本文關鍵詞： 矮牽牛 八氫番茄紅素脫氫酶基因 基因沉默 shRNA　出處：《園藝學報》2017年02期 　論文類型：期刊論文

【摘要】：以矮牽牛(Petunia hybrida)自交系‘Mitchell Diploid’(MD)為材料,通過3′RACE和5′RACE獲得了其八氫番茄紅素脫氫酶基因(PhPDS)的cDNA序列,進而通過PCR擴增出編碼區(qū)的基因組序列。分析結果顯示,PhPDS的cDNA序列全長2 182 bp,編碼區(qū)序列1 746 bp,編碼582個氨基酸。推測的蛋白質序列具有類胡蘿卜素脫氫酶的保守結構域特征。編碼區(qū)的基因組序列長7 609 bp,結構與番茄和擬南芥等PDS基因相似,具有14個外顯子和13個內含子。利用博德研究所的小發(fā)夾RNA(shRNA)設計程序設計了兩個針對PhPDS的shRNA,構建植物表達載體轉化矮牽牛后,從其中一個載體的轉化子中獲得白色愈傷組織和幼枝,對白色幼枝的RT-PCR檢測表明,其PDS基因的cDNA累積量減少,表明shRNA基因沉默技術能在矮牽牛中應用。以PDS基因為靶基因比用花色素合成基因研究基因沉默技術能更快地分析基因沉默的效果,全長序列的獲得將有助于PDS基因在矮牽�；虺聊夹g研究中的應用。
[Abstract]:Petunia hybrida, an inbred line, was used as material. The cDNA sequence of its octahydrolycopene dehydrogenase gene, PhPDS, was obtained by 3H race and 5H race. Then the genomic sequence of the coding region was amplified by PCR. The results showed that the cDNA sequence of PhPDS was 2 182 BP in length and the coding region was 1 746 BP. It encodes 582 amino acids. The deduced protein sequence has the conserved domain of carotenoid dehydrogenase. The genome sequence of the coding region is 7 609 BP long. The structure is similar to the PDS gene of tomato and Arabidopsis thaliana. With 14 exons and 13 introns, two shRNA targeting PhPDS were designed using the Bod Institute's small hairpin RNA-shRNAs. After the plant expression vector was constructed and transformed into petunia, the white callus and young branches were obtained from the transformants of one of the vectors, and the RT-PCR detection of white young shoots showed that. The cDNA accumulation of PDS gene decreased. The results showed that shRNA gene silencing technique could be used in Petunia hybrida, and the effect of gene silencing could be analyzed more quickly by using PDS gene as target gene than by using anthocyanin synthesis gene to study gene silencing. The acquisition of full-length sequence will be helpful to the application of PDS gene in the study of petunia gene silencing.
【作者單位】： 西南大學園藝園林學院重慶市花卉工程技術中心南方山地園藝學教育部重點實驗室;
【基金】：中央高校基本科研業(yè)務費專項資金項目(XDJK2016D062) 重慶市科委項目(cstc2014yykf B80008)
【分類號】：S681.9
【正文快照】： 類胡蘿卜素是光合作用過程中的輔助色素,能夠猝滅單線態(tài)氧從而保護葉綠素不受到傷害(Qin et al.,2007)。八氫番茄紅素脫氫酶(phytoene desaturase,PDS)是類胡蘿卜素合成途徑中的關鍵酶,PDS基因功能的失活會導致類胡蘿卜素不能有效合成,葉綠素在光照條件下會被破壞,使本應呈綠

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