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聚合酶链式反应快速检测E型肉毒神经毒素基因 |
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王兴民
作者单位： 730046 卫生部兰州生物制品研究所第五研究室
本课题受甘肃省中青年科研基金资助， 项目号为ZQ-94-024
【 摘要 】目的 E型肉毒中毒是人类肉毒中毒的主要型别之一，本试验旨在建立用于Ｅ型肉毒
梭菌鉴定的PCR方法。 方法 用人工合成的寡核苷酸引物扩增E型肉毒神经毒素基因的一段3
68bp的DNA片段，快速检测E型肉毒神经毒素基因，对梭状芽胞杆菌属的40株保藏菌株及3
3份土壤标本进行了鉴定，用E型肉毒梭菌CMCC(B)64501对其灵敏度进行了检查。 结果 从
所有E型神经毒素原性菌株或标本均能扩增出目的片段，且能用特定的限制性内切酶切成相应
的片段。其它菌株均未能扩增出目的片段。灵敏度试验可从30个菌体扩增出目的片段。结论
此方法用于E型神经毒素原性梭菌的鉴定具有较高的灵敏度及特异性。
【 主题词 】 E型肉毒神经毒素 基因 聚合酶链反应
Rapid detection of type E botulinal neurotoxin gene by polymerase chai
n reaction Wang Xingmin,Meng Xiaoqi,Zou Kaiyong,et al.Department o
f Anaerobic Bacteriology,Lanzhou Institute of Biological Products,
Ministry of Public Health,Lanzhou 730046
【 Abstract 】 Objective Type E botulism is one of main types of botulism
in human beings.A polymerase chain reaction (PCR) method was establish
ed for rapid detection of type E botulinal neurotoxin gene and identifi
cation of Clostridium botulinum type E. Methods The PCR was developed
by using a pair of oligonucleotide primers which were designed to ampli
fy a fragment of 368bp on the heavy chain of type E neurotoxin gene.Forty
strains belonging to 10 clostridial species and thirty-three soil samp
les were detected using the PCR.The sensitivity of the PCR was determin
ed by using Clostridium botulinum strain CMCC(B)64501. Results All
of type E neurotoxigenic strains and samples whose cultures containin
g type E neurotoxin were positive but the others were negative.The rest
riction endonuclease digestion patterns of amplified products were i
n conformity with the sizes estimated from the sequence data.In test of
sensitivity,target DNA fragment could be amplified from 30 CFU by the P
CR. Conclusion The PCR is specific and sensitive for detection and ide
ntification of type E neurotoxigenic clostridial species.
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