长穗偃麦草DREB类基因EeAP2.2的克隆与序列分析

Cloning and Sequence Analysis of DREB-like Gene EeAP2.2 in Thinopyrum Ponticum

结合RACE和Genome Walking技术从十倍体长穗偃麦草中克隆了AP2家族的一个全长cDNA序列,命名为Ee-AP2.2。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%和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。

By combining RACE and Genome Walking techniques, a full-length cDNA sequence of AP2 family was cloned from Thinopyrum Ponticum （2n = 70） , and named as EeAP2. 2. Sequence analysis of EeAP2. 2 revealed a single open reading frame （ORF） of 837bp encoding a protein of 279 amino acids, and a highly conserved AP2 domain. Gene EeAP2. 2 was classified 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 HvDREBI-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% and 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.