Barenbrug Holland BV, Stationsstraat 40, NL-6678, AC Oosterhout, The Netherlands.
Theor Appl Genet. 1992 Aug;84(5-6):763-70. doi: 10.1007/BF00224182.
Techniques have been developed for the production of cybrids in Lolium perenne (perennial ryegrass). Gamma-irradiated protoplasts of a cytoplasmically male-sterile breeding line of perennial ryegrass (B200) were fused with iodoacetamide-treated protoplasts of a fertile breeding line (Jon 401). After fusion 25 putative cybrid calli were characterized to determine mitochondrion type and composition of the nuclear genome. Analysis of phosphoglucoisomerase isozyme profiles and determination of the ploidy level by flow cytometry indicated that all of the calli tested essentially contained the nuclear DNA of the fertile line. However, the presence of parts of the nuclear DNA from the sterile line could not be excluded. Southern blotting of total DNA isolated from the parental lines and putative cybrids combined with hybridizations using the mitochondrial probes cox1 and atp6 revealed that the mitochondria of the calli originated from the fertile line (5 calli), the sterile line (5 calli) or from both parental lines (15 calli). The hybridization patterns of the mtDNA from the cybrid calli showed extensive quantitative and qualitative variation, suggesting that fusion-induced inter- or intramolecular mitochondrial recombination had taken place.
已经开发出了生产 Lolium perenne（黑麦草）细胞杂种的技术。用碘乙酰胺处理的可育黑麦草（Jon 401）原生质体与细胞质雄性不育育种群（B200）的辐照原生质体融合。融合后，对 25 个假定的细胞杂种愈伤组织进行了鉴定，以确定线粒体类型和核基因组的组成。磷酸葡糖异构酶同工酶谱分析和流式细胞术测定的倍性水平表明，所有测试的愈伤组织基本上都含有可育系的核 DNA。然而，不能排除不育系部分核 DNA 的存在。从亲本系和假定的细胞杂种中分离出的总 DNA 的 Southern 印迹杂交，并结合使用线粒体探针 cox1 和 atp6 的杂交表明，愈伤组织的线粒体来源于可育系（5 个愈伤组织）、不育系（5 个愈伤组织）或两个亲本系（15 个愈伤组织）。细胞杂种 mtDNA 的杂交模式显示出广泛的定量和定性变化，表明融合诱导的线粒体间或分子内重组已经发生。
