大叶落地生根谷氨酸脱羧酶基因(KdGAD)的克隆与表达

c DNA cloning and expression analysis of glutamate decarboxylase gene(KdGAD) in Kalanchoe daigremontiana

为研究大叶落地生根中胎生苗发育的分子机制,利用RACE-PCR技术从大叶落地生根中克隆了1个新的谷氨酸脱羧酶基因(Kd GAD)。该基因开放阅读框长度为1 509 bp,编码502个氨基酸残基,分子量为5.657×104,等电点为5.43。其氨基酸序列与蓖麻的同源性最高,与人参的进化关系最近,含有保守的Ser(S)-x-x-Lys(K)基序和Trp(W)残基。实时荧光定量PCR分析结果表明,该基因在大叶落地生根的茎中表达量最高,且受渗透胁迫(甘露醇处理)的诱导下调表达。

To better understand the molecular mechanisms of plantlet formation involved in Kalanchoe daigremontiana,a glutamate decarboxylase（ GAD） gene,Kd GAD,was identified using rapid amplification of c DNA end（ RACE）PCR. Kd GAD gene consisted of an ORF of 1 509 bp,which was predicted to encode a 502 amino acid residue protein of5.657×104with an isoelectric point of 5.43. The sequence analysis of the Kd GAD revealed homology to Ricinus communis.Phylogenetic analysis showed that Kd GAD protein was most related to Panax ginseng. In addition,Kd GAD protein possessed a Ser（ S）-X-X-Lys（ K） active site and a single tryptophan（ W） residue. Real-time PCR analysis revealed that Kd GAD transcript was expressed highly in stem and down-regulated under osmotic stress（ treatment with mannitol）.