[Establishment and application of a standard IS6110-RFLP method in the study of molecular genotyping analysis on Mycobacterium tuberculosis].
OBJECTIVE
To develop a standardized IS6110-restriction fragment length polymorphism (RFLP) method, used for evaluating the capacity of genotyping.


METHODS
IS6110-RFLP of 78 Mycobacterium (M.) tuberculosis strains were studied by bio-molecular techniques including DNA isolation, PCR, restriction endonuclease enzyme analysis, southern blotting, agarose gel electrophoresis, together with data analysis by software Gel-Pro analyzer 3.1 and BioNumerics (Version 5.0).


RESULTS
IS6110-RFLP method was established and standardized successfully, including DNA isolation, PCR, restriction endonuclease enzyme analysis, southern blotting, agarose gel electrophoresis and usage of the analysis software with standard parameters. By this method, 78 M. tuberculosis isolates were classified into 75 genotypes which belonged to 11 different clusters. Of all the isolates, 66.7% (52/78) belonged to a main cluster.


CONCLUSIONS
Standard IS6110-RFLP method was established successfully. This method had powerful capacity for genotyping and strain level identification and could be used for the surveillance on pathogens of M. tuberculosis in China.
Liu
Jing-hua
JH
State Key Laboratory for Infectious Diseases Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China.
Kremer
Kristin
K
Pourcel
Christine
C
Mulder
Arnout
A
Liu
Zhi-guang
ZG
Zhao
Xiu-qin
XQ
Wan
Kang-lin
KL
D015373:Bacterial Typing Techniques / Q000379:methods; D004269:DNA, Bacterial; D016680:Genome, Bacterial; D005838:Genotype; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D012150:Polymorphism, Restriction Fragment Length
D004269:DNA, Bacterial