Restriction Enzyme Pattern Analysis of Mycobacteria DNA by Capillary Electrophoresis with Laser-induced Fluorescence Detection

• 来源期刊: 中国高等学校学术文摘·化学
• 关键词: capillary electrophoresis laser-induced fluorescence detection restriction enzyme pattern Mycobacterium
• 年，卷（期）: 2006,（1）
• 页码范围: 27-33
• 页数: 7页
• 语种: 英文

摘要

A new method for rapidly detecting restriction enzyme patterns of Mycobacterium DNA using capillary electrophoresis with laser-induced fluorescence detection (CE-LIFD)was developed.Polymerase chain reaction was used to amplify a 439-bp fragment of a 65,000-kDa(Mr)heat shock protein gene(hsp65)of Mycobacterium.After digesting amplification products by BstEII and HaeIII,patterns of enzyme cleavage products were detected by both CE-LIFD and agarose gel electrophoresis(AGE),respectively.Experimental parameters of CE were optimized.Restriction enzyme patterns of Mycobacterium DNA were detected in optimum electrophoresis conditions:a coated capillary column with a length of 50 cm and an internal diameter of 100 μm,an electrophoresis buffer of 45 mmol/1 Tris-boric acid-ethylenediaminetetraacetic acid,and a running voltage of 11 kV.The restriction enzyme patterns for eight species of mycobacteria were studied.Relative standard deviations of the relative migration times of DNA segments were＜3.6%.Compared with AGE,CE is more outstanding in resolution and detection time,and it can be applied as a more effective means to DNA restriction enzyme pattern analysis.