摘要
‘长选3S’是由‘培矮64S’通过辐射诱变选育出的长穗颈温敏核不育水稻,以‘长选3S’二核花粉期穗颈节间作为目标群体,‘培矮64S’二核花粉期穗颈节间作为对照群体,进行抑制差减杂交,用经过‘培矮64S’cDNA差减的‘长选3S’cDNA构建了一个含有大约130个独立克隆的差减文库;采用差减前的‘培矮64S’cDNA和‘长选3S’cDNA以及正向/反向差减杂交后的cDNA为模板标记探针,对随机挑取的96个重组质粒进行差示筛选,获得了20个阳性候选克隆。从这些阳性候选克隆中随机挑取了8个进行Northern blot分析,证实其中1个候选克隆代表了在‘长选3S’穗颈节间中特异表达的基因,序列分析和同源性比较表明它与信号传导有关。这一在‘长选3S’穗颈节间特异表达的cDNA片段有助于进一步揭示其穗颈节间伸长的分子机制,并为水稻不育系的遗传改良提供有用的素材。
‘Changxuan 3S＇ is a long-spike neck and thermo-sensitive genic male sterility rice variety bred with the rice variety ‘Peiai 64S＇ by 350 GY60 y-ray radiation mutagenesis. A suppression subtractive hybridization between the internodes of ‘Changxuan 3S＇ at the biuncleate pollen stage as the testers populations and the internodes of ‘Peiai 64S＇ at the binucleate pollen stage as the drivers was conducted and then a subtracted cDNA library composed of 130 independent clones was constructed with cDNA of ‘Changxuan 3S＇ modified with cDNA of ‘Peiai 64S＇ ;randomly-chosen 96 recombinant plasmids were screened with cDNA of ‘Peiai 64S＇ and cDNAs of ‘Changxuan 3S＇ before suppression subtraction hybridization and their cDNAs after forward and backward-direction suppression subtractive hybridizations as the template-marking probes and twenty positive candidate clones were chosen;eight candidate clones taken from these positive candidate clones were analyzed by Northern blotting and one of them were proved to represent the specific expression gene of the internodes beneath the spike of ‘Changxuan 3S＇ ;the sequencing and homology comparison of the gene indicated that it was related to signal transmission. This specifically expressed cDNA fragment is helpful to further reveal the elongation molecular mechanism of the internodes beneath the spikes and has laid clown a foundation for improving sterility rice lines.
出处
《西北植物学报》
CAS
CSCD
北大核心
2006年第11期2257-2264,共8页
Acta Botanica Boreali-Occidentalia Sinica
基金
湖南省教育厅资助项目(04C417)
关键词
抑制差减杂交
温敏核不育水稻
隐性长穗颈
穗颈节间
CDNA文库
特异表达
suppression subtractive hybridization
thermo-sensitive genic male sterility rice
recessive elongated spike neck
internode beneath spike
cDNA library
specific expression
