| Comparative Quantification of LacZ (β-galactosidase) Gene from a Pure
Cultured Escherichia coli K-12 |
| Ji-sun Han, and Chang-gyun Kim† |
| Department of Environmental Engineering, Inha University, Incheon, South Korea |
Corresponding Author:
Chang-gyun Kim ,Tel: +82-32-860-7561, Fax: +82-32-865-1425, Email: cgk@inha@ac.kr
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Received: June 14, 2008; Accepted: January 17, 2009. |
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| ABSTRACT |
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Escherichia coli K-12 (E. coli K-12) is a representative indicator globally used for distinguishing and monitoring dynamic fates of pathogenic
microorganisms in the environment. This study investigated how to most critically quantify lacZ (β-galactosidase) gene in E. coli K-12 by two
different real-time polymerase chain reaction (real-time PCR) in association with three different DNA extraction practices. Three DNA extractions,
i.e., sodium dodecyl sulfate (SDS)/proteinase K, magnetic beads and guanidium thiocyanate (GTC)/silica matrix were each compared for extracting
total genomic DNA from E. coli K-12. Among them, GTC/silica matrix and magnetic beads beating similarly worked out to have the highest (22-23
ng/μL) concentration of DNA extracted, but employing SDS/proteinase K had the lowest (10 ng/μL) concentration of DNA retrieved. There were
no significant differences in the quantification of the copy numbers of lacZ gene between SYBR Green I qPCR and QProbe-qPCR. However,
SYBR Green I qPCR obtained somewhat higher copy number as 1×108 copies. It was decided that GTC/silica matrix extraction or magnetic beads
beating in combination with SYBR Green I qPCR can be preferably applied for more effectively quantifying specific gene from a pure culture of
microorganism. |
| Keywords:
E. coli K-12 | DNA extraction | Real-time PCR | QProbe-qPCR | SYBR Green I qPCR |
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