三角褐指藻DGAT1基因生物信息学分析及表达调控研究
PDF下载 (379)王何瑜,龚一富,郑小恽,李申睿.三角褐指藻DGAT1基因生物信息学分析及表达调控研究[J].宁波大学学报(理工版),2021,34(4):1-7.DOI:
WANG Heyu,GONG Yifu,ZHENG Xiaoyun,LI Shenrui.Bioinformation analysis and expression regulation of DGAT1 gene in Phaeodactylum tricornutum[J].Journal of Ningbo University(Natural Science & Engineering Edition),2021,34(4):1-7.DOI:
| Title: | Bioinformation analysis and expression regulation of DGAT1 gene in Phaeodactylum tricornutum |
| 作者: | 王何瑜, 龚一富, 郑小恽, 李申睿 |
| Author(s): | WANG Heyu, GONG Yifu, ZHENG Xiaoyun, LI Shenrui |
| 关键词: | 三角褐指藻; DGAT1基因; 克隆; 生物信息学; 基因表达调控 |
| Keywords: | Phaeodactylum tricornutum; DGAT1 gene; cloning; bioinformatics; gene expression regulation |
| 分类号: | S917.3; Q943.2 |
| 文献标识码: | A |
| 摘要: | 本研究采用转录组测序获得了三角褐指藻DGAT1基因cDNA全长序列(GeneBank登录号: 7200924), 并对其进行生物信息学分析和表达调控研究. 结果表明, 三角褐指藻DGAT1基因cDNA序列全长为1438bp, 开放阅读框(ORF)为1098bp, 编码365氨基酸序列, 含有脂肪酸蛋白特性(I)和DAG结合位点(II)等功能结构域. 预测三角褐指藻DGAT1蛋白为亲水性蛋白, 含有6个跨膜结构域和7个超强跨膜螺旋区, 无信号肽. 进化树分析表明, 三角褐指藻与海链藻DGAT1蛋白同源性最高. RT-qPCR结果表明, 光照强度和温度均显著促进三角褐指藻DGAT1基因的表达. 随着光照强度和温度的增加, 三角褐指藻DGAT1基因的表达量呈先升高后降低趋势, 在光照强度为2500lx或温度为25℃时, 三角褐指藻DGAT1基因的表达量达到最大, 这与三角褐指藻总脂含量的变化趋势一致, 同样揭示DGAT1蛋白与三角褐指藻总脂生物合成与积累密切相关. |
| Abstract: | In the current study, the full-length cDNA sequence of DGAT1 (GeneBank Accession Number: 7200924) in Phaeodactylum tricornutum was cloned by transcriptome sequencing. Bioinformatics analysis and expression difference were also studied. The results showed that the full length of the DGAT1 gene from P. tricornutum was 1438bp, had an open reading frame (ORF) of 1098bp with the fatty acid-binding protein signature (I) and DAG binding site (II) domains, encoding 365 amino acids. The protein prediction results showed that the DGAT1 protein from P. tricornutum was a hydrophilicity protein including 6 transmembrane structure domains and 7 super transmembrane helical regions, without signal peptide. Phylogenetic analysis results showed that DGAT1 protein from P. tricornutum had closed homologous relationship with the DGAT1 protein of Thalassiosira pseudonana. RT-qPCR results showed that light intensity and temperature could significantly promote the expression of DGAT1 gene of P. tricornutum. With the increase of light intensity and temperature, the expression quantity of DGAT1 gene from P. tricornutum showed an increased first and then decreased trend. The expression quantity of DGAT1 gene reached the maximum at light intensity of 2500 lx or temperature of 25℃. The expression of DGAT1 gene was consistent with the total fat content. The results indicated that DGAT1 protein might be one of the key enzymes in the biosynthesis and accumulation of total lipid in P. tricornutum. |
| 参考文献 /References: | [1] Gouveis L, Liveira A C. Microalgae as a raw material for biofuels production[J]. Journal of Industrial Microbiology & Biotechnology, 2009, 36(2):269-274. [2] Chisti Y. Biodiesel from microalgae[J]. Biotechnology Advances, 2007, 25(3):294-306. [3] 马海明, 施启顺, 柳小春. DGAT相关基因研究进展[J]. 遗传学报, 2005, 32(12):1327-1332. [4] 唐桂英, 柳展基, 单雷. 二酰基甘油酰基转移酶(DGAT)研究进展[J]. 