Cloning of the chick hsp 90 cDNA in expression vector.
A cDNA clone for the 90kDa heat-shock protein, which we have recently identified as a component of steroid hormone receptors in their heteromeric 8S form, was isolated by direct immunological screening of a chicken smooth muscle cDNA expression library, prepared in the expression plasmids pUC8 and pUC9. Using polyclonal and monoclonal antibodies against the 90kDa protein a colony was identified that reacted with both antibodies. Plasmid 9.11 (p9.11, approximately 1100 base pair insert) was found to hybrid-select mRNA for the 90kDa heat-shock protein. Northern blot analysis revealed that RNA isolated from various chicken tissues contain a single transcript of approximately 3 Kb hybridizing to a [32P]labelled cDNA probe made from p9.11. Heat-shock treatment of chick embryonic fibroblasts resulted in increased steady-state levels of a 3 Kb transcript in both poly A+ and poly A- RNA fractions. Southern blot analysis of chicken genomic DNA indicated that the cDNA hybridizes to a single copy sequence. Sequence data show that the p9.11 cDNA displays a high degree of homology with the 5' portion of yeast heat shock protein 90 cDNA.
D000595:Amino Acid Sequence; D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D002645:Chickens; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D005822:Genetic Vectors; D006360:Heat-Shock Proteins / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013045:Species Specificity
D006360:Heat-Shock Proteins; D012333:RNA, Messenger; D004247:DNA