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为研究甜樱桃在高温下胚囊发育的分子机理,以甜樱桃品种‘拉宾斯’为试材,采用RT-PCR结合RACE方法克隆胚囊发育相关的MADS-box SEP类基因c DNA全长,使用半定量PCR法进行特异性表达分析,通过温室和露地环境PaMADS4实时荧光定量分析不同温度下的表达。克隆得到一个基因全长999 bp,命名为PaMADS4(Gen Bank登录号:JQ686726),包含一个735 bp的开放阅读框,编码244个氨基酸;序列比对和进化分析表明,PaMADS4与拟南芥SEP基因家族亲缘关系最近;组织特异性分析表明PaMADS4在四轮花器官中均表达;开花后PaMADS4在露地中的表达量明显大于温室。
In order to study the molecular mechanism of embryo sac development in sweet cherry at high temperature, the full length c cDNA of MADS-box SEP gene was cloned by RT-PCR and RACE using the sweet cherry cultivar ’Labins’ , Specific expression analysis was carried out by semi-quantitative PCR method, and the expression of PaMADS4 was analyzed by real-time fluorescence quantitative analysis of different temperatures through the greenhouse and the open field environment. The full-length cDNA of PaMADS4 was cloned and named as PaMADS4 (Gen Bank accession number: JQ686726), containing a 735 bp open reading frame encoding 244 amino acids. Sequence alignment and phylogenetic analysis showed that PaMADS4 interacts with the Arabidopsis SEP gene Family close relationship; tissue specificity analysis showed that PaMADS4 were expressed in four flower organs; expression of PaMADS4 in the open field after flowering was significantly greater than the greenhouse.