Linked Life Data

3012280

Source:http://linkedlifedata.com/resource/pubmed/id/3012280

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
1986-7-14
pubmed:abstractText
The Saccharomyces diastaticus structural gene STA2, encoding an exracellular glucoamylase (1,4-alpha-D-glucan glycohydrolase, EC 3.2.1.3.), has been cloned by complementation of a stao strain. A genomic library was initially constructed from a STA2 yeast strain in the yeast Escherichia coli shuttle cosmid vector pYCl. The Sta+ complementing function was further delimited to an 8.3 kb BglII fragment whose restriction map was found to be similar to related genomic regions of STA1 and STA3. Fusions of several DNA fragments derived from the 8.3 kb BglII fragment with a truncated E. coli beta-galactosidase gene resulted in two overlapping fragments that could direct the production of large fusion proteins in E. coli. These fusion proteins were immunoprecipitable by anti-glucoamylase II antibodies, confirming that the Sta+ complementing fusion was due to the expression of a gene that coded for a yeast glucoamylase. Measurements of the STA1, STA2 and STA3 RNA transcripts by RNA-DNA hybridization using an internal fragment of the cloned STA2 gene as the probe indicated that a common transcript of 2.5 kb is produced by each of the STA genes. Integrative disruption of the STA2 gene through homologous recombination was achieved by transforming a STA2 yeast strain to Sta- using an in vitro constructed donor DNA fragment that has the URA3 gene inserted within the coding region of the cloned glucoamylase gene. This was confirmed by tetrad analysis of crosses between strains carrying a disrupted STA2 and a functional STA2. Southern blot analysis using BamHI digested genomic DNA from 15 tetrads demonstrated consistent co-segregation and Mendelian inheritance of the Sta- phenotype with STA2::URA3. These data further confirm that the cloned DNA that showed Sta+ complementing activity carries a functional STA2 gene that encodes the yeast extracellular glucoamylase II.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
0026-8925
pubmed:author
pubmed:issnType
Print
pubmed:volume
203
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
29-35
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1986
pubmed:articleTitle
Molecular cloning and characterization of the STA2 glucoamylase gene of Saccharomyces diastaticus.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S.