番荔枝科植物鹰爪内生真菌rDNA ITS-PCR扩增条件的优化

探索一种简单、快速的获得番荔枝科植物鹰爪内生真菌rDNA ITS目的片断的方法。采用改良的Rodrigues组织培养表面消毒技术和CTAB法提取植物样品总DNA,利用真核生物通用引物LH2/Sm73扩增rDNA ITS目的片段,正交法优选ITS-PCR扩增条件。PCR反应可同时获得两条600 bp和700 bp产物,分别为植物和内生真菌的rDNA ITS目的片断。改良的组织表面消毒技术可排除外源DNA污染。正交法可快速获得植物内生真菌目的片断。该法简单、快速,为番荔枝科植物内生真菌生物多样性研究提供分子生物学基础。

Optimization for ITS-PCR Amplification Conditions on Endophytic Fungi of Artabortrys hexapetalus (L.f.) Bhandari

To develop a simple,rapid method for amplifying fungal rDNA ITS regions within plant tissues of Artabortrys hexapetalus(L.f.) Bhandari.The healthy leaves were surface sterilized following a procedure modified from Rodrigues.The total genomic DNA were extracted using CTAB method with modifications.Using the optimization condition of PCR based on orthogonal design,rDNA ITS regions were amplified with a pair of eukaryote universal primers LH2/ Sm73 for endophytic fungi within Artabortrys hexapetalus(L.f.) Bhandari.PCR amplifications of the rDNA ITS regions were shown being two distinguished and closed fragments which the 700 bp is angiosperms and 600 bp is corresponding a typical rDNA ITS size for fungi.DNA contamination can be prevented by the sterile methods using modified Rodrigues for tissue surface.The optimization of orthogonal design is a simple,rapid method for endophytic fungi of angiosperms.