铁皮石斛HSP70基因的克隆及冷胁迫表达分析

Cloning and expression analysis of HSP70 gene from Dendrobium officinale under low temperature stress

目的:揭示铁皮石斛热激蛋白HSP70基因冷胁迫表达,为耐低温品系选育提供分子生物学基础。方法:根据已获得的HSP70基因片段序列,采用RACE方法从铁皮石斛中克隆到HSP70基因全长cDNA序列。预测其编码蛋白的结构与功能,并运用实时定量PCR进行冷胁迫下表达分析。结果:核苷酸序列分析表明该基因全长2 296 bp,包含一完整的1 944 bp的开放阅读框,编码647个氨基酸。氨基酸序列具有典型的HSP70特征并且与其他植物的HSP70有很高的同源性。冷胁迫表达分析说明该基因确实能被低温诱导表达。结论:首次克隆获得受低温诱导表达的铁皮石斛HSP70基因,为进一步研究铁皮石斛健康栽培与抗寒品系的选育奠定基础。

Objective： To investigate HSP70 gene expression from Dendrobium officinale under low temperature stress, which will provide the molecular biological foundation for breeding the low temperature resistant strain. Method： HSP70 gene full length cDNA was cloned by rapid amplification of cDNA ends （RACE） on the basis of HSP70 gene fragment sequences, and the structure and function of HSP70 gene were deduced. The expression of HSP70 under low temperature stress was detected by RT-PCR. Result： The full length of HSP70 gene cDNA was 2 296 bp containing a 1 944 bp open reading frame （ORF） that encoded a protein of 647 amino acids. Its amino acids sequence had typical HSP70 characteristics and high homology with other plant＇s HSP70. Cold stress expression analysis showed that expression of the HSP70 gene could be induced by low temperature. Conclusion ： The HSP70 gene of D. officina- le was successfully cloned and reported for the first time which proved that the expression could be induced by low temperature. The cloning of HSP70 gene provides a stable foundation for further study of D. officinale cultivation and the breeding of the cold resistance strains.