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Development of a System for Identification of Hybrid Plants

杂交植物识别系统的开发

基本信息

批准号：
01860002
负责人：
HIRAI Atsushi
金额：
$ 3.52万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Developmental Scientific Research (B).
财政年份：
1989
资助国家：
日本
起止时间：
1989 至 1990
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01860002/
关键词：
Cell fusion Hybrid identification Simple method for DNA isolation Nonradioactive label rRNA gene

项目摘要

A simple and efficient method for identification of hybrids after cell fusion was established by combining a method for the efficient extraction of DNA from very small amounts of plant tissue and a Southern blotting with nonradioactively labeled rDNA from rice as a probe. One hundred milligrams of leaf or ca llus tissue, are sufficient for identification of the hybrid, and the amount of material can be reduced to as little as 10 mg. Since this method permits selection of parasexual hybrids not only at the plantlet stage but also at the callus stage, hybrids can be identified at the early stages of the development of the products of cell fusion.Segments of mitochondrial DNA carrying the gene for the alphasubunit of ATPase (atpA) were detected by Southern hybridization with atpA from pea as probe. In the case of Nicotiana langsdorffii, we identified four fragments that are derived from combinations of two different 5' and two different 3' franking regions of atpA. All of the four types share the coding region, suggesting that the four types can be generated as a result of homologous recombination in the coding region of atpA. By contrast, N. Glauca generated only one fragment which indicated the existence of only a single type of atpA. In the case of somatic hybrids between N. Langsdorffii and N. Glauca, three new fragments were detected in addition to parental fragments. These new fragments can be explained by homologous recombination within the coding region of atpA. These results show that the coding region of atpA is involved not only in intragenomic homologous recombination but can also be involved in homologous recombination between two parental mitochondrial genomes of somatic hybrids.

以非放射性标记的水稻rDNA为探针，结合从植物组织中提取DNA的方法和Southern杂交技术，建立了一种简单有效的鉴定细胞融合后杂种的方法。100毫克的叶或胼胝体组织足以鉴定杂种，材料的量可以减少到10毫克。由于这种方法不仅可以在植株阶段而且可以在愈伤组织阶段选择准性杂种，因此可以在细胞融合产物发育的早期阶段鉴定杂种。在烟草Langsdorffii的情况下，我们鉴定了来自atpA的两个不同的5'和两个不同的3'侧翼区的组合的四个片段。所有的四种类型共享的编码区，这表明这四种类型可以作为同源重组的结果产生的atpA的编码区。相比之下，N. Glauca只产生了一个片段，表明只存在一种类型的atpA。在N. Langsdorffii和N. Glauca中，除了亲本片段外，还检测到三个新片段。这些新的片段可以通过atpA编码区内的同源重组来解释。这些结果表明，atpA的编码区不仅参与基因组内的同源重组，但也可以参与体细胞杂种的两个亲本线粒体基因组之间的同源重组。