and energy metabolism etc. were obtained. Five positive clones were observed from 6 genes assayed by quantitative PCR, the expression levels of which were 1.35.5 folds higher in mammary gland of either middle or late lactation stage. The results showed that the forward and reverse subtracted cDNA libraries of mammary gland at middle and late lactation stages were constructed using SSH, the screening of 24 differentially expressed genes laid the foundation for the further studies on gene regulation in the mammary gland of dairy goats.

武会娟;罗军;赵旺生;王伟. 西农萨能奶山羊泌乳中期和后期乳腺组织差异基因的筛选[J]. 畜牧兽医学报, 2010, 41(1): 16-21.

WU Huijuan;LUO Jun;ZHAO Wangsheng;WANG Wei . Screening of Differentially Expressed Genes in Mammary Gland of Xinong Saanen Dairy Goat at Middle and Late Lactation Stages[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2010, 41(1): 16-21.

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