摘要
构建了ura3缺失基因载体pEMT-△ura3,采用农杆菌介导法转化里氏木霉QM9414,在含尿嘧啶核苷(1.87 mg.mL-1)和五氟乳氰酸(5-FOA,5 mg.mL-1)的筛选培养基上筛选转化子,经表型验证和PCR检测,获得ura3基因缺失突变体1株,ura3基因的同源重组率为16.6%。采用农杆菌介导法将黑曲霉的pyrA基因导入ura3基因缺失突变体中,使其回复野生型表型,从而建立了以pyrA基因作为筛选标记的里氏木霉受体菌株。
A binary vecter pEMT-△ura3 with deleted ura3 gene was constructed in this study.The deleted ura3 gene was transformed into Trichoderma reesei QM9414 by Agrobacterium tumefaciens-mediated transformation.Transformants were selected on culture medium containing 1.87 mg·mL-1 uracil and 5 mg·mL-1 5-FOA.One △ura3 auxotrophic strain was obtained by phenotype and PCR analysis.The level of homgous recombination was 16.6%.Furthermore pyrA gene from A.niger was transformed into uridine auxotrophic ura3 mutant Trichoderma reesei,which can restore the phenotype.On the based of this work a transform acceptor was obtained using pyrA gene as a selectable marker.
出处
《东北农业大学学报》
CAS
CSCD
北大核心
2011年第7期147-153,共7页
Journal of Northeast Agricultural University
基金
山东大学微生物技术国家重点实验室开放项目资助(M2008-17)
关键词
营养缺陷型
农杆菌介导法
同源重组
ura3基因
auxotrophic
Agrobacterium tumefaciens-mediated transformation
homologous re-combination
ura3 gene
