Western blot detection of PMI protein in transgenic rice  

作　　者：RONG Rui-juan WU Peng-cheng LAN Jin-ping WEI Han-fu WEI Jian CHEN Hao SHI Jia-nan HAO Yu-jie LIU Li-juan DOU Shi-juan LI Li-yun WU Lin LIU Si-qi YIN Chang-cheng LIU Guo-zhen 

机构地区：[1]College of Life Sciences, Hebei Agricultural University, Baoding 071001, P.R. China [2]Beijing Protein Innovation Co. Ltd., Beijing 101318, P.R.China [3]Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, P.R. China

出　　处：《Journal of Integrative Agriculture》2016年第4期726-734,共9页农业科学学报（英文版）

基　　金：supported in part by the Natural Science Foundation of Beijing, China (5121001);the Cultivate New Varieties of Genetically Modified Organisms Technology Major Projects, the Ministry of Science and Technology of China (2009ZX08012-006B)

摘　　要：Phosphomannose isomerase （PMI） encoding gene manA is a desirable selective marker in transgenic research. Under- standing of its expression patterns in transgenic plant and establishing highly sensitive detection method based on immunoassay have great impacts on the application of PMI. In this study, PMI-specific monoclonal antibodies were generated using recombinant protein as immunogen, and could be used in Western blot to detect as little as 0.5 ng His-tagged PMI protein or rice expressed PMI protein in sample accounted for 0.4% of single rice grain （about 0.08 mg）. PMI protein driven by CaMV-35S promoter was detected in dozens of tested tissues, including root, stem, leaf, panicle, and seed at all developmental stages during rice growing, and PMI protein accounted for about 0.036% of total protein in the leaves at seedling stage. The established method potentially can be used to monitor PMI protein in rice grains.Phosphomannose isomerase （PMI） encoding gene manA is a desirable selective marker in transgenic research. Under- standing of its expression patterns in transgenic plant and establishing highly sensitive detection method based on immunoassay have great impacts on the application of PMI. In this study, PMI-specific monoclonal antibodies were generated using recombinant protein as immunogen, and could be used in Western blot to detect as little as 0.5 ng His-tagged PMI protein or rice expressed PMI protein in sample accounted for 0.4% of single rice grain （about 0.08 mg）. PMI protein driven by CaMV-35S promoter was detected in dozens of tested tissues, including root, stem, leaf, panicle, and seed at all developmental stages during rice growing, and PMI protein accounted for about 0.036% of total protein in the leaves at seedling stage. The established method potentially can be used to monitor PMI protein in rice grains.

关 键 词：transgenic rice protein expression CaMV-35S promoter phosphomannose isomerase （PMI） Western blot 

分 类 号：S511[农业科学—作物学] Q786[生物学—分子生物学]