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首页> 外文期刊>African Journal of Biotechnology >Development of a competitive PCR assay for the quantification of total Escherichia coli DNA in water
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Development of a competitive PCR assay for the quantification of total Escherichia coli DNA in water

机译：研发竞争性PCR分析方法，用于定量分析水中的总大肠杆菌DNA

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Standard health-related microbial water testing relies on the culturability ofEscherichia coli?(E. coli) to?estimate their numbers. Competitive PCR (c-PCR) offers the potential to estimate the?E. coli?level of a water source without culturing. The aim was to investigate the use of c-PCR reaction to detect and quantify, without prior enrichment,?Escherichia coli?in water samples. The?E. coli?malate dehydrogenase?Mdh?house-keeping gene was modified and used as an internal control and competitor DNA for the c-PCR.?E. coli?cell concentration equivalents ranging from 20 to 2 x 104?cells ml-1?could be quantified with the c-PCR. Fifty-three water samples from various sources were tested with the DNA extraction and c-PCR protocol. Due to PCR inhibition?E. coli?Mdh?gene copies could only be determined for 20 of the 53 samples (38%).?Of the 20 samples tested 15% gave comparable results for competitive PCR and culturable?E. coli?numbers; 55% obtained higher values with competitive PCR and 30% obtained higher values with the culture based experiments.?The c-PCR successfully estimated?E.?coli?numbers that gave comparable results with the culture based microbiological data obtained.
机译：标准的与健康相关的微生物水测试依赖于大肠杆菌的可培养性来估计其数量。竞争PCR（c-PCR）提供了估计ΔE的潜力。大肠杆菌水平，无需培养。目的是研究使用c-PCR反应来检测和定量水样品中的大肠杆菌而无需事先富集。 ？E。大肠杆菌？苹果酸脱氢酶？Mdh？持家基因被修饰并用作c-PCR的内部对照和竞争者DNA。大肠杆菌细胞的浓度当量范围从20到2 x 104？细胞ml-1 ?，可以通过c-PCR进行定量。使用DNA提取和c-PCR方案对来自各种来源的53个水样进行了测试。由于PCR抑制作用？只能在53个样品中的20个（38％）中确定大肠杆菌的Mdh基因拷贝。在20个样品中，有15％的PCR和可培养E值可比。大肠杆菌数量; 55％的竞争性PCR获得了更高的值，而30％的基于培养物的实验获得了更高的值。c-PCR成功地估算了大肠杆菌的大肠杆菌数，该数值与获得的基于培养物的微生物学数据具有可比性。

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