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基于重复 DNA 指纹的大肠杆菌 SCAR 引物的开发。

Development of SCAR primers based on a repetitive DNA fingerprint for Escherichia coli detection.

机构信息

Department of Biology, Faculty of Science, Mahasarakham University, Mahasarakham 44150, Thailand.

出版信息

J Microbiol. 2013 Feb;51(1):31-5. doi: 10.1007/s12275-013-2244-4. Epub 2013 Mar 2.

Abstract

The present study aimed to use enterobacterial repetitive intergenic consensus (ERIC) fingerprints to design SCAR primers for the detection of Escherichia coli. The E. coli strains were isolated from various water sources. The primary presumptive identification of E. coli was achieved using MacConkey agar. Nineteen isolates were selected and confirmed to be E. coli strains based on seven biochemical characteristics. ERIC-PCR with ERIC 1R and ERIC 2 primers were used to generate DNA fingerprints. ERIC-PCR DNA profiles showed variant DNA profiles among the tested E. coli strains and distinguished all E. coli strains from the other tested bacterial strains. A 350 bp band that predominated in five E. coli strains was used for the development of the species-specific SCAR primers EC-F1 and EC-R1. The primers showed good specificity for E. coli, with the exception of a single false positive reaction with Sh. flexneri DMST 4423. The primers were able to detect 50 pg and 10(0) CFU/ml of genomic DNA and cells of E. coli, respectively.

摘要

本研究旨在利用肠杆菌基因间重复一致序列(ERIC)指纹图谱设计 SCAR 引物,用于检测大肠杆菌。从各种水源中分离出大肠杆菌菌株。使用麦康凯琼脂初步推定鉴定大肠杆菌。根据七项生化特征,选择了 19 个分离株并确认其为大肠杆菌株。使用 ERIC 1R 和 ERIC 2 引物进行 ERIC-PCR 以生成 DNA 指纹图谱。ERIC-PCR DNA 图谱显示,测试的大肠杆菌菌株之间存在不同的 DNA 图谱,并将所有大肠杆菌菌株与其他测试的细菌菌株区分开来。在五个大肠杆菌菌株中占主导地位的 350bp 带被用于开发种特异性 SCAR 引物 EC-F1 和 EC-R1。引物对大肠杆菌具有良好的特异性,除与 Shigella flexneri DMST 4423 有一个假阳性反应外。引物能够分别检测到 50pg 和 10(0)CFU/ml 的大肠杆菌基因组 DNA 和细胞。

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