紫苏PfPDCT基因序列特征及表达分析
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摘要
采用生物信息学技术和在线分析软件对紫苏PfPDCT基因序列及所编码的蛋白质进行分析,并且利用实时荧光定量PCR技术研究该基因在晋紫苏1号种子发育不同时期的表达特性,旨在探究磷脂酰胆碱甘油二脂转磷酸胆碱酶(PDCT)在植物脂肪酸代谢过程的作用。结果表明,紫苏PfPDCT基因c DNA全长序列为2 098 bp,开放阅读框为1 683 bp,编码560个氨基酸残基。生物信息学分析结果表明,PfPDCT蛋白分子量约为59.315 ku,等电点为9.48,不稳定系数为35.00,推测其为稳定蛋白,与已知赤藓PDCT蛋白高度同源。通过荧光定量PCR分析可知,紫苏PfPDCT基因在不同发育时期的种子中均有表达,其中,在开花后20 d表达量最高,为开花后10 d的1.92倍。研究结果可为进一步阐明紫苏PfPDCT基因的功能及作用机制奠定基础。
This study was conducted to explore the role of PfPDCT enzyme in the process of the plant fatty acids metabolism. The PfPDCT gene and the encoded protein were analyzed using bioinformatics techniques and online software. And the PfPDCT gene expression characteristics in Perilla frutescens seeds from different development stages of Jinzisu 1 were analyzed. The results showed that the full-length c DNA sequence of PfPDCT gene was 2 098 bp, with a 1 683 bp ORF, which encoded 560 amino acid residues. Bioinformatics analysis results showed that the predicted molecular quality of PfPDCT protein was about 59.315 ku, with a theoretical isoelectric point of9.48, and the instability coefficient of this protein was 35.00. Phylogeny analysis revealed that PfPDCT was closest to that from Erythranthe guttatus. The expression pattern of PfPDCT gene at different development stages of seeds was analyzed by semi-quantitative PCR. The results showed that the expression level of PfPDCT gene was the highest at 20 d after flowering, which was 1.92 times as much as that of 10 d after flowering. This study will provide a basis for further clarifying the function and mechanism of PfPDCTT gene.
This study was conducted to explore the role of PfPDCT enzyme in the process of the plant fatty acids metabolism. The PfPDCT gene and the encoded protein were analyzed using bioinformatics techniques and online software. And the PfPDCT gene expression characteristics in Perilla frutescens seeds from different development stages of Jinzisu 1 were analyzed. The results showed that the full-length c DNA sequence of PfPDCT gene was 2 098 bp, with a 1 683 bp ORF, which encoded 560 amino acid residues. Bioinformatics analysis results showed that the predicted molecular quality of PfPDCT protein was about 59.315 ku, with a theoretical isoelectric point of9.48, and the instability coefficient of this protein was 35.00. Phylogeny analysis revealed that PfPDCT was closest to that from Erythranthe guttatus. The expression pattern of PfPDCT gene at different development stages of seeds was analyzed by semi-quantitative PCR. The results showed that the expression level of PfPDCT gene was the highest at 20 d after flowering, which was 1.92 times as much as that of 10 d after flowering. This study will provide a basis for further clarifying the function and mechanism of PfPDCTT gene.
引文
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[19]谭太龙,冯韬,罗海燕,等.甘蓝型油菜磷脂二酰甘油酰基转移酶(BnPDAT1)cDNA的克隆和功能鉴定[J].作物学报,2016,42(5):658-666.
[2] YAN H U,SUNL W,ZHANG Y X,et al. MOKGOLODI N C. Prima ry identifications and palynological observations of Perilla in China[J]. Journal of Systematics and Evolution,2010,48(2):133-145.
[3]谭美莲,严明芳,汪磊,等.国内外紫苏研究进展概述[J].中国油料作物学报,2012,34(2):225-231.
[4]李璐,梁倩,安茜,等.紫苏脂肪酸硫酯酶基因PfFatA生物信息学及其表达特性分析[J].华北农学报, 2017,32(2):104-108.
[5]LEE K R,LEE Y,KIM E H,et al. Functional identification of oleate12-desaturase andω-3 fatty acid desaturase genes from Perilla frutescens var. frutescens[J]. Plant Cell Reports,2016,35(12):1-15.
[6]梁倩,李璐,安茜,等.紫苏脂肪酸去饱和酶基因PfFAD2的生物信息学及表达特性分析[J].山西农业科学,2018,46(3):316-319.
[7]禹山林.花生脂肪酸代谢关键酶基因的克隆与表达分析[D].南京:南京农业大学,2008.
[8] BROUN P,GETTNER S,SOMERVILLE C. Genetic engineering of plant lipids[J]. Annual Review of Nutrition,1999,19(1):197-216.
[9] DAWN JO'BYRNE,KNAUFT D A,SHIREMAN R B,et al. Low fat-monounsaturated rich diets containing high-oleic peanuts improve serum lipoprotein profiles[J]. Lipids,1997,32(7):687-695.
[10]王树彦,韩冰,周四敏,等.胡麻脂肪酸含量与相关基因差异表达[J].中国油料作报,2016,38(6):771-777.
[11] LU C,XIN Z,REN Z,et al. An enzyme regulating triacyl-glycerol composition is encoded by the ROD1 gene of Ara-bidopsis[J]. Proc Nat Acad Sci,2009,106(44):18837-18842.
[12] WICKRAMARATHNA A D,SILOTO R M P,MIETKIEWSKA E,et al. Heterologous expression of flax PHOSPHOLIPID:DIACYLGLYCEROL CHOLINEPHOSPHOTRANSFERASE(PDCT)increases polyunsaturated fatty acid content in yeast and Arabidopsis seeds[J]. Bmc Biotechnology,2015,15(1):63.
[13] GUAN R,LI X,HOFVANDER P,et al. RNAi Targeting putative genes in phosphatidylcholine turnover results in significant change in fatty acid composition in crambe abyssinica seed oil[J]. Lipids,2015,50(4):407-416.
[14]张玲慧.紫苏种子油脂代谢及限速酶DGAT1基因的表达分析[D].太谷:山西农业大学,2014.
[15] BATES P D,DURRETT T P,OHLROGGE J B,et al. Analysis of acyl fluxes through multiple pathways of triacylglycerol synthesis in developing soybean embryos[J]. Plant Physiology,2009,150(1):55-72.
[16] BATES P D,OHLROGGE J B,POLLARD M. Incorporation of newly synthesized fatty acids into cytosolic glycerolipids in pea leaves occurs via acyl editing[J]. Journal of Biological Chemistry,2007,282(43):31206-31216.
[17] WU Y,LI R,HILDEBRAND D F. Biosynthesis and metabolic engineering of palmitoleate production,an important contributor to human health and sustainable industry[J]. Progress in Lipid Research,2012,51(4):340-349.
[18] HU Z,REN Z,LU C. The phosphatidylcholine diacylglycerol cholinephosphotransferase is required for efficient hydroxy fatty acid accumulation in transgenic arabidