蜡梅结瘤素基因CpNOD的克隆与表达分析

Cloning and Expression Analysis of the CpNOD Gene from Chimonanthus praecox(L.)

根据已构建的蜡梅cDNA文库中得到的片段,通过cDNA末端克隆得到了一个早期结瘤素基因,命名为Cp-NOD(GenBank登录号:JQ622290),该基因cDNA序列全长611个碱基,包含一个71bp 5-UTR区,一个333bp的开放阅读框ORF和一个207bp的3-UTR区,该基因由111个氨基酸组成,其中丙氨酸组分为多,占18.2%,序列比对分析表明,CpNOD具有典型的蛋白质家族ENOD93特征,属于ENOD93家族早期结瘤素蛋白.实时荧光定量PCR表达检测显示,CpNOD在蜡梅根、茎、花器中都有表达,其中以花中瓣表达最高;在花的不同阶段表达结果是从萌动期、花蕾期、盛开期到衰败期,表达量呈正相关,以衰败期表达量最高.转录表达分析结果推测,CpNOD在蜡梅的开花生理和氮元素响应方面起着积极作用.

Based on the eDNA library constructed from Chimonanthus praecox L. , a new early nodulin gene was isolated, using the rapid amplification of eDNA ends （RACE） technology, and the new gene was designated as CpNOD （GenBank accession No. JQ622290）. The full-length eDNA of CpNOD was 611 bp, with an open reading frame （ORF） of 333 bps, a 71-bp 5＇ untranslated region （5＇-UTR）, and a 207-bp 3＇-untranslated region （3＇-UTR）. CpNOD encoded a Ill-amino-acid polypeptide, in which Ala was the richest and its abundance reached 18.2%. Sequence alignments and phylogenetic analysis revealed that CpNOD belonged to the ENOD93 superfamily of early nodulin protein gene, which had a typical motif character. Real-time quantitative PCR showed that CpNOD constitutively expressed in roots, stems and flowers under normal conditions, and it expressed at the highest level in the middle flaps. During the blooming stage, the expression of CpNOD was correlated with the maturity of the flower and it expressed at the highest level in the withering period. The gene expression analysis results suggested that CpNOD might play a positive role in blooming physiology of C. praecox （L.） and N responses.