花生△9-硬脂酰-ACP脱氢酶基因(SAD)的序列分析

摘要

The peanut (Arachis hypogaea L.) cultivars are an allotetraploid consisting of A and B genomes. △9-Stearoyl-ACP desaturase (SAD) is a key enzyme that catalyzes the conversion of stearoyl-ACP to oleoyl-ACP, and finally controls the content of oleic acid as well as the proportion of saturated to unsaturated fatty acids. By using the primers based on the peanut cDNA sequence of SAD (AF172728), genomic DNAs were amplified from the wild diploid species A. Duranensis and A. Ipaensis, and from the cultivated accessions of peanut Fcnghua 2, Shanhua 7 and Puyangdatuoyang, respectively. Two isoforms of the genomic SAD were identified and named as FhgSAD-1 and FhgSAD-2 from the cultivated peanut accessions. In addition, two isoforms of SAD cDNA were isolated and named as FhrSAD-1 and FhrSAD-2 from Fenghua 2. Comparison of the genomic sequences with cDNAs revealed that there were two introns in the SAD genomic sequences. Sequences alignment showed that the similarity between FhgSAD-1 and FhgSAD-2 in nucleotide level was over 97.5%, with 69 different sites in total, including 62 SNP sites and six variation sites of endonuclease recognition. The cDNA sequence similarity between FhrSAD-1 and FhrSAD-2 was 98.6%, with 98.9% nucleotides identity in coding region. Deduced amino acid sequences revealed that only one difference occurred in serine gathering area of "PSSSSSSSSSSFSL30. gSAD-1 shared 99.9% nucleotide sequence homology with gSAD-A, while gSAD-2 was the same as gSAD-B. According to the phylogenetic tiee, it is assumed that gSAD-1 and gSAD-2 may come from the A and B genome, respectively. The results revealed the characteristics of SAO sequences from different genomes of peanut, and provided important basis for exploring gene expression regulation and fatty acid component improvement in peanut seeds.%利用同源克隆技术获得花生区组二倍体野生种A.duranensis和A.ipaensis的△9-硬脂酰-ACP脱氢酶基因SAD(命名为gSAD-A和gSAD-B)及3个栽培品种的SAD,每个栽培品种有2个SAD(命名为gSAD-1和gSAD-2).同时获得丰花2号SAD的两条全长cDNA(命名为FhrSAD-1和FhrSAD-2).丰花2号的FhgSAD-1和FhgSAD-2均含有2个内含子,二者同源性97.5％,共有69个变异位点,其中62个是SNP位点、6个特异性酶切位点.FhrSAD-1和FhrSAD-2间核苷酸序列同源性98.6％,其中编码区序列同源性98.9％,共有12个变异位点,编码的Ah-SAD2氨基酸序列与Ah-SAD1相比在N端的17PSSSSSSSSSSFSL30丝氨酸聚集区少一个丝氨酸.gSAD-1和gSAD-A同源性为99.9％,存在4个SNP位点；gSAD-2和gSAD-B同源性为100％.推测gSAD-1和gSAD-2分别来自花生栽培品种的A、B2个染色体组.研究明确了花生不同染色体组SAD的序列特征,为进一步探讨SAD的表达及其在控制花生籽仁脂肪酸组分中的作用提供了重要的参考.