不同样本处理方法对显色法芯片检测结核分枝杆菌利福平和异烟肼耐药基因的影响  

Effect of Varied Sample Treatment Procedures on Detection of the Resistance Genes of Rifampin and Isoniazid in Mycobacterium Tuberculosis DNA by Using Coloration Genes Chips

作　　者：王万相 郭乃洲 严爱华 马力 周步全 马达 景奉香[3] 唐向荣[3] 庄严[4] 

机构地区：[1]盐城市第一人民医院,江苏盐城224006 [2]盐城市第二人民医院,江苏盐城224002 [3]中国科学院上海微系统与信息技术研究所,上海200050 [4]江苏大学京江学院,江苏镇江212013

出　　处：《实用医技杂志》2006年第21期3742-3745,共4页Journal of Practical Medical Techniques

基　　金：盐城市科技局;卫生局科技发展计划项目(编号YK2005105)

摘　　要：目的:选择一种较为理想的从痰标本中直接抽提结核分枝杆菌DNA的方法,用于显色法芯片检测结核分枝杆菌利福平(RFP)和异烟肼(INH)的耐药基因。方法:选择3份含结核分枝杆菌分别为4.325×103、6.857×104、5.356×105Copies/ml的痰液,分别用Chelex100树脂法、煮沸法、蛋白酶K消化-异丙醇沉淀法、蛋白酶K消化-酚/氯仿抽提法、碱裂解法、异硫氰酸胍裂解法、盐酸胍裂解法、离心柱法抽提结核分枝杆菌DNA作多重聚合酶链反应,显色法芯片检测结核分枝杆菌RFP和INH耐药基因。结果:离心柱法检测结果阳性,其余方法检测结果均为阴性,离心柱法检测灵敏度为2.743×103Copies/ml。结论:离心柱法是一种较为理想的从痰标本中直接抽提结核分枝杆菌DNA的方法,可用于显色法芯片检测结核分枝杆菌RFP和INH耐药基因。Objective To choose a more ideal method to directly extract My cobacterium tuberculosis DNA from sputum sample to detect the resistance genes of rifampin （RFP） and isoniazid （INH） in Mycobaeterium tuberculosis by using Coloration gene chips. Methods Choose three sputum samples whose Myeobacterium tuberculosis respectively were 4. 325 × 10^3 ,6. 857 × 10^4 and 5. 356 × 10^5 Copies/ml. Respectively use chelex 100 resinate, boiling extraction, proteinase K digest/isopropanol precipitation, proteinase K digest/phenoL/ehloroformextra etion, alkaline lysis, guanidine thiocyanate lysis, guanidine hydrochloride lysis, spin eolumr/method to extract Myeobaeterium tuberculosis DNA for mulfiple-polymerase chain eraction. Results The result of spin column method was positive. Others were negative. The sensitivity of spin column method was 2. 743 × 10^3 Copies/ml. Conclusion The spin column is a more ideal method to extract Myeobacterium tuberculosis DNA from sputum sample, which can be used to detect resistance genes of RFP and INH in Mycobacterium tuberculosis by using coloration gene chips.

关 键 词：痰标本 结核分枝杆菌DNA 抽提 显色法 耐药基因 基因芯片 

分 类 号：R450[医药卫生—治疗学]