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KMID : 0578320060220030336
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Molecules and Cells
2006 Volume.22 No. 3 p.336 ~ p.342
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Determination of Neurotoxin Gene Expression in Clostridium botulinum Type A by Quantitative RT-PCR
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Shin Na-Ri
Shin Ji-Hun
Chun Jeong-Hoon
Yoon So-Yeon
Kim Bong-Su
Oh Hee-Bok
Rhie Gi-Eun
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Abstract
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Real time reverse transcription (RT)-PCR was used to quantify the expression of the botulinum neurotoxin type A (BoNT/A) gene (cntA) by normalization with the expression of 16S rRNA. The method were confirmed by monitoring the mRNA levels of cntA during growth in five type A strains. In all but one of the strains the expression of cntA mRNA was maximal in the late exponential phase, and approximately 35-fold greater than in the early exponential phase. The concentration of the extracellular BoNT/A complex detected by ELISA was highest in stationary phase. Sodium nitrite and sorbic acid completely inhibited growth at 20 ppm and 4 mg ml-1, respectively. CntA expression became lower in proportion to the concentration of sorbic acid, and this reduction was confirmed by mouse bioassay. Our results show that real time RT-PCR can be used to quantify levels of C. botulinum type A neurotoxin transcripts and to assess the effects of food additives on botulinal risk.
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KEYWORD
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Botulinum Neurotoxin Type A Gene, Clostridium botulinum, RT-PCR, Sodium Nitrite, Sorbic Acid
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