不同花色矮牵牛细胞色素b5蛋白的cDNA克隆及序列分析

cDNA Cloning and Sequencing of Cytochrome b5 Protein from Petunia hybrida Strains with Different Flower Colors

以云南不同花色矮牵牛的花瓣为材料 ,提取总RNA ,用Oligo (dT)作为引物反转录合成cDNA第一链。以此为模板 ,用根据国外报道的矮牵牛细胞色素b5蛋白的cDNA序列设计合成的引物进行PCR扩 ,均扩增到一条约 4 5 0bp的片段 ,分别克隆到pGEM -T载体上。对重组克隆进行序列分析 ,结果表明所克隆到的矮牵牛细胞色素b5蛋白的cDNA的编码区均含有 4 4 7个核苷酸 ,编码 149个氨基酸残基 ,与国外报道的一致 ;但其核苷酸及氨基酸的序列与国外报道的有所不同 ,即与国外的相比 ,紫红色、蓝紫色矮牵牛中的该cDNA的核苷酸有 1个不同 ,而氨基酸完全相同 ;粉红色、白色矮牵牛中的有 3个核苷酸不同 ,并导致了2个氨基酸的不同。暗示该基因对花色的调控可能与其编码cDNA的一级结构有关。

Total RNAs were extracted from the petals of Petunia hybrida strains with different flower colors.Using Olig(dT) as pimer,the total RNAs were synthesized into the first strand of cDNA by reverse transcription reaction.By Polymerase Chain Reaction(PCR)amplification,we obtained a cDNA fragment in length of about 450bp from different Petunia strains.The primes used for PCR amplification were designed according to the cDNA encoding region of cytochrome b5 protein that were reported previously.The PCR products were cloned into pGEM T vector,respectively.We analyzed the sequences of different recombinant plasmids and compared them with the reported cDNA sequence of Cyt b5.We found that their encoding areas all contain 447 nucleotides and encode 149 amino acid residues.However,the cDNAs from violet and blue Petunia have one different nucleotide,but without different amino acid residues from the reported cDNA;those from pink and white Petunia have three different nucleotides and two different amino acid residues from the reported cDNA.These results imply that the Cyt b5 gene regulation on flower color may be relevant to the primary structure of its encoding cDNA.