杉木炭疽菌遗传转化及附着孢发育过程的细胞核行为观察

Efficient genetic transformation and nuclear behavior during the process of appressorial development in Colletotrichum gloeosporioides

【目的】建立高效的杉木炭疽菌遗传转化体系,并观察附着胞发育过程中的细胞核行为。【方法】通过菌丝体酶解的方法制备原生质体,PEG介导的原生质体转化法将含有伯莱霉素抗性的细胞核表达质粒NL1∶∶GFP转入受体材料杉木炭疽菌SMCG1#C菌株,通过荧光显微镜跟踪观察附着胞发育过程中的孢子、芽管和附着胞等结构中的细胞核行为。【结果】获得了稳定表达NL1∶∶GFP质粒的杉木炭疽菌阳性转化子;杉木炭疽菌附着胞发育过程伴随着细胞的有丝分裂与细胞核的转移;附着孢发育成熟后,孢子和芽管菌丝中的细胞核仍然保持完整。【结论】建立了高效的杉木炭疽菌遗传转化体系;杉木炭疽菌与稻瘟病菌、希金斯炭疽菌的附着胞发育过程的细胞生物学特性显著不同。

[ Objective] This study was aimed to establish an efficient genetic transformation system in CoUetotrichum gloeosporioides, and to observe the nuclear behavior during the process of appressorial development. [ Method ] Proto- plasts were prepared using cell-wall lyric enzymes. Protoplast transformation was mediated by PEG, and the nucleus-spe- cific expression plasmid NL1 ： ：GFP with zeocin-resistance gene was transformed into the C. gloeosporioides strain SMCGI#C. During the process of appressorial development, nuclear behavior in the conidia, germ tubes, and appresso- rium of C. gloeosporioides was observed under a fluorescence microscope. [ Result] Positive C. gloeosporioides transforma- nts expressing NL1 ： ：GFP were obtained. Appressorial development in C. gloeosporioides was accompanied by mitosis and cell nuclear transfer. The nuclear in the conidia and germ tubes remained intact without nuclear degration after the ap- pressoria were developed ripely. [ Conclusion] An efficient genetic transformation system was established in C. gloeospori- oides. During the process of appressorial development, there were considerable differences in cell-biological characteris- tics among C. gloeosporioides, Magnaporthe oryzae and C. higginsianum.