中国博士后科学基金(No.20060390479),河北农大博士启动基金(200805),转基因生物新品种培育重大科研专项基金(No.2009ZX08002-012B)
由于一些基因的特殊碱基序列限制,使得应用一种技术获得基因的全长cDNA序列比较困难。本研究结合RACE和Genome Walking技术从十倍体长穗偃麦草(Elytrigia elongata,2n=70)中克隆了AP2家族的一个全长cDNA序列,命名为EeAP2.2。序列分析表明,该基因具有一个837bp的开放阅读框,编码279个氨基酸残基,含有一个保守的AP2结构域,是AP2大家族的一个新成员。该基因编码的氨基酸序列与GenBank已有的普通小麦AP2家族两个同源基因编码蛋白TaDREB1和TaDREBW50(登录号分别为:AAL01124.1和AAY44605.1)具有98%的氨基酸序列一致性, 与大麦AP2蛋白HvDREB1-a(登录号AAY25517.1),高羊茅AP2蛋白FaDREB2A (登录号CAG30547.1) 及水稻OsDREB2.2(登录号AY064403)的氨基酸序列一致性分别为 93%、86%、69%。说明该基因与小麦AP2家族基因的同源性最高。本研究除获得了长穗偃麦草一个重要抗逆转录因子基因EeAP2.2的全长cDNA序列外,也提供了一种快速、有效克隆功能基因的方法。
Due to the restriction of special nucleotide sequences in some genes, it’s difficult to obtain their full-length cDNA sequences by using only one method. In this paper, by combining RACE and Genome Walking techniques, a full-length cDNA sequence of AP2 family was cloned from Elytrigia elongata (2n=70), and named as EeAP2.2. Sequence analysis of EeAP2.2 revealed a single open reading frame (ORF) of 837 bp encoding a protein of 279 amino acids, and a highly conserved AP2 domain which classifies EeAP2.2 as a new member of AP2 superfamily. Further comparison analysis through NCBI blast showed that EeAP2.2 has 98%, amino sequence identities with TaDREB1 (Accession no. AAL01124.1) and TaDREBW50 (Accession no. AAY44605.1) from Triticum aestivum, and its amino sequence identities with HvDREB1-a (Accession no. AAY25517.1) from Hordeum vulgare, FaDREB2A (Accession no. CAG30547.1) from Festuca arundinacea and OsDREB2.2 (Accession no. AY064403) from Oryza sativa are 93%, 86%, 69%, respectively. The result indicates that EeAP2.2 has the highest homology with AP2 family members from Triticum aestivum. This study also provided a fast and effective method for cloning full length cDNA sequences in plants.
默韶京,刘桂茹,郎明林.长穗偃麦草DREB类基因EeAP2.2 的克隆与序列分析[J].植物遗传资源学报,2011,12(5):764-769.
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