Decay of mRNA in Escherichia coli: investigation of the fate of specific segments of transcripts.
Decay of mRNA in Escherichia coli: investigation of the fate of specific segments of transcripts.
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DOI：
10.1073/pnas.80.3.653
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发表时间：
1983-02
影响因子：
11.1
通讯作者：
A. von Gabain;J. Belasco;J. Schottel;A. C. Chang;S. Cohen
中科院分区：
文献类型：
--
作者：
A. von Gabain;J. Belasco;J. Schottel;A. C. Chang;S. Cohen
建立了一种检测大肠杆菌中β-内酰胺酶（bla）和ompA基因转录物特定片段的方法。用能够切割单链DNA的核酸内切酶处理在噬菌体M13中克隆的DNA探针，将产生的片段与总细胞RNA退火，并将所得RNA。DNA杂交体进行S1核酸酶处理和凝胶分级分离。通过使用该测定，获得了ompA基因编码的长寿命mRNA衰变中3 '至5'方向性的直接证据，并且对于翻译的mRNA片段与未翻译的mRNA片段没有观察到优先稳定性。在bla mRNA的情况下，全长转录物的初始切割是限速的，并且没有检测到衰变中间体。对于具有变体3'或5'末端的bla转录物，未观察到降解速率的差异。
An assay was developed to investigate the fate of specific segments of beta-lactamase (bla) and ompA gene transcripts in Escherichia coli. DNA probes cloned in bacteriophage M13 were treated with an endonuclease capable of cleaving single-stranded DNA, the fragments produced were annealed with total cellular RNA, and the resulting RNA . DNA hybrids were subjected to S1 nuclease treatment and gel fractionation. By using this assay, direct evidence was obtained for 3'-to-5' directionality in the decay of the long-lived mRNA encoded by the ompA gene, and no preferential stability was observed for translated versus untranslated mRNA segments. In the case of bla mRNA, initial cleavage of the full-length transcript was rate limiting, and no decay intermediates were detected. No difference in degradation rate was seen for bla transcripts having variant 3' or 5' termini.