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Saccharomyces cerevisiae cells in both the memory associated with the cAMP signaling pathway , respectively, by heterotrimeric G proteins and small G protein Ras mediated . Saccharomyces cerevisiae Ras-cAMP signaling pathway in yeast cell metabolism , proliferation, differentiation, and plays an important role in stress resistance . Ras-cAMP signaling pathway downstream of the direct target is the cAMP-dependent protein kinase A is PKA; when Ras protein activity is too high , PKA activity has also come increased. PKA as a serine / threonine kinase that may make it a target protein serine or threonine residues phosphorylated . By means of phosphorylation , PKA regulate cell nutrition and pressure on the outside of the stress reaction conditions . Studies have shown that yeast cells can affect the activity of PKA thermal shock resistance of the cell ; PKA activity in the case of high , Saccharomyces cerevisiae cells, thermal shock resistance significantly lower than the wild type . Further investigate the activity of PKA high thermal shock resistance of the cell between , this study strain W303-1a YCp22RASval19 starting library of random mutagenesis using mTn-lacZ/LEU2 inserted into its chromosome form any part of the transposon . mTn library into strain W303-1a YCp22RAS2val19 , if the transposon insertion sites are resistant ORF genes or cis- acting element , then the formation of thermal shock resistant mutants may have changed . Purified transformants test its thermal shock resistance , thermal shock identified six enhanced resistance mutants ; among ZL7, ZL9 and ZL13 heat shock tolerance time up to 6min, and ZL10, ZL11 and ZL16 of heat shock tolerance time also increased to 3min. Over pRSQ2-URA3 plasmid was then recovered on a chromosome identification transposon insertion sites , confirmed mutated gene ; wherein ZL7 NOP7 and RAS2 mutant of strain double inactivation , ZL9 inactivation mutant strains of ULP2 , ZL10 and ZL16 are NFS1 inactivation mutant strains , ZL11 inactivation mutant strains of TPK1 , ZL13 as TIM9 inactivation mutant strains . Finally, the building YCp33ULP2, YCp33TIM9, and with the existing YCp33TPK1 and plasmids were transformed into the corresponding YCp33NOP7 deficient strain , revertant strains formed , which restored the wild -type strain of thermal shock resistance , indicating ZL7, ZL9, ZL11 and ZL13 other four strains mutation confirmation .
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