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Position: Home > Articles > Cloning and Bioinformatics Analysis of Gene Sequence Encoding ACC Synthase from Paeonia suffruticosa Journal of Henan Agricultural Sciences 2013,42 (3) 100-102,106

牡丹ACS基因克隆及序列分析

作  者：
高水平;魏春梅;刘改秀;王丽娜;孔祥生;范丙友
单  位：
河南省洛阳国家牡丹园;河南科技大学农学院;河南科技大学林学院;河南省洛阳市牡丹研究所
关键词：
牡丹;ACC合成酶基因;基因序列;克隆;生物信息学分析
摘  要：
ACC合成酶(ACS)是乙烯生物合成的关键酶.为了明确牡丹ACC合成酶基因的结构,基于报道的牡丹ACS cDNA (DQ337250)序列,设计一对特异性PCR扩增引物,以洛阳红牡丹(Paeonia suf futicosa Andr.)新鲜叶片总DNA为模板,用高保真DNA聚合酶KOD-Plus成功地扩增出长度约为2 200 bp的PCR产物;用pMD18-T载体对PCR产物进行了克隆,测序结果表明,牡丹ACS基因序列全长2 251 bp,含4个外显子和3个内含子,剪接位点均为保守的5 '供位GU与3'受位AG模式,4个外显子分别居于1-174、267-398、483-643、1 240-2 251 bp;序列已提交至GenBank数据库,其登录号为FJ769773;牡丹ACS蛋白由492个氨基酸组成,与苹果、枇杷、西洋梨、天竺葵等ACS的相似性达74%~76%;生物信息学分析预测牡丹ACS蛋白的分子量为54.9 kD,等电点为6.8,是一个定位于细胞质的不稳定的非分泌亲水蛋白.
译  名：
Cloning and Bioinformatics Analysis of Gene Sequence Encoding ACC Synthase from Paeonia suffruticosa
摘  要：
ACC synthase is the key enzyme in ethylene biosynthsis.Based on reported cDNA sequence(DQ337250) encoding ACC synthase from Paeonia suffruticosa Andr.,a pair of specific PCR primers was designed.With the total DNA extracted from fresh leaves of Paeonia suffruticosa cv.Luoyanghong as DNA template,PCR products about 2 200 bp in length were successfully amplified by high-fidelity KOD-Plus DNA polymerase.Then the PCR products were cloned to pMD18-T vector.The sequencing result showed that the full-length of the genomic sequence of ACSisolated from Paeonia suffruticosa Andr.was 2 251 bp containing 4 extrons and 3 introns,and all the splicing sites were conservative at 5′ donor with GU and 3′ receptor with AG.The 4 extrons were located from 1 to 174,267 to 398,483 to 643 and 1 240 to 2 251 bp,respectively.The sequence information has been submitted to GenBank with the accession number of FJ769773.The ACS protein contained 492 amino acids and its similarities to other ACS in Malus*domestica,Eriobotrya japonica,Pyrus communis and Pelargonium*hortorum were up to 74%-76%.Bioinformatic analysis revealed that the molecular weight and pI of ACS was 54.9 kD and 6.8,respectively.It also showed that ACS was an unstable,non-secretory hydrophilic protein,being located in cytoplasm.

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