On the fidelity of mRNA translation in the nuclease-treated rabbit reticulocyte lysate system.
AUTOR(ES)
Dasso, M C
RESUMO
As a test of the fidelity of the rabbit reticulocyte lysate system, we have examined the products of translation of various different influenza virus mRNAs, produced by in vitro transcription. A common finding with all mRNA species was that the ratio of full-length translation product to incomplete products decreased with increasing mRNA concentration. These short products are a mixture of (i) polypeptides initiated at the authentic initiation site but terminated prematurely, and (ii) polypeptides initiated at internal sites and terminated at the correct site. Analysis of mRNA stability during the translation assay showed very little degradation, quite insufficient to be the principle cause of incomplete product synthesis. Investigation of the influence of various parameters on the ratio of full-length to incomplete products leads to the conclusion that a high fidelity of translation can be obtained provided certain precautions are followed: the use of capped, rather than uncapped, mRNAs at low concentrations, with KCl concentrations about 20 mM above the level that gives maximum incorporation.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=317719Documentos Relacionados
- Translation of HTLV (human T-cell leukemia virus) RNA in a nuclease-treated rabbit reticulocyte system.
- Reconstitution of functional mRNA-protein complexes in a rabbit reticulocyte cell-free translation system.
- Proteins associated with rabbit reticulocyte mRNA caps during translation as investigated by photocrosslinking.
- Low ionic strength extraction of nuclease-treated nuclei destroys the attachment of transcriptionally active DNA to the nuclear skeleton.
- Unwinding protein specific for mRNA translation fractionated together with rabbit reticulocyte initiation factor 3 complex
