【摘要】：黃烷酮3-羥化酶基因(flavanone 3-hydroxylase,F3H)在植物花青素合成途徑中發(fā)揮重要作用,可作為山茶花色遺傳育種的候選基因。本研究在前期從Gen Bank數(shù)據(jù)庫中篩選得到白花茶樹CsF3H、黃花金花茶CnF3H和紅花浙江紅山茶CcF3H三個基因。然而,它們的分子遺傳與變異信息仍然缺乏,不利于最大正向效應(yīng)基因的選擇利用。本研究系統(tǒng)探討了CsF3H、CnF3H和CcF3H基因的分子特征、系統(tǒng)進(jìn)化和蛋白三維結(jié)構(gòu)。結(jié)果發(fā)現(xiàn)CsF3H、CnF3H和CcF3H三者之間存在高度序列多樣性,共含37個核苷酸和9個氨基酸差異,且與CsF3H和CnF3H相比,CcF3H序列發(fā)生了更多的變異。系統(tǒng)進(jìn)化結(jié)果表明CsF3H、CnF3H和CcF3H蛋白與獼猴桃AcF3H具有共同祖先,但在后期進(jìn)化過程中,CcF3H率先與CsF3H和CnF3H二者發(fā)生分化。序列比對和保守結(jié)構(gòu)域分析發(fā)現(xiàn)CsF3H、CnF3H和CcF3H蛋白均含有一個保守的依賴2-酮戊二酸和二價鐵離子的雙加氧酶超家族特征區(qū)域,該區(qū)域內(nèi)的組氨酸H~(218)和H~(276)以及天冬氨酸D220是Fe~(2+)結(jié)合位點(diǎn),精氨酸R~(286)和絲氨酸S~(288)是2-酮戊二酸的重要結(jié)合位點(diǎn)。本研究還發(fā)現(xiàn)了3個茶屬F3H蛋白的保守結(jié)構(gòu)域中存在一個差異位點(diǎn),與CsF3H和CnF3H第200位異亮氨酸I200相比,CcF3H對應(yīng)氨基酸變異為纈氨酸V~(200),Web logo3.4分析揭示V200在植物F3H蛋白中更為保守�？臻g結(jié)構(gòu)分析顯示CcF3H中氨基酸V~(200的替換導(dǎo)致它和賴氨酸K197的分子作用力增強(qiáng)。本研究結(jié)果表明CcF3H是更為保守的植物F3H蛋白,纈氨酸V200可能是一個重要的功能位點(diǎn)。上述研究也為山茶屬F3H基因提供了新的信息,為今后選擇CcF3H基因作為山茶花色育種最適候選基因提供了理論支撐。
[Abstract]:Flavanone 3-hydroxylase gene (F3H) plays an important role in plant anthocyanin synthesis pathway and can be used as a candidate gene for floral color genetics and breeding of Camellia oleifera. In this study, three genes of white flower tea CsF3H, yellow flower gold tea CnF3H and safflower Zhejiang Hongshan tea CcF3H were screened from Gen Bank database. However, their molecular genetic and variation information is still lacking, which is not conducive to the selection and utilization of the maximum positive effect gene. In this study, the molecular characteristics, systematic evolution and protein three-dimensional structure of CsF3H,CnF3H and CcF3H genes were systematically discussed. The results showed that there was a high degree of sequence diversity between CsF3H,CnF3H and CcF3H, which contained 37 nucleotides and 9 amino acids, and CcF3H sequences varied more than CsF3H and CnF3H. The results of systematic evolution showed that CsF3H,CnF3H and CcF3H proteins had a common ancestor with kiwifruit AcF3H, but in the later evolution process, CcF3H first differentiated from CsF3H and CnF3H. Sequence alignment and conserved domain analysis showed that both CsF3H,CnF3H and CcF3H proteins contained a conserved dioxygenase superfamily region dependent on 2-ketoglutaric acid and divalent iron ions. Histidine H218 and H276 and aspartic acid D220 were Fe~ (2) binding sites, arginine R286 and serine S288were important binding sites of 2-ketoglutaric acid. Compared with CsF3H and CnF3H 200th isoleucine I200, the variation of CcF3H corresponding amino acid to valine V200 revealed that V200 was more conserved in plant F3H protein. Spatial structure analysis showed that the substitution of amino acid V200 in CcF3H resulted in the enhancement of the molecular force between V200 and lysine K197. The results showed that CcF3H was a more conservative plant F3 H protein and valine V200 might be an important functional site. These studies also provide new information for Camellia F3 H gene and provide theoretical support for the selection of CcF3H gene as the most suitable candidate gene for Camellia flower color breeding in the future.
【作者單位】： 樂山師范學(xué)院生命科學(xué)學(xué)院;
【基金】：樂山師范學(xué)院科研項目(Z1201)資助
【分類號】：Q943.2

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