| 摘要: |
| 【目的】探索建立以G418抗性为筛选标记的农杆菌介导的深绿木霉遗传转化方法，为木霉菌突变菌株的筛选和基因功能研究奠定基础。【方法】测试供试野生型深绿木霉菌菌株T23对G418的敏感性，构建以G418抗性为筛选标记的质粒载体p1300-neo，并将该质粒通过农杆菌导入深绿木霉T23菌株中，利用G418抗性平板筛选获得转化子，通过PCR对稳定转化子进行鉴定。【结果】T23菌株对G418具有高度敏感性，25 μg/mL G418可完全抑制T23菌株的生长；构建的p1300-neo载体是可行的G418抗性标记，通过该载体获得了多株具有G418抗性的深绿木霉遗传转化子。【结论】G418抗性可以作为深绿木霉菌有效的遗传筛选标记，其在PDA培养基中的使用量为25 μg/mL。 |
| 关键词: G418抗性 根癌农杆菌 深绿木霉 |
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| 基金项目:国家自然科学基金项目（31440025，41001189）；安徽省自然科学基金项目（1408085MC44）；阜阳师范学院科技成果孵化基金项目（2013KJFH04）；安徽省大学生创新创业训练计划项目（AH201310371046，AH201310371047） |
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| Agrobacterium tumefaciens mediated transformation of Trichoderma atroviride with G418 as screening marker |
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TANG Jun,MA Yue,WANG Shu-mei,et al
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| Abstract: |
| 【Objective】The paper aimed to establish a method of Agrobacterium-mediated Trichoderma atroviride transformation,which will lay the foundation for the study of gene function through gene knockout and gene complement techniques.【Method】Firstly,sensitivity analysis of Trichoderma atroviride to G418 was done,then the plasmid vector p1300-neo with G418 resistance marker was constructed and transformed into Trichoderma atroviride T23 by Agrobacterium mediated transformation methods.G418-resistant transforrmants were screened and further identified by PCR.【Result】Trichoderma atroviride T23 showed high sensitivity to G418,and strain T23 were wholly inhibited with 25 μg/mL G418.The constructed vector p1300-neo was feasible as a resistant marker.G418-resistant transformants were obtained.【Conclusion】G418-resistance was an effective genetic screening marker for Trichoderma atroviride and its proper dose in PDA medium was 25 μg/mL. |
| Key words: G418-resistance Agrobacterium tumefaciens Trichoderma atroviride |
