通过逆转录-聚合酶链反应检测mRNA作为大肠杆菌细胞活力的指标。
Detection of mRNA by reverse transcription-PCR as an indicator of viability in Escherichia coli cells.
作者信息
Sheridan G E, Masters C I, Shallcross J A, MacKey B M
机构信息
Institute of Food Research, Reading, United Kingdom.
出版信息
Appl Environ Microbiol. 1998 Apr;64(4):1313-8. doi: 10.1128/AEM.64.4.1313-1318.1998.
Abstract
The relationship between the detection of mRNA and cellular viability in Escherichia coli was investigated in cells killed by heat or ethanol. Reverse transcription-PCR (RT-PCR) methods were developed for detecting mRNA from rpoH, groEL, and tufA genes. mRNA from all three genes was detected immediately after the cells had been killed by heat or ethanol but gradually disappeared with time when dead cells were held at room temperature. In heat-killed cells, some mRNA targets became undetectable after 2 to 16 h, whereas after ethanol treatment, mRNA was still detected after 16 h. In contrast, 16S rRNA was detected by RT-PCR in all samples containing dead cells and did not disappear during a subsequent incubation of 16 h at room temperature. Of the different types of nucleic acid, mRNA is the most promising candidate for an indicator of viability in bacteria, but its persistence in dead cells depends on the inactivating treatment and subsequent holding conditions.
摘要
研究了热或乙醇处理致死的大肠杆菌中mRNA检测与细胞活力之间的关系。开发了逆转录聚合酶链反应（RT-PCR）方法来检测来自rpoH、groEL和tufA基因的mRNA。在细胞被热或乙醇杀死后，立即检测到来自所有三个基因的mRNA，但当死细胞在室温下放置时，mRNA会随着时间逐渐消失。在热致死细胞中，一些mRNA靶点在2至16小时后无法检测到，而乙醇处理后，16小时后仍能检测到mRNA。相比之下，在所有含有死细胞的样品中，通过RT-PCR都能检测到16S rRNA，并且在随后室温下16小时的孵育过程中不会消失。在不同类型的核酸中，mRNA是细菌活力指标最有前景的候选者，但其在死细胞中的持久性取决于灭活处理和随后的保存条件。
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本文引用的文献
1
Improved Method for Recovery of mRNA from Aquatic Samples and Its Application to Detection of mer Expression.从水生样本中提取 mRNA 的改良方法及其在 mer 表达检测中的应用。
Appl Environ Microbiol. 1994 Jun;60(6):1814-21. doi: 10.1128/aem.60.6.1814-1821.1994.
2
Viability and isolation of marine bacteria by dilution culture: theory, procedures, and initial results.海洋细菌的稀释培养法的生存力和隔离：理论、程序和初步结果。
Appl Environ Microbiol. 1993 Mar;59(3):881-91. doi: 10.1128/aem.59.3.881-891.1993.
3
Gene expression per gene dose, a specific measure of gene expression in aquatic microorganisms.每个基因剂量的基因表达，一种水生微生物中特定的基因表达衡量标准。
Appl Environ Microbiol. 1993 Feb;59(2):451-7. doi: 10.1128/aem.59.2.451-457.1993.
4
Effects of Hg, CH(3)-Hg, and Temperature on the Expression of Mercury Resistance Genes in Environmental Bacteria.汞、甲基汞和温度对环境细菌中汞抗性基因表达的影响。
Appl Environ Microbiol. 1990 Nov;56(11):3266-72. doi: 10.1128/aem.56.11.3266-3272.1990.
5
Sensitive detection of viable Listeria monocytogenes by reverse transcription-PCR.通过逆转录聚合酶链反应灵敏检测存活的单核细胞增生李斯特菌
Appl Environ Microbiol. 1997 Nov;63(11):4441-8. doi: 10.1128/aem.63.11.4441-4448.1997.
6
An effective method of RNA extraction from bacteria refractory to disruption, including mycobacteria.一种从难以破碎的细菌（包括分枝杆菌）中提取RNA的有效方法。
Nucleic Acids Res. 1997 Feb 1;25(3):675-6. doi: 10.1093/nar/25.3.675.
7
Detection of viable Vibrio cholerae by reverse-transcriptase polymerase chain reaction (RT-PCR).通过逆转录聚合酶链反应（RT-PCR）检测霍乱弧菌活菌。
Mol Biotechnol. 1996 Feb;5(1):1-10. doi: 10.1007/BF02762407.
8
Probes and polymerase chain reaction for detection of food-borne bacterial pathogens.用于检测食源性病原体的探针和聚合酶链反应
Int J Food Microbiol. 1995 Nov;28(1):1-78. doi: 10.1016/0168-1605(94)00159-4.
9
In situ PCR for visualization of microscale distribution of specific genes and gene products in prokaryotic communities.原位聚合酶链反应用于可视化原核生物群落中特定基因和基因产物的微观分布。
Appl Environ Microbiol. 1995 Nov;61(11):4074-82. doi: 10.1128/aem.61.11.4074-4082.1995.
10
Application of reverse transcriptase PCR for monitoring expression of the catabolic dmpN gene in a phenol-degrading sequencing batch reactor.逆转录聚合酶链反应在监测酚降解序批式反应器中分解代谢dmpN基因表达方面的应用。
Appl Environ Microbiol. 1995 Nov;61(11):3981-5. doi: 10.1128/aem.61.11.3981-3985.1995.
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