里氏木霉外源表达载体的构建（摘要）.pdfVIP

AgriculturalScienceTechnology．2011，12(2)：308—312 Copydght⑥2011，InformationInstituteofHAASAllrightsreserved AgriculturalBiotechnology Construction ofExogenous Expression Vector‘of Trichoderma reesei ZHANG Xiao-xuan ，WANG An-xue 1．ChengdongCollege，NortheastAgriculturalUniversity，Harbin150030；2，CollegeofLifeSciences，NortheastAgriculturalUniversity Har- ， bin150030 Abstract [Objective]ThestudywastoconstructaexogenousexpressionvectorforTrichodermareeseL[Method]UsingCBHJpromoterand terminatorof reeseistrain40359．weconstructedanexpressionvectorof reeseistrain40359forexpressingHptgeneandgotsixstrains capableofgrowingonbasicmediumcontaining175mg／LofhygromycinB，furtherconductedhygromycinresistancetest．[Results]Incompad— sonwiththeoriginalstrain(wildtype)，hygromycinresistancethesixengineeredstrainswasincreasedby75％；thehygromycinresistancecouId jnhentstably．[Conclusion]0urresultsJaidbasisforbiologicalstudyon7-,feeseiatmolecularandgeneticallyengineeringIevels． Keyworde Trichodermareesei；Vector；Genetictransformation CBHIpromoterfrom Trichodermareeseijsaverystrong Mediarecipe(for1L) Basalmedium：20gglucose。15g promoterthatisusuallyusedforvectorconstructionforgenet— KHPO ，5g(NH )SO ，0．6gCaCI2·2HO，0．6gMg- icimprovementof-／-．reesei． e．．insertingthegeneofinter— SO4·7H2O，0．005gFeSO4·7H2O，0．0016gMnSO4·H2O， est(GOI)jntothespacebetweenCBH』promoterandtermi— 0．0014gZnSO4·7H2O，0．002gCoCI2·6 O，pH=5。5；auto- natorsequence，Therecombinedfragmentistransformedinto clavingat121oC for30 min faddition of2％ ofagarforsolid filamentouslungiprotoplast．and lOcaled and integrated Into medium and1％ agarforsemi-solid medium、．Regeneration chromosomeby usingthe method oftargetedgene replace— media：(1)Iowerlayermedium：additionof1moVLofsorbI-