关键词: 杜梨(Pyrus betulaefolia), 发根农杆菌(Agrobacterium rhizogenes), 毛状根, 遗传转化

Abstract: Pyrus betulaefolia was used as the research material to establish a simple,rapid and efficient method for genetic transformation of pear hairy roots by Agrobacterium rhizogenes.The expression vector pROK2 containing mCherry gene was introduced into Agrobacterium rhizogenes K599 to infect the seedlings of Pyrus betulaefolia.The function of the expression vector was detected by the three methods,namely,injection of tobacco leaves,routine PCR and stereoscopic fluorescence microscope observation.By transient expression in infected tobacco leaves,it was proved that mCherry could be expressed in plant cells.After infected by Agrobacterium rhizogenes infected 34 Pyrus betulaefolia seedlings regenerated hairy roots and the regeneration rate was 85%.Through the identification by PCR,there were 6 of the 34 transgenic plants with positive transgenic hairy roots,with a positive rate of 17.6%.The red fluorescence of mCherry can be observed directly in the transgenic root by stereoscopic fluorescence microscope.In this study,by using the method of Agrobacterium rhizogenes directly infecting Pyrus betulaefolia,the expression of foreign gene in the new adventitious root of pear was successfully realized.This method overcomes the difficulties such as complex genetic background,long childhood,simple operation,high transformation rate and short cycle.It can transform roots stably in a relatively short time,which provides a new tool for pear gene function research and genetic improvement.This work provides a foundation for improving the success rate of genetic transformation of pear trees.

Key words: Pyrus betulaefolia, Agrobacterium rhizogenes, hairy roots, genetic transformation