PAN Zhuo

doi:10.3967/bes2016.030

Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus

文章导航 > Biomedical and Environmental Sciences > 2016 > 29(3): 233-238

PAN Zhuo. Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus[J]. Biomedical and Environmental Sciences, 2016, 29(3): 233-238. doi: 10.3967/bes2016.030

Citation:

PAN Zhuo. Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus[J]. Biomedical and Environmental Sciences, 2016, 29(3): 233-238. doi: 10.3967/bes2016.030

doi: 10.3967/bes2016.030

基金项目: This study was supported by the National Special Fund for the Development of Major Research Equipment and Instruments (2012YQ09019706)from the Ministry of Science and Technology, and by the Priority Project on Infectious Disease Control and Prevention (2012ZX10004215 and 2008ZX10004-008) from the National Health and Family Planning Commission of the Peoples Republic of China

Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus

PAN Zhuo

Microbiology Laboratory, Shijiazhuang Center for Disease Control and Prevention, Shijiazhuang 050016, Hebei, China

Funds: This study was supported by the National Special Fund for the Development of Major Research Equipment and Instruments (2012YQ09019706)from the Ministry of Science and Technology, and by the Priority Project on Infectious Disease Control and Prevention (2012ZX10004215 and 2008ZX10004-008) from the National Health and Family Planning Commission of the Peoples Republic of China

摘要: In order to develop a rapid and reliable method forB. cereusgenotyping, factors influencing PFGE results, including preparation of bacterial cells embedded in agarose, lysis of embedded cells, enzymatic digestion of intact genomic DNA, and electrophoresis parameters allowing for reproducible and meaningful DNA fragment separation, were controlled. Optimal cellular growth (Luria-Bertani agar plates for 12-18 h) and lysis conditions (4 h incubation with 500 μg/mL lysozyme) produced sharp bands on the gel. Restriction enzymeNotI was chosen as the most suitable. Twenty-two isolates were analyzed by NotI digestion, using three electrophoretic parameters (EPs). The EP-a was optimal for distinguishing between isolates. The optimized protocol could be completed within 40 h which is a significant improvement over the previous methods.
- /
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- Bacillus cereus /
- DNA fragment /
- genomic DNA

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Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus

doi: 10.3967/bes2016.030

PAN Zhuo

刊出日期: 2016-03-20

关键词:

Bacillus cereus /

DNA fragment /

genomic DNA

摘要: In order to develop a rapid and reliable method forB. cereusgenotyping, factors influencing PFGE results, including preparation of bacterial cells embedded in agarose, lysis of embedded cells, enzymatic digestion of intact genomic DNA, and electrophoresis parameters allowing for reproducible and meaningful DNA fragment separation, were controlled. Optimal cellular growth (Luria-Bertani agar plates for 12-18 h) and lysis conditions (4 h incubation with 500 μg/mL lysozyme) produced sharp bands on the gel. Restriction enzymeNotI was chosen as the most suitable. Twenty-two isolates were analyzed by NotI digestion, using three electrophoretic parameters (EPs). The EP-a was optimal for distinguishing between isolates. The optimized protocol could be completed within 40 h which is a significant improvement over the previous methods.

English Abstract

PAN Zhuo. Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus[J]. Biomedical and Environmental Sciences, 2016, 29(3): 233-238. doi: 10.3967/bes2016.030

Citation:

PAN Zhuo. Optimization of Pulsed-field Gel Electrophoresis Procedure for Bacillus cereus[J]. Biomedical and Environmental Sciences, 2016, 29(3): 233-238. doi: 10.3967/bes2016.030