|Budget Amount *help
¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1988: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1987: ¥1,700,000 (Direct Cost: ¥1,700,000)
A relationship between cytoplasmic male strility (CMS) of Cruciferae plants and variation in cytoplasmic DNAs has been investigated by a restriction endonuclrase analysis and with nine Cruciferae plants with or without CMS to elucidate the nature of CMS at the molecular level, which is useful to produce F1 hybrid plants with better characteristics in agriculture and horticulture.
Chloroplasts were purified by a sucrose-gradient centrifugation method after homogenizing young, green and intact leaves of nine Cruciferae plants. Chloroplast DNA (ct-DNA) was prepared by a DNase treatment to remove a contaminated nuclear-DNA, followed by a deproteinizing treatment with proteinase K and chlorolorm-phenol mixture. Since the ct-DNA obtained thus was not digested by three types of restriction endonucleases, the nine ct-DNAs was further purified by using a commercial kit of GENECLEAN from Funakoshi Co..
variation in cytoplasmic DNAs was examined by the treatment of three restriction endonucleases (
Hind III, EcoR I, Sal I) and consequently 1% agarose gel electrophoresis detecting total number of fragments and mobility of each fragment in the digest products.
Ct-DNAs from leaves of the nine plants showed a very similar electrophoretical results each other, but a few bands of fragments were recognized to be different among them. Especially, between two species, radish and turnip, there was a distinct difference in the both electrophoretical patterns. No difference was recognized between diploid and tetraploid, and CMS species and normal species in the Cruciferae plants. In a hybrid plant, Raphanobrassica was very similar to radish at the restiction endonuclease digest patterns. Finally, it is suggested that there is no variation in ct-DNAs being related to CMA.
Mitochondrial DNAs (mt-DNA) were purified from main roots of nine Cruciferae plants. Although last year they were tried to purify from seedlings, the try was unsuccessful because of difficulty to get a large numer of seeds. The yields of mt-DNAs from main roots differed much depending on the age of main roots. Some of them too low to analyze the variation in mt-DNAs. The variation in mt-DNAs is likely to be larger than it in ct-DNAs and it may be related to CMS. Less