Katsuya S, Kaneko I, Owaki M, Ishikawa K, Tsujimoto T, Tsuge T
School of Agricultural Sciences, Nagoya University, Japan.
Genetics. 1997 May;146(1):111-20. doi: 10.1093/genetics/146.1.111.
We found the presence of plasmid DNA in strain T88-56 of the Japanese pear pathotype of Alternaria alternata, which causes black spot of certain cultivars of Japanese pear by producing host-specific AK-toxin. The plasmid, designated pAAT56, was identified to be an approximately 5.4-kilobase (kb) circular molecule by electron microscopic observation and restriction endonuclease mapping. Southern blot analysis showed that pAAT56 DNA had no homology with either nuclear or mitochondrial DNA. Cultures of strain T88-56 grown at 26 degrees showed markedly reduced plasmid levels relative to those grown at lower temperatures. The strain was completely cured of pAAT56 during growth at 29 degrees. Temperature-dependent curing of pAAT56 was confirmed by using single-protoplast isolates from mycelia grown at 23 degrees, most of which maintained the plasmid, and from mycelia grown at 29 degrees, most of which had lost the plasmid. Northern blot analysis detected the presence of three RNA species (approximately 1.7, 2.7 and 5.4 kb) transcribed from pAAT56. The biological function of pAAT56 was observed using single-protoplast isolates from mycelia that either contained or had been cured of pAAT56. The plasmid-containing isolates tended to be reduced in AK-toxin production and pathogenicity compared with the plasmid-cured isolates.
我们在链格孢菌日本梨致病型的T88 - 56菌株中发现了质粒DNA，该致病型通过产生宿主特异性AK毒素导致某些日本梨品种出现黑斑病。通过电子显微镜观察和限制性内切酶图谱分析，鉴定出该质粒为约5.4千碱基（kb）的环状分子，命名为pAAT56。Southern杂交分析表明，pAAT56 DNA与核DNA或线粒体DNA均无同源性。在26℃下培养的T88 - 56菌株的培养物，其质粒水平相对于在较低温度下培养的培养物明显降低。在29℃生长期间，该菌株完全消除了pAAT56。通过使用从23℃生长的菌丝体中分离的单个原生质体（其中大多数保留了质粒）和从29℃生长的菌丝体中分离的单个原生质体（其中大多数失去了质粒），证实了pAAT56的温度依赖性消除。Northern杂交分析检测到从pAAT56转录的三种RNA种类（约1.7、2.7和5.4 kb）的存在。使用来自含有pAAT56或已消除pAAT56的菌丝体的单个原生质体分离物观察pAAT56的生物学功能。与消除质粒的分离物相比，含有质粒的分离物的AK毒素产生和致病性往往降低。
