摘要
以红色蝴蝶兰为材料,利用SMARTer技术构建蝴蝶兰全长cDNA文库,分析从文库中获得的EST序列,获得蝴蝶兰类黄酮3′羟化酶基因(PhF3′H)的全长,并对其进行生物信息学分析;采用实时荧光定量PCR方法,研究PhF3′H基因在红色蝴蝶兰和黄色蝴蝶兰花瓣中的表达情况。结果表明:(1)从构建的蝴蝶兰文库中随机挑选24个单克隆进行测序,结果显示均含有插入片段,重组率为100.0%,其中扩增片段长度在750bp以上的克隆有22个,占91.7%。(2)获得的PhF3′H基因编码的氨基酸序列与大丽花(Dahlia pinnata,ADB_77826.1)、毛油点草(Tricyrtis hirta,BAH_22519.1)等多种观赏植物F3′H编码的氨基酸序列均具有较高的一致性。(3)实时荧光定量PCR检测结果显示,PhF3′H在红色蝴蝶兰中的表达量大约是黄色蝴蝶兰的19倍,说明该基因对蝴蝶兰花青素苷的积累有重要影响。该研究结果为蝴蝶兰新基因的挖掘及花色育种奠定了理论基础。
cDNA library was constructed from the red Phalaenopsis petals by SMARTer technique.The F3′ H unigene in Phalaenopsis was obtained through analyzing the EST sequences from library and it was analyzed using bioinformatics.At last,the expression patterns of F3′H in red and yellow Phalaenopsis petals were analyzed using real-time PCR.The results indicated that：（1）The recombinant percentage of the library was determined as 100.0% by random screening 24 clones.Among these clones,there were 22 inserts that were longer than 750 bp,up to 91.7%.（2）The amino acid sequence encoded by PhF3′H obtained from the library had a high identity with that of Dahlia pinnata（ADB_77826.1）,Tricyrtis hirta（BAH_22519.1）and other ornamental plants.（3）The result of RT-PCR showed that PhF3′H expression abundance of red Phalaenopsis was about 19 times than that of the yellow Phalaenopsis,which proved that the gene plays an important role in the accumulation of anthocyanins in Phalaenopsis.In brief,the results will provide a theoretical basis to Phalaenopsis gene mining and color breeding.
出处
《西北植物学报》
CAS
CSCD
北大核心
2013年第9期1731-1738,共8页
Acta Botanica Boreali-Occidentalia Sinica
基金
国家自然科学基金(31101574)
南京农业大学青年科技创新基金(KJ2011008)
