基于cry1Ac表達(dá)調(diào)控元件的蘇云金芽孢桿菌表達(dá)載體構(gòu)建

nng—hai,sunming—論文發(fā)表專家一趣|中國(guó)學(xué)木期測(cè)^jj^www.qikanwang,nel基于crylAc表達(dá)調(diào)控元件的蘇云金芽抱桿菌表達(dá)載體構(gòu)建摘要：通過(guò)克隆cry/lac基因bti—btii啟動(dòng)子、sd序列以及終止子，并引入pet28a的his標(biāo)簽和多克隆位點(diǎn)，在穿梭載體pht304的基礎(chǔ)上構(gòu)建了一個(gè)蘇云金芽抱桿菌表達(dá)載體pbmbla。將crylac以及具有非典型bti—btii啟動(dòng)子的cry2ab、cry5ba、cry6aa、cry7ba、cry55aa6個(gè)基因裝載到該表達(dá)載體上，轉(zhuǎn)入bmb171構(gòu)建了重組菌株，通過(guò)復(fù)紅簡(jiǎn)單染色后的顯微鏡鏡檢結(jié)果表明，重組菌株bmb1a-1ac、bmb1a-5babmb1a-7ba和bmb1a-55aa均能形成正常晶體，sds—page結(jié)果也證實(shí)這4個(gè)重組菌株的重組質(zhì)粒均能表達(dá)出目的蛋白。同時(shí)，選取重組菌株bmb1a-1ac來(lái)考察his標(biāo)簽對(duì)crylac殺蟲活性的影響，生物活性測(cè)定結(jié)果顯示重組菌株的lc50值與對(duì)照菌株相比無(wú)明顯差異。該載體可用于快速克隆表達(dá)不同類別的cry蛋白。關(guān)鍵詞：蘇云金芽抱桿菌；表達(dá)載體；cry基因；bti—btil啟動(dòng)子constructionofbacillusthuringiensisexpressionvectorbyusingregulatoryelementsfromcrylacgenezhengwen,yewei—xing,pengdo醫(yī)一論文發(fā)袤專家一J中國(guó)黠斛網(wǎng)f^www.qikanwang.nelcollegeoflifescienceandtechnology/statekeylaboratorechnology/statekeylaboratoryofagriculturalmicrobiology,huazhongagriculturaluniversity,wuhan430070,china)abstract:abacillusthuringiensisexpressionvectornamedpbmblawasconstructedbycloningbti—btiipromoter,sd—sequence,andtranscriptionterminatorofcrylacgene,andaddinghis—tagandthefollowingmultiplecloningsites(mcs)frompet28aintotheshuttlevectorpht304.crylacandotherfivecrygeneswithatypicalbti—btiipromoterincludingcry2ab,cry5ba,cry6aa,cry7ba,cry55aawereclonedintopbmbla.microscopicobservationshowedthattherecombinantstrainscontaini

醫(yī)一論文發(fā)袤專家一J中國(guó)黠斛網(wǎng)f^www.qikanwang.nelngcrylac,cry5ba,cry7baandcry55aacouldngcrylac,cry5ba,cry7baandcry55aacouldformparasporalinc1usionbodies;andsds—pageproveducesecticidalcrystalproteinsbands.meanwhile,crylacwasselectedtodeterminethefectofhis—tonitsinsectidalactivitandthebioassresultshowehattherewasveducesecticidalcrystalproteinsbands.meanwhile,crylacwasselectedtodeterminethefectofhis—tonitsinsectidalactivitandthebioassresultshowehattherewassignifcantfferencebetweenthelthatallofthemcouldprodthecorrespondingmajorinc50oftherecombinantstrainandthecontrolstrain.theexpressionvectorconstructedwassuitableforrapidcloningandexpressionofdifferentcrygen