甜菜幼苗低温和长日照诱导表达基因的克隆和序列分析

 以代表性差异分析cDNA-RDA(representational difference analysis of cDNA)方法获得在低温和长日照诱导甜菜(Beta vulgaris L)幼苗茎尖中差异表达的基因片段DP3，并进行了cDNA的末端快速扩增（rapid amplification of cDNA ends），经RT-PCR 扩增和基因组DNA中PCR扩增，克隆测序后获得1个全长609 bp的cDNA和855 bp的DNA序列，分别命名为Ty7Br600 和Ty7Br900，并在GenBank注册，登录号分别为AY324115和AY324114。在GenBank中比较未发现同源序列，可能为新基因。序列分析发现，Ty7Br600具有1个编码136个氨基酸的开放读码框（ORF）, Ty7Br900序列中存在1个内含子。分别以克隆的低温诱导甜菜幼苗cDNA为探针，分别进行Northern印记和Southern印记杂交。结果表明，cDNA差异片段只在低温诱导的甜菜中表达，在甜菜基因组中以2个拷贝或低拷贝形式存在。

Cloning and Sequence Analysis of Cold and Longday Induced Expressed Genes from Sugar Beet (Beta vulgaris L.) Seedlings

s:cDNA representational difference analysis (cDNA-RDA) was applied to clone genes specifically expressed in cold and longday induced sugar beet (Beta vulgaris L.).One cDNA fragment was obtained after 3 rounds of subtractive hybridization and amplified by 3'-RACE and 5'-RACE.A sequence of 609 bp cDNA named Ty7Br600 and 855 bp DNA named Ty7Br900 were obtained respectively by RT-PCR and PCR after cloning and sequencing.Blasting in GenBank shows no homology with them,suggesting that it may be a novel gene of sugar beet.Results of Northern blotting and Southern blotting showed that the cDNA clone was only expressed in the cold induced sugar beet and low copy in sugar beet genome.