Optimization of Recombination Proteins of Ethylene Receptor ETR1 from Feicheng Peach Fruit and Effect of Nitric Oxide on Expression of ETR1

In this study a ripening related ethylene receptor ETR1 gene in Feicheng peach fruits was cloned and the recombination protein of ETR1 was obtained for the further researches on the functions of ETR1 and improving the storage and transport quality of Feicheng peach fruits.Using cDNA reverse-transcribed from total RNA in mature Feicheng peach fruits as template,the specific PCR product of ETR1 was obtained,then it was ligated to pEASY-T1 vector.The sequencing data showed that the PCR product was about 2 500 bp.After digested by BamHⅠ and NdeⅠ,the sequence was ligated to pET15b and transformed to E.coli.Recombinant ETR1 was induced by IPTG,and the expression of ETR1 was optimized.And with the cDNA as templates,perform real-time quantitative fluorescence PCR,determine the influence of exogenous NO on expression of ETR1.Comparison with the cDNA,sequence of AF124527 indicated the homology was 99%,and amino acid identity was 99%.The proper conditions for recombinant ETR1 protein were that the mutant host strains E.coli was C43(DE3).The final concentration of IPTG was 0.5 mmol/L.The proper temperature was 30℃ and the induction time was 8 h.Recombinant ETR1 was highly expressed under the optimized conditions,which was helpful for studying the functions of ETR1 in vitro.And,the inhibition by NO on the expression of ETR1 depended on the dose of NO.