摘要
由尖镰孢古巴专化型1号生理小种Fusarium oxysporumf.sp.cubenserace1(Focr1)引起的香蕉枯萎病是粉蕉类香蕉品种(AAB)不能规模化种植的最主要因素,其致病机理至今尚不十分清楚。本实验室前期通过T-DNA插入获得Focr1致病性丧失突变体Focr1-328,从Focr1全基因组序列中定位了因T-DNA插入失活的基因,并从野生型菌株Focr1-N2中敲除了该致病相关基因,获得了敲除突变体△Focr1-328。为了明确该致病相关基因的生物学功能,通过活体叶片、活体根部、孢子悬浮液等接种方法对敲除突变体△Focr1-328的致病性进行了测定,研究了该突变体与野生型菌株Focr1-N2在PDA培养基上菌落、菌丝和分生孢子形态上的差异;在不同碳、氮源培养基上的生长速率及形态差异;在培养不同时间后菌体生物量、pH值、OD值的变化差异及玻璃纸穿透能力的差异等。致病性测定结果显示4种接种方法都未见△Focr1-328发病症状,表明其丧失了致病力;表型测定结果显示:在PDA培养基上,△Focr1-328的生长速率、分生孢子产量、分生孢子萌发率、培养不同时间的菌丝干重均明显低于野生型菌株;在不同培养时间的培养液中,△Focr1-328的pH值、OD值均极显著低于野生型菌株Focr1-N2;敲除突变体和野生型菌株在不同碳、氮源上生长差异显著,最适碳源分别为山梨醇和麦芽糖,最适氮源均为硝酸钠;突变体菌株不能穿透玻璃纸生长,而野生型可以。上述实验结果表明:T-DNA插入失活的致病相关基因与菌株的碳源利用、产酸调控及菌丝穿透能力有关。
Fusarium wilt of banana （Musa spp.）, caused by Fusarium oxysporum f. sp. cubense race 1, is the most important factor limiting the plantation of plantain banana （AAB Group Fenjao） on the large scale. The pathogenic mechanism is still not very clear yet. A nonpathogenic mutant Focrl-328 with a T-DNA insertion was recently obtained. The inactivated gene resulting from a T-DNA insertion was located in the genomic sequences of Focrl. Subsequently, the gene was knocked out and a deletion mutant △Focrl-328 was obtained by homologous recombination. In order to investigate the function of the pathogenic related gene, the pathogenicity of the wild type Focrl-N2 and the deletion mutant △Focrl-328 was tested by inoculating living leaves and roots with spore suspension. The differences in the colony morphology, mycelial growth and conidial production on PDA, the growth rates and morphology on different carbon and nitrogen sources and the biomass, potential of hydrogen value, optical density value as well as cellophane penetrating capability were studied. The results of pathogenicity experiments showed that the deletion mutant △Focrl-328 was nonpathogenic, indicating that △Focrl-328 lost the pathogenic capability. The growth rate, the quantity and germination rate of conidia and the biomass of mycelia of the deletion mutant △Focrl-328 were significantly lower than those of wild type. The best carbon sources for the mutant and the wild type growth was sorbitol and maltose, respectively. The optimum Nitrogen source was sodium nitrate in both cases but the growth rates and colors of mutant and wild type were different on various C and N media. The wild strain penetrated cellophane readily, but the mutant could not. The results indicated that pathogenic related gene which was inactivated by T-DNA insertion is related to the utilization of C source, acid production and mycelial penetration capability.
出处
《菌物学报》
CAS
CSCD
北大核心
2012年第4期593-607,共15页
Mycosystema
基金
国家自然科学基金(No.30860160)
国家公益性行业(农业)科研专项课题(No.200903049-2)
海南大学“211工程”建设项目
海南大学环境与植物保护学院研究生创新平台项目
关键词
香蕉枯萎病
致病性
敲除突变体
生物学特性
banana Fusarium wilt, pathogenicity, knock out strain, biological characteristics
