Linked Life Data

7599645

Source:http://linkedlifedata.com/resource/pubmed/id/7599645

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
6
pubmed:dateCreated
1995-8-8
pubmed:abstractText
A convenient in vitro transcription system using monocot and dicot whole cell extracts and circular DNA templates has been developed. The system consists of incubating template and whole cell extract to generate initiation complexes, followed by addition of nucleotide triphosphates to support elongation, and primer extension assay to detect authentic transcripts. This in vitro transcription system required circularized templates and was essentially inactive with linearized templates. Accurate in vitro transcription of a rice phenylalanine ammonia-lyase (PAL) promoter-beta-glucuronidase (GUS) gene fusion and a tobacco sesquiterpene cyclase promoter-GUS gene fusion was examined in their homologous whole cell extracts, and the optimal concentrations for several reaction components, including DNA template, whole cell extract, monovalent and divalent cations, were determined for specific initiation from the in vivo start site. Transcription was inhibited by low concentrations of alpha-amanitin, demonstrating that the reaction was mediated by RNA polymerase II. Accurate transcription initiation was dependent on the TATA-box motif within the respective promoters. Based on the effect of delayed addition of sarkosyl at a concentration sufficient to inhibit transcription initiation but not elongation, three to four rounds of transcription were initiated in standard assays.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0960-7412
pubmed:author
pubmed:issnType
Print
pubmed:volume
7
pubmed:geneSymbol
GUS, PAL
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
1021-30
pubmed:dateRevised
2009-11-19
pubmed:meshHeading
pubmed:year
1995
pubmed:articleTitle
Accurate in vitro transcription from circularized plasmid templates by plant whole cell extracts.
pubmed:affiliation
Plant Biology Laboratory, Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
pubmed:publicationType
Journal Article, In Vitro, Research Support, Non-U.S. Gov't