半定量RT-PCR法分析猪肌肉组织细胞谷胱甘肽过氧化物酶mRNA表达水平  

RT-PCR method for the semiquantitative analysis of cGPX mRNA level in pig muscle

作　　者：徐春兰[1] 汪以真[1] 

机构地区：[1]浙江大学饲料科学研究所,浙江杭州310029

出　　处：《中国兽药杂志》2005年第8期3-6,23,共5页Chinese Journal of Veterinary Drug

基　　金：国家973重点基础研究发展计划资助项目(2004CB117506)

摘　　要：细胞谷胱甘肽过氧化物酶(cGPX)是动物体内一种重要的抗氧化酶。本实验室构建一优化的半定量RTPCR法,以18srRNA为内标,研究猪肌肉组织中cGPX基因mRNA表达水平。提取猪肌肉组织总RNA,经反转录后进行扩增,先寻求PCR线性扩增范围,确定RTPCR的最佳循环次数和Mg2+浓度,扩增后用VDS摄像系统扫描PCR扩增产物的电泳条带灰度。通过cGPXPCR产物的灰度与18srRNAPCR产物的灰度之比,即可计算出cGPXmRNA的相对含量。Cellar glutathione peroxidase （cGPX） is an important antixidase in animals. In this article RT-PCR method is developed for the semiquantitative analysis of cGPX mRNA. 18s rRNA is used as internal standard. Total RNA is extracted from pig muscle tissue. The mRNA is reverse-transcripted and the cDNA fragments of cGPX and 18s rRNA gene are amplified by RT-PCR using the gene specific primers. The key conditions in PCR process, the optimal cycle number and Mg^2＋ concentration, are determined by studying the linear range of PCR amplification products. The integrated optical density （IOD） is showed by scanning the eletrophoresis bands of PCR products with VDS system. The relative concentration of cGPX mRNA can be calculated by the IOD ratio of cGPX and 18S rRNA.

关 键 词：RT-PCR cGPX 猪 肌肉 

分 类 号：Q503[生物学—生物化学]