中国油料作物学报, 2010, 32(2):320-328. [5] Shockey J M, Gidda S K, Chapital D C, et al. Tung tree DGAT1 and DGAT2 have nonredundant functions in triacylglycerol biosynthesis and are localized to different subdomains of the Endoplasmic reticulum[J]. Plant Cell, 2006, 18(9):2294-2313. [6] Jako C, Taylor D C. Seed-specific over-expression of an Arabidopsis cDNA encoding a diacylglycerol acyltrans-ferase enhances seed oil content and seed weight[J]. Plant Physiology, 2001, 126(2):861-874. [7] Lardizabal K, Effertz R, Levering C, et al. Expression of Umbelopsis ramanniana DGAT2A in seed increases oil in soybean[J]. Plant Physiology, 2008, 148(1):89-96. [8] Gómez-Loredo A, Benavides J, Rito-Palomares M. Growth kinetics and fucoxanthin production of Phaeodactylum tricornutum, and Isochrysis galbana, cultures at different light and agitation conditions[J]. Journal of Applied Phycology, 2016, 28(2):849-860. [9] Jaeyon L, Chan Y, Soyoung J, et al. Comparison of several methods for effective lipid extraction from microalgae[J]. Bioresource Technology, 2009, 101(1):75-77. [10] 李书霞, 刘伟, 李威, 等. 玉米DGAT基因家族的全基因组分析[J]. 核农学报, 2015, 29(4):643-650. [11] 刘贵芹, 邵群, 黄荣峰, 等. 大豆DGAT基因家族的鉴定和表达分析[J]. 中国农学通报, 2013, 29(12):55-61. [12] 李书霞. 玉米和棉花DGAT基因家族的全基因组分析[D]. 杭州: 浙江大学, 2015. [13] 刘凯. 油茶DGAT1基因的全长cDNA克隆与原核表达[D]. 长沙: 中南林业科技大学, 2012. [14] 吕文兵, 刘红涛, 薛乐勋. 不同盐浓度和光照强度对杜氏盐藻psbA基因表达的影响[J]. 华中师范大学学报(自然科学版), 2010, 44(2):288-292. [15] 尹航, 袁岚瑛, 俞凯, 等. 三角褐指藻GPAT基因生物信息学及表达差异分析[J]. 核农学报, 2019, 33(8):1483-1489. [16] 华雪铭, 周洪琪, 丁卓平. 温度和光照对微藻的生长、总脂肪含量及脂肪酸组成的影响[J]. 上海海洋大学学报, 1999, 8(4):309-315. [17] 叶丽, 蒋霞敏, 毛欣欣, 等. 温、光、盐对三角褐指藻紫外诱变株生长、总脂及脂肪酸的影响[J]. 生态学杂志, 2015, 34(2):454-462. [18] 蒋霞敏, 郑亦周. 14种微藻总脂含量和脂肪酸组成研究[J]. 水生生物学报, 2003, 27(3):243-247. [19] 吕军, 张颖, 冯立芹, 等. 生物信息学工具BLAST的使用简介[J]. 内蒙古大学学报(自然科学版), 2003, 34(2):179-191. [20] Gasteiger E, Hoogland C, Gattiker A, et al. Protein Identification and Analysis Tools on the ExPASy Server[M]//The Proteomics Protocols Handbook. Totowa, NJ: Humana Press, 2005:571-607. [21] 余舜武, 刘鸿艳, 罗利军. 利用不同实时定量PCR方法分析相对基因表达差异[J]. 作物学报, 2007, 33(7):1214-1218. [22] Bligh E G, Dyer W J. A rapid method of total lipid extraction and purification[J]. Canadian Journal of Biochemistry & Physiology, 1959, 37(8):911-917. [23] 马燕斌, 吴霞, 王霞, 等. 甘蓝型油菜BnDGAT1基因表达的特异性分析[J]. 西北植物学报, 2013, 33(10):1958-1963. [24] 周丹. 大豆二酰甘油酰基转移酶(DGAT)基因克隆及其功能的初步研究[D]. 南京: 南京农业大学, 2012. [25] Lung S C, Weselake R J. Diacylglycerol acyltransferase: A key mediator of plant triacylglycerol synthesis[J]. Lipids, 2006, 41(12):1073-1088. [26] Guo Z, Cromley D, Billheimer J T, et al. Identification of potential substrate-binding sites in yeast and human acyl-CoA sterol acyltransferases by mutagenesis of conserved sequences[J]. Journal of Lipid Research, 2001, 42(8):1282-1291. [27] Weselake R J, Madhavji M, Szarka S J, et al. Acyl-CoA-binding and self-associating properties of a recombinant 13.3 kDa N-terminal fragment of diacylglycerol acyl-transferase-1 from oilseed rape[J]. BMC Biochemistry, 2006, 7(1):24-36. [28] 郝敬云, 周广航, 邵雪梅, 等. 莱茵衣藻DGAT2基因家族的鉴定与功能分析[J]. 分子植物育种, 2016, 14(9):2343-2352. [29] 张晓琼, 乔琳, 胡利宗, 等. 棉花与模式植物DGAT2基因的鉴定与分析[J]. 江苏农业科学, 2016, 44(10):69-73. [30] 沈城, 刘楚吾, 刘丽. 温度胁迫及恢复初期稀杯盔形珊瑚共生虫黄藻Hsp70、Hsp90、psaA、psbA基因表达分析[J]. 热带海洋学报, 2016, 35(3):72-78. [31] 周洪琪, Renaud S M, Pany D L, 等. 温度对新月菱形藻、铲状菱形藻和杷夫藻的生长、总脂肪含量以及脂肪酸组成的影响[J]. 水产学报, 1996, 20(3):235-240. |
| 备注/Memo: | 收稿日期: 2019－11－20. 宁波大学学报（理工版）网址: http://journallg.nbu.edu.cn/ 基金项目: 浙江省科技厅重点科技创新团队项目（2012R10029-07, 2010R50029）; 宁波市科技攻关项目（2014C91023, 2013C10018）; 宁波市社发重大项目（2017C510002）. 第一作者: 王何瑜（1982－）, 女, 重庆江津人, 助理实验师, 主要研究方向: 藻类生物技术. E-mail: wangheyu@nbu.edu.cn *通信作者: 龚一富（1973－）, 男, 重庆开县人, 博士/副教授, 主要研究方向: 植物次生代谢产物及机理. E-mail: gongyifu@163.com 宁波大学学报（理工版）网址：http://journallg.nbu.edu.cn/ |
