| 农杆菌介导法构建里氏木霉ura3基因突变体的研究 |
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| 引用本文： | 闫作梅,张旭,李杰.农杆菌介导法构建里氏木霉ura3基因突变体的研究[J].东北农业大学学报,2011,42(7):147-153. |
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| 作者姓名： | 闫作梅 张旭 李杰 |
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| 作者单位： | 东北农业大学生命科学学院,哈尔滨,150030 |
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| 基金项目： | 山东大学微生物技术国家重点实验室开放项目资助 |
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| 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。
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| 关 键 词： | 营养缺陷型 农杆菌介导法 同源重组 ura3基因 |
Construct of ura3 gene auxotrophic of Trichoderma reesei by Agrobacterium tumefaciens-mediated transformation |
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| YAN Zuomei,ZHANG Xu,LI Jie.Construct of ura3 gene auxotrophic of Trichoderma reesei by Agrobacterium tumefaciens-mediated transformation[J].Journal of Northeast Agricultural University,2011,42(7):147-153. |
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| Authors: | YAN Zuomei ZHANG Xu LI Jie |
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| Institution: | (College of Life Sciences,Northeast Agricultural University,Harbin 150030,China) |
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| Abstract: | 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. |
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| Keywords: | auxotrophic Agrobacterium tumefaciens-mediated transformation homologous re-combination ura3 gene |
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