摘要
为筛选最佳牛支原体(mycoplasma bovis,Mb)膜蛋白的提取工艺,本试验以牛支原体W70株为代表,分别比较了4种浓度的TritonX-100、TritonX-114、TritonX-100和TritonX-114;3种浓度碘化钾(KI)复合提取;反复冻融和超声裂解等6种Mb膜蛋白提取工艺和参数。各方法所获膜蛋白经SDS-PAGE和Bradford定量分析表明:不同方法的提取物中蛋白多肽和蛋白浓度各异;其中以3%TritonX-114和0.4%TritonX-100结合0.6M KI两法提取的蛋白条带清晰度较好,蛋白浓度及蛋白收集率较高,蛋白多肽条数24～28条,蛋白相对分子质量分别在170 000～12 200和167 000～11 400之间;超声裂解及反复冻融法提取效果一般;而1.8 M KI法效果最差。研究发现3%TritonX-114或0.4%TritonX-100结合0.6M KI是Mb膜蛋白较优的提取方法。
For choos!ng and comparing extraction process and parameters of membrane proteins of ycoplasma boris （Mb）. In the study, taking W70 strain for representative strains, six kinds of mbrane protein extraction methods of Mb were compared, which include four concentrations of ＂itonX-100 and TritonX-114 extraction alone, TritonX-100 and TritonX-114 combine with three ncentrations of KI, the repeated freezing and ultrasonication extraction and so on. Membrane oteins obtained from each method were analyzed by SDS-PAGE and bradford quantitative meth- The results showed that there were different kinds of p eptides and concentrations and the thod with 3% TritonX-114 or 0.4 TritonXq00 combined with 0.6M KI were much better, e clarity of protein bands was better, the protein concentration and protein collection rate were ,herthere were 24-28 peptide bands and protein relative moleculer mass were among 170 000-- 200 and 167 000--11 400,respectively. The method with repeated freezing and ultrasonication ts not very good,and the worst effects was 1.8 M KI methods. Therefore, that the better mem- ane protein extraction methods of Mb was with 3% TritonX-114 or 0.4% TritonX-100 combine th 0.6M KI.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2013年第6期855-860,共6页
Chinese Journal of Veterinary Science
基金
重庆市科委重大攻关资助项目(CSTC
2010AA1021)
