摘要
蛋白质分离纯化技术是生化及分子生物学实验技术的重要组成部分,在本科实验教学中占有重要地位。为了探索合适的实验方法和实验条件,优化本科实验教学中的溶菌酶分离纯化实验,以获得更好的纯化结果和教学效果,该文使用离子交换层析、硫铵沉淀、分子筛层析等方法成功地从蛋清中纯化出高纯度的溶菌酶,用SDS-聚丙烯酰胺凝胶电泳方法鉴定了样品纯度,并用溶壁微球菌做为底物测定了纯化酶的活力及比活力。结果显示,使用改进后的方法能够得到较高纯度和比活力的溶菌酶,整个实验重复性好、易于操作,适合作为本科教学实验项目。
Protein isolation and purification technology is one of the basic technologies in biochemical and molecular biology studies,which also is an important part of experiment teaching at the undergraduate level.In order to develop an efficient protein separation and purification scheme to establish a more optimal experimental teaching model for undergraduate students,the ion-exchange column chromatography,ammonium sulphate precipitation and gel filtration chromatography are applied to purify lysozyme from egg white step by step.After each step of purification procedure,the fractions obtained are analyzed by SDS-polyacrylamide gel electrophoresis to assess protein purity.The activity and specific vitality of purified enzyme are determined by using micrococcus as substrate.The results show that the lysozyme with high purity and specific vitality can be obtained by using the improved method.The whole experiment has good repeatability and is easy to operate.It is suitable for undergraduate teaching experiment.
作者
刘奕君
赵宇煊
高松民
王雅
郝帅林
杨志伟
陈志玲
刘晓晴
王海龙
LIU Yijun;ZHAO Yuxuan;GAO Songmin;WANG Ya;HAO Shuailin;YANG Zhiwei;CHEN Zhiling;LIU Xiaoqing;WANG Hailong(College of Life Science,Capital Normal University,Beijing 100048,China)
出处
《实验技术与管理》
CAS
北大核心
2020年第8期200-203,共4页
Experimental Technology and Management
基金
国家重大科学研究计划“DNA损伤修复蛋白的作用机制”(2015CB910602)
北京市自然科学基金面上项目“CtIP在MicroRNA加工及肿瘤转移中的功能研究”(5182003)。
关键词
溶菌酶
蛋白质分离纯化技术
实验教学
lysozyme
protein separation and purification technology
experimental teaching
