Yu Yi-Fan, Guo Juan, Su Ping, Ma Ying, Zhao Yu-Jun, Huang Lu-Qi
State Key Laboratory Breeding Base of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China.
School of Pharmacy, Jiangsu University, Zhenjiang 212013, China.
Zhongguo Zhong Yao Za Zhi. 2018 May;43(10):2048-2052. doi: 10.19540/j.cnki.cjcmm.20180307.003.
The electroporation method was performed to transfer plasmid DNA of PBI-1300 carrying GFP gene into Agrobacterium rhizogenes C₅₈C₁ strains. Mediated by A. rhizogenes C₅₈C₁, the GFP gene were transformed into Erigeron breviscapus aseptic leaves by leaf disc method, then the hairy roots were induced and the infected hairy roots were screened by hygromycin resistance. The chromosomal DNA of the hairy root was used as the templates for the PCR amplification with the GFP-specific primers and then the expected amplified DNA bands appeared, the green fluorescent of GFP in the cut hairy roots was observed by two-photon microscope. These results indicated that GFP gene was integrated into the genome of E. breviscapus and was expressed stably. This study laid the groundwork for foreign gene high-efficiency expression inthe genetic transformation system for hairy root culture of E. breviscapus.
采用电穿孔法将携带绿色荧光蛋白（GFP）基因的PBI-1300质粒DNA转入发根农杆菌C₅₈C₁菌株。以发根农杆菌C₅₈C₁为介导，通过叶盘法将GFP基因转化到灯盏花无菌叶片中，然后诱导毛状根并通过潮霉素抗性筛选感染的毛状根。以毛状根的染色体DNA为模板，用GFP特异性引物进行PCR扩增，出现预期的扩增DNA条带，用双光子显微镜观察切割后的毛状根中GFP的绿色荧光。这些结果表明GFP基因已整合到灯盏花基因组中并稳定表达。本研究为灯盏花毛状根培养遗传转化体系中外源基因的高效表达奠定了基础。
