基因枪法导入GUS基因获得小麦条锈菌突变体

Virulent transgenic mutants obtained by particle bombardment transformation of Puccinia striiformis f. sp. tritici with the GUS gene

采用基因枪转化法用带有β-葡糖醛酸酶(GUS)报告基因的质粒pGUS6L20转化小麦条锈菌,建立了优化的转化体系;在转化当代的条锈菌中得到了GUS基因瞬时表达的菌株;连续继代转接培养后,在转化后第1代(T1)和第2代(T2)条锈菌中检测到了GUS基因稳定表达的菌株;利用GUS报告基因和毒性标记相结合的方法,经过3代筛选鉴定,在小麦条锈菌中国鉴别寄主尤皮Ⅱ号和抗引655上,各获得1个稳定的毒性突变体,经PCR及PCR-southern b lot证实外源片段已整合到毒性突变菌株的基因组中。该研究表明基因枪转化法是获得条锈菌毒性突变体的有效方法。

To obtain virulent mutant stains, the optimized transformation system was established and plasmid pGUS6L20 containing the GUS gene was transformed into urediniospores of Puccinia striiformis f. sp. tritici via particle bombardment. The transient expression of GUS gene was identified in the transformed urediniospores. The GUS-carrying spores were cultured and increased consecutively. GUS gene was expressed in the uredia and urediniospores of T1 and T2 generation of the stripe rust fungus. Two insertion mutants of increased virulence were identified on wheat cultivars Jubilejina I1 and Kangyin 655, respectively. The presence of GUS gene was detected by polymerase chain reaction （PCR） and Southern blot in these two virulent mutants. The results showed that the particle bombardment was an effective way to obtain the virulent strains of P. striiformls.