烟粉虱MED隐种蜕皮激素受体基因cDNA克隆、转录与组织表达

Molecular cloning,sequence analysis and expression of ecdysone receptor gene from Bemisia tabaci Mediterranean(Hemiptera: Aleyrodidae)

蜕皮激素受体(ecdysone receptor,Ec R)是调控昆虫蜕皮、变态和生殖等过程中基因表达的重要调控因子。为研究Ec R在烟粉虱Bemisia tabaci生长发育中的作用,利用RT-PCR和RACE等技术扩增了烟粉虱MED隐种Ec R基因的c DNA全长序列,并利用实时荧光定量PCR技术检测其在烟粉虱不同发育时期相对表达量的变化。Bt Ec R c DNA序列全长2 844 bp,含有一个1 518 bp的开放阅读框,共编码505个氨基酸,预测蛋白分子量为56 k D,等电点为6.43,含有特殊结构功能域:DNA结合域(DBD_Ec R)和配体结合域(LBD_Ec R),该序列编码的蛋白质序列与其它已报道的昆虫Ec R蛋白序列具有很高的相似性,命名为Bt Ec R(Gen Bank登录号:KR534774)。Bt Ec R在MED隐种若虫、雌成虫各发育时期均有表达,若虫期在伪蛹前期达到最高值,成虫期呈现先升高后降低的趋势,且在羽化后第7天达到最高值,约为羽化第1天的23倍。研究结果为揭示Bt Ec R在烟粉虱整个生长发育过程中的作用提供了重要依据。

Ecdysone receptor（ Ec R） is a dominant regulator of development, metamorphosis and reproduction in insects. In order to define the function of Ec R gene in Bemisia tabaci Mediterranean（ MED）,the full-length c DNA sequence of an Ec R gene was cloned by both reverse transcription polymerase chain reaction（ RT-PCR） and rapid amplification of c DNA ends（ RACE） methods,and the relative expression levels at different developmental stages of MED was analyzed using real-time quantitative PCR（ q PCR）. The complete c DNA（ 2 844 bp） of Ec R gene in B. tabaci MED species complex contained an open reading frame（ ORF） of 1 518 bp,encoding 505 amino acids,with a molecular weight of 56 k D and a theoretical isoelectric point of 6. 43. This gene had two special domains：the DNA-binding domain of ecdysone receptor（ DBD_Ec R） and the ligand-binding domain of ecdysone receptor（ LBD_Ec R）. Multiple sequence alignment revealed that the deduced amino acid sequence had high identity with Ec R from other insect species. This c DNA was named as Bt Ec R（ Gen Bank accessionno. KR534774）. q PCR results indicated that Bt Ec R transcripts were detected at all instars of nymph and developmental stages of female adults. The highest relative expression level of Ec R gene was detected at pseudo-pupae stage among different instar nymphs. Peak level was detected at 7 d after eclosion,which was about 23 times the level at 1 d after eclosion. The present results provided valuable information for revealing the role of Bt Ec R underlying the development of whitefly species.