长春花丙二烯氧化物合酶基因的克隆与表达

Cloning and Expression of Allene Oxide Synthase Gene from Catharanthus roseus

利用RT-PCR和RACE相结合的方法,从长春花中克隆了丙二烯氧化物合酶(AOS)基因。结果显示:长春花AOS基因(CrAOS)cDNA全长为2 118bp,包括5′和3′非翻译区,polyA尾和一个长1 638bp的开放阅读框,其基因组中不含内含子;CrAOS基因编码的蛋白含545个氨基酸。多重比对表明CrAOS蛋白与其他的AOS蛋白具有较高的相似性,CrAOS蛋白序列中含有AOS家族应有的保守氨基酸残基。Southern杂交表明:CrAOS基因在长春花中为低拷贝。qRT-PCR结果显示:CrAOS在各个组织均有表达但表达量存在差异,在老叶中最高,在幼花中表达最低。对长春花幼苗进行不同处理,结果表明:伤害、低温、甲基茉莉酸、乙烯利处理等可使CrAOS基因表达量显著提高,水杨酸处理对基因表达影响不大。

In this study, a full-length cDNA of allene oxide synthase （AOS） gene （named as CrAOS, JQ364955） was cloned from Catharanthus roseus. The gene was 2 118 bp in size containing an open reading frame （1 638 bp） encoding 545 amino acids. Comparative and bioinformatic analysis revealed that the de- duced protein of CrAOS was highly homologous to AOSs from other plant species. Southern blot analysis revealed that it was a low-copy gene. Real-time Quantitative PCR （qRT-PCR） analysis showed that CrAOS mRNA accumulated most abundantly in old leaves and least in young alabastrums. The qRT-PCR analysis revealed that wound, low temperature, methyl jasmonic acid, ethylene treatments significantly enhanced CrAOS transcript expression,and salicylic acid had no influence.