柑橘钙调蛋白cDNA的克隆及序列分析

Cloning and Sequence Analysis of Citrus Calmodulin cDNA

以温州蜜柑(CitrusunshiuMarc.)幼果cDNA第1链为模板,参照大麦钙调蛋白基因序列(AccessionNo.M27303)合成5端和3端引物,以PCR的方法扩增得到柑橘钙调蛋白cDNA,将其克隆到PMD18-T载体上并进行测序。序列分析表明,柑橘钙调蛋白cDNA全长453bp,共编码148个氨基酸,与大麦和大豆钙调蛋白基因的同源性均高达83%以上,所编码氨基酸序列同源性达97%以上。以该cDNA作探针进行点杂交,得到良好的杂交信号。

Using the first strand cDNA from the immature fruits of Citrus unshiu Marc. as template, we amplified and cloned the cDNA of Citrus calmodulin gene in this study. The PCR primers were designed according to the sequences of barley calmodulin gene obtained from GenBank （Accession No. M27303 ）. The PCR product was cloned into PMD18-T vector and then sequenced. Sequence analysis indicated that the cDNA of Citrus calmodulin gene contains 453 nucleotides coding for 148 amino acid residues. The nucleotide sequences and their deduced amino acid sequences of the Citrus calmodulin cDNA show an identity of above 83% and 97%, respectively, to those of barley and soybean. Southern dot blot analysis using the cloned cDNA as probe confirmed the results of sequencing analysis.