茶陵野生稻冷响应基因OrCrZFl的克隆与生物信息学分析

Cloning and Bioinformatics Analysis of a Novel Cold Responsive Gene (OrCrZFl) in ‘Chaling’ Common Wild Rice

OrCrZFl基因是基于水稻基因表达芯片分析从茶陵野生稻中筛选出的一个受低温诱导、编码类锌指蛋白的基因。以茶陵野生稻为材料,用RT-PCR方法扩增获得了包含其完整ORF的cDNA克隆。根据其ORF进行预测,该基因编码一个包含492个氨基酸残基的蛋白,其理论分子量为51.895 kD,pI=6.21。经蛋白质相似性比对,其编码蛋白与日本晴第8号染色体上Os08g0536300基因编码的蛋白质(GenBank登录号:XP_015649825.1,zinc finger protein CONSTANS-LIKE 14)氨基酸序列一致性为98.37%;其第145位到第492位氨基酸序列与籼稻93-11的预测蛋白(EEC83950.1,hypothetical protein OsI_30045)一致性为96.55%;其第16位到第492位氨基酸序列与谷子(Setaria italica)、高粱(Sorghum bicolor)、玉米(Zea mays)、粗山羊草(Aegilops tauschii subsp.tauschii)及穿心莲(Panicum hallii)编码的蛋白(zinc finger protein CONSTANS-LIKE 14/15)一致性为67.35%~72.47%。对其可能的启动子区域序列分析,发现多个可能与逆境或逆境激素反应有关的顺式作用元件。基于以上结果,我们认为该基因为一新的野生稻耐冷候选基因。

OrCrZFl is a novel cold responsive gene screened from‘Chaling’common wild rice(CWR)based on the global genome expression profiling of‘Chaling’CWR subjected to cold stress.The sequence of its cDNA containing a complete ORF(open reading frame)was cloned by RT-PCR by using‘Chaling’CWR as material.Sequence analysis showed that the cDNA of the gene codes a protein of 492 amino acid residues with M.W.=51.895 kD and pI=6.21.Performing Blastp in GenBank showed that the sequence of its deduced amino acids was 98.37%identical with that of a protein(zinc finger protein CONSTANS-LIKE 14,GenBank accession:XP_015649825.1)encoded by Os08g0536300 located at chromosome 8 of japonica rice Nipponbare,the sequence of its deduced amino acids from 145 th to 492 nd(the last)was 96.55%identical with that of hypothetical protein OsI_30045 of indica rice 93-11,and the sequence of its deduced amino acids from 16 th to 492 nd(the last)was 67.35%~72.47%identical with those of similar proteins(zinc finger protein CONSTANS-LIKE 14 or zinc finger protein CONSTANS-LIKE 15)of Setaria italica,Sorghum bicolor,Zea mays,Aegilops tauschii,and Panicum hallii.Analysis of the putative promoter region for candidate cis-elements by using PlantCARE software identified some matches related to cold stress or other stresses or stress phytohormone responses.Based on the above results,we thought that OrCrZFl is a novel candidate cold tolerance gene in wild rice.