豇豆单孢锈菌中病毒样颗粒中双链RNA的存在

PRESENCE OF DOUBLE-STRANDED RNA IN THE VIRUS-LIKE PARTICLES OF VROMYCES VIGNAE BARCL

从北京西郊清华园附近田间豇豆上采集的豇豆单孢锈菌(Uormyces vignal Barcl)夏孢子。萌发后提取双链RNA,电泳分析可测出300—8000碱基对的三组双链RNA。从萌发的孢子中通过差迷离心提取病毒样颗粒,可获得二种类型的病毒样颗粒,一种直径为35—40nm的等轴颗粒。另一种为长短不等的棒状颗粒,用提纯物提取核酸电泳分析与直接从孢子中提取的双链RNA有相同的核酸带,从而证明这些双链RNA存在于病毒样颗粒中。

Electron microscope observations revealed the presence of isometric virus-like particles of 35-40nm diameter (Plate A). These are about the same size as those found in other cowpea rust isolates, Melampsora lint and Puccinia spp. Rod-shaped particles of unequal length were also observed in these cowpe rust spores (Plate B). Previously, McDonald and Heath (1978) had reported three different types of rod-shaped particles present in two other cowpea rust isolates. In Melampsora lini, rod-shaped particles of different lengths have also been found Plate C demonstrates the separation of cowpea rust dsRNAs after agarose gel electrophoresis. The nucleic acids in lane 2 were resistant to DNasel and RnaseA digestion, indicating their identities as dsRNAs. In total there were 7 or more dsRNA segments in the cowpea rust spores, with 4 of those heaving large (5-8kb), 1 medium (2-3kb) and 2, small MW (less than 0.7kb) in size. The pattern of dsRNAs recovered in a total nucleic acid extract (Plate C. lenns 1 and 2), was identical to that found for the dsRNAs isolated from purified VLPs (Plate C. Lenes 3), indicating that all the dsRNAs in the rust spores were encapsidated. Previous work has shown a similar organization for dsRNAs in puccinia spp.. However in Melampsor lini, dsRNAs are both unencapsidated and encapsidated. The detailed study on flax rusts by Dickinson and Pryor (1989a) demonstrated that dsRNA was detected only in isometric VLPs and not in rod-shaped particles. LMS, a strain of M. lini growing on the Australian endemic host plant Linum marginale, was free of detectable dsRNA but still contained the rod-shaped particles. No conclusion can be drawn at this stage for the rod -shaped particles in cowpea rust since no effort has been made to purify the particles or their contents in the present study. The cowpea rust spores used in this experiment were collected from field and so may not represent a pure strain, there by contributing to the complexity of the size and number of dsRNAs observed. However pure isolates of rusts of the genus puccinia show similar dsRNA patterns.