【摘要】：番茄紅素β-環(huán)化酶(lycopene beta-cyclase,LCYB)能催化番茄紅素形成β-胡蘿卜素及其氧化物,是處于植物類胡蘿卜素合成途徑中的一個(gè)分支點(diǎn)。為研究三角褐指藻(Phaeodactylum tricornutum)在不同外源誘導(dǎo)下lcyb基因的表達(dá)情況及其對巖藻黃素積累的影響,本研究通過從頭測序(de novo測序)獲得三角褐指藻lcyb基因c DNA全長并對其進(jìn)行生物信息學(xué)分析,進(jìn)一步對經(jīng)過不同濃度的茉莉酸甲酯(methyl jasmonate,Me JA)、乙酰水楊酸(acetylsalicylic acid,ASA)、花生四烯酸(arachidonic acid,AA)和硫酸鈰銨(ammonium cerous sulfate,ACS)處理的三角褐指藻lcyb基因表達(dá)進(jìn)行分析,并對其巖藻黃素含量進(jìn)行測定。結(jié)果表明,本研究通過轉(zhuǎn)錄組獲得的三角褐指藻lcyb基因序列與NCBI上已知序列(Gen Bank No.XM002176576)相同,其c DNA全長為2 060 bp,含1個(gè)長度為1 980 bp的開放閱讀框(open reading fragment,ORF),編碼659個(gè)氨基酸,該蛋白在N端含有1個(gè)雙核苷酸結(jié)合位點(diǎn),在C端含有1個(gè)螺旋跨膜區(qū)域,存在葉綠體導(dǎo)肽和信號肽,定位于葉綠體類囊膜上。系統(tǒng)發(fā)育進(jìn)化樹顯示,三角褐指藻等硅藻共處1個(gè)進(jìn)化支,親緣關(guān)系較近。誘導(dǎo)轉(zhuǎn)錄結(jié)果表明,50μmol/L Me JA、10 mg/L ASA、0.1 mg/L AA和0.4 mg/L ACS處理后的三角褐指藻lcyb基因在本實(shí)驗(yàn)范圍內(nèi)轉(zhuǎn)錄表達(dá)水平最高,且三角褐指藻中的巖藻黃素含量隨著lcyb基因轉(zhuǎn)錄量的增加而增加,并呈正相關(guān),說明該酶在巖藻黃素合成過程中起到重要作用,誘導(dǎo)子能增強(qiáng)該基因轉(zhuǎn)錄從而達(dá)到提高巖藻黃素含量的目的。本研究為巖藻黃素的合成調(diào)控提供理論指導(dǎo),也為進(jìn)一步通過代謝工程手段提高三角褐指藻中的巖藻黃素含量提供參考。
[Abstract]:Lycopene 尾-cyclase (lycopene beta-cyclase,LCYB) can catalyze lycopene to form 尾-carotene and its oxides, which is a branch of plant carotenoid synthesis pathway. In order to study the expression of lcyb gene in Phaeodactylus triangularis (Phaeodactylum tricornutum) induced by different exogenous sources and its effect on the accumulation of alginin, In this study, the full length of lcyb gene c DNA was obtained by ab initio sequencing (de novo sequencing, and its bioinformatics analysis was carried out. Methyl jasmonate,Me JA), acetylsalicylic acid (acetylsalicylic acid, with different concentrations was further studied. The lcyb gene expression of ASA), arachidonic acid (arachidonic acid,AA) and ammonium cerium sulfate (ammonium cerous sulfate,ACS) was analyzed and the content of alginin was determined. The results showed that the lcyb gene sequence of Phaeodactylus triangularis obtained from the transcriptional group was the same as that of the known sequence (Gen Bank No.XM002176576 on NCBI. The c DNA was 2060 bp, with an open reading frame (open reading fragment,ORF of 1.980 bp in length. It encodes 659 amino acids. The protein contains a dinucleotides binding site at the N-terminal and a spiral transmembrane region at the C-terminal. Chloroplast conduction peptides and signal peptides are located on the chloroplast capsule. The phylogenetic tree showed that diatoms such as Phaeodactylus triangularis coexisted with one evolutionary branch and were closely related to each other. The results of induced transcription showed that the lcyb gene of Phaeodactylus triangularis treated with 50 渭 mol / L Me JA,10 mg/L ASA,0.1 mg/L AA and 0.4 mg/L ACS had the highest transcriptional expression level in this experiment. The content of alginin in Phaeodactylus triangularis increased with the increase of lcyb gene transcription, and there was a positive correlation, which indicated that the enzyme played an important role in the synthesis of alginin. The inducer can enhance the transcription of the gene and increase the content of alginin. This study provides theoretical guidance for the synthesis and regulation of alginin, and also provides a reference for further improving the content of alginin in Phaeodactylus triangularis by means of metabolic engineering.
【作者單位】： 寧波大學(xué)海洋學(xué)院;
【基金】：國家星火計(jì)劃項(xiàng)目(No.2015GA701001) 浙江省科技廳重點(diǎn)科技創(chuàng)新團(tuán)隊(duì)項(xiàng)目(No.2012R10029-07和No.2010R50029) 寧波科技攻關(guān)項(xiàng)目(No.2013C10018和No.2014C91023)
【分類號】：Q943.2

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