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Open AccessJournal ArticleDOI

Eucaryotic RNA polymerase conditional mutant that rapidly ceases mRNA synthesis.

Michael L. Nonet, +3 more
- 01 May 1987 - 
- Vol. 7, Iss: 5, pp 1602-1611
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TLDR
The isolation of a yeast conditional mutant which rapidly ceases synthesis of mRNA when subjected to the nonpermissive temperature indicates that the mutant possesses a functionally defective, rather than an assembly-defective, RNA polymerase II.
Abstract
We have isolated a yeast conditional mutant which rapidly ceases synthesis of mRNA when subjected to the nonpermissive temperature. This mutant (rpb1-1) was constructed by replacing the wild-type chromosomal copy of the gene encoding the largest subunit of RNA polymerase II with one mutagenized in vitro. The rapid cessation of mRNA synthesis in vivo and the lack of RNA polymerase II activity in crude extracts indicate that the mutant possesses a functionally defective, rather than an assembly-defective, RNA polymerase II. The shutdown in mRNA synthesis in the rpb1-1 mutant has pleiotropic effects on the synthesis of other RNAs and on the heat shock response. This mutant provides direct evidence that the RPB1 protein has a functional role in mRNA synthesis.

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Dissecting the regulatory circuitry of a eukaryotic genome

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Cryptic Pol II Transcripts Are Degraded by a Nuclear Quality Control Pathway Involving a New Poly(A) Polymerase

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Codon Optimality Is a Major Determinant of mRNA Stability

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Precision and functional specificity in mRNA decay

TL;DR: No simple correlation between mRNA half-lives and ORF size, codon bias, ribosome density, or abundance is found, but the decay rates of mRNAs encoding groups of proteins that act together in stoichiometric complexes were generally closely matched.
References
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Journal ArticleDOI

A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding

TL;DR: This assay is very reproducible and rapid with the dye binding process virtually complete in approximately 2 min with good color stability for 1 hr with little or no interference from cations such as sodium or potassium nor from carbohydrates such as sucrose.
Book

Molecular Cloning: A Laboratory Manual

TL;DR: Molecular Cloning has served as the foundation of technical expertise in labs worldwide for 30 years as mentioned in this paper and has been so popular, or so influential, that no other manual has been more widely used and influential.
Journal ArticleDOI

Screening lambdagt recombinant clones by hybridization to single plaques in situ

TL;DR: A rapid, direct method for screening single plaques of Agt recombinant phage is described, which allows at least 10(6) clones to be screened per day and simplifies physical containment of recombinants.
Journal ArticleDOI

A positive selection for mutants lacking orotidine-5'-phosphate decarboxylase activity in yeast: 5-fluoro-orotic acid resistance.

TL;DR: Mutations at the URA3 locus of Saccharomyces cerevisiae can be obtained by a positive selection, based on the loss of orotidine-5′-phosphate decarboxylase activity, and seems applicable to a variety of eucaryotic and procaryotic cells.
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