甘蓝型油菜Tic21基因克隆、表达及拷贝数鉴定

Isolation andexpression analysis of Brassica napus L.Tic21 gene and its copy number determination

根据拟南芥Tic21基因编码区设计引物,在甘蓝型油菜中扩增并克隆到其cDNA同源基因,命名为BnTic21(GenBank登录号HQ613273),测序结果显示该基因含有5个外显子,4个内含子,cDNA编码区大小为885 bp,编码294个氨基酸,蛋白理论分子量约为31.24kD,等电点11.10.与拟南芥、豌豆等物种氨基酸序列相似性在60.40%～87.75%之间.实时定量PCR对其不同组织特异性表达分析表明:BnTic21在油菜不同部位均有表达,茎尖最高,子叶、真叶、下胚轴次之,根表达量最低.用实时定量PCR检测表明BnTic21在甘蓝型油菜基因组中约有2个拷贝.

A cDNA sequence with primers based on Arabidopsis （Arabidopsis thaliana L. ） was obtained from Brassica napus L. , which was named as BnTic21 （GenBank accession number HQ613273） and could be delineated into 5 exons and 4 introns. An ORF （Open Reading Frame） of 885 bp encoded a 294 predicted amino acid residues with a molecular weight of 31.24 kD, and an isoelectric point of 11. 10. The amino acid sequence similarity between the BnTic21 gene and some genes from other plants in Gen- Bank were 60.40~87.75~. Real-time PCR was analyzed in different tissues. The results showed that the BnTic21 gene＇s expression level in various organs were found as follows： shoot apex〉 cotyledonS〉 leaf〉 hypocotyl 〉 root. Real-time PCR analysis showed that there were about two copies of Bn- Tic21gene in the genome of Brassica napus L. Key words： Brassica napus L. , Tic21, Real-time PCR, copy number