水稻CYP81A6蛋白原核表达与活性检测

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中图分类号：S511.01 文献标识码：A 文章编号： 1000-4440（2025）11-2081-07

Abstract：In plants，the plant cytochrome P45O gene familyis involved inavarietyof physiological activities.Our previoustranscriptome sequencing analysis revealed that the relative expresion level ofthe CYP8IA6 gene was up-regulated bythe inductionwithsalicylicacidandriceblastfungus.Althoughitisknown thattheCYP8IA6genehasherbicide resistance，therehavebeennoreportssofarontheinvolvementof thisgene inplant diseaseresistanceresponses.Therefore，in thisstudy，rice Nipponbareleaves wereusedas experimental materials.TheCYP8lA6gene fragmentof rice was amplifiedbyPCR technologySubsequently，thesuccessfullyamplified gene fragment wascloned into the prokaryoticexpression vector pET32a.Based on the above experiments， theprotein expression and purification were performed for thisgene.In order to obtain more soluble recombinant protein，screening experiments for the induction conditions of CYP81A6 proteinwere carried out.The results showed that theoptimal protein expression was achieved under induction conditions of O.7 mmol/L isopropyl -β D -thiogalactoside（IPTG）at 16‰ for 20h .Meanwhile，the CYP81A6 protein could be successfully eluted and purified under the actionof 5O mmol/L imidazole.Further，the purified CYP81A6 protein wasused for enzymaticreaction with nicotinamide adenine dinucleotide phosphate（NADPH），and the enzyme activityofCYP8IA6 was measured tobe1.518 U/mL by profesionaldetectionmethods.Insummary，thisstudyhaslaidafoundation for in-depth explorationof the functionand mechanism of the CYP8lA6 gene inricethrough cloning，expression，purification and enzyme activitydetermination.

Key words： rice；CYP8IA6；cytochrome P45O；prokaryotic expression；protein purification；rice blast

水稻作为全球重要的粮食作物，在生产上极易遭受稻瘟病病菌（Magnaportheoryzae）的感染。（剩余12557字）