摘要
以野生大豆Glycine soja叶片总RNA为模板,根据其他双子叶植物DREBP/AP2结构域的保守氨基酸序列设计简并引物,通过降落和巢式PCR进行扩增,获得1条190 bp的cDNA片段。以此序列设计引物,采用RACE扩增3’和5’末端未知序列,最终克隆到野生大豆DREB全长基因(GsDREB)。该基因1 191bp编码395个氨基酸,基因内部没有内含子。氨基酸分析表明,其N-末端具有核定位信号(nuclear localiza-tion signal,NLS),并具有由58个氨基酸编码的、能够与DNA结合的保守区域-DREBP/AP2结构域。通过多序列比对分析,该基因与拟南芥Arabidopsis thalianaDREB2C氨基酸序列相似性最高,推测其为DREB2亚家族的成员。
The total RNA in leaf of Glycine soja was treated as cyclostyle. The degenerate primers were designed based on the conservative amino acids sequence of DREBP/AP2 domain from other dicotyledonous. A cDNA fragment of 191bp from G. soja was obtained by touchdown PCR and nested- PCR. Based on this sequence, the nest primers were designed, than 3＇ and 5＇ cDNA sequences were obtained by 3＇and 5＇ rapid amplification of cDNA ends PCR （RACE-PCR）, and the complete DREB gene was constructed in G. soja （designated as GsDREB）. The whole sequence of GsDREB was 1191 bp open reading frame （ORF） encoding a 395 amino acids, without any intron. Amino acid analysis indicated that, the predicted GsDREB protein contained a N-terminal nuclear localization signal （NLS）, and a putative AP2 DNA-binding domain containing 58 conserved amino acids. Alignment of multiple showed that GsDREB had much similarity with AtDREB2, so it might be a member of DREB2 subgroup of DREB family.
出处
《草业科学》
CAS
CSCD
北大核心
2009年第8期17-23,共7页
Pratacultural Science
基金
国家"973"重点基础研究前期专项资助项目(2003CCA03500)
国家自然科学基金资助项目(30471059)
关键词
野生大豆
DREB
同源克隆
RACE
非生物胁迫
Glycine soja
dehydration responsive element binding protein （DREB）
homologous cloning
RACE
abiotic stress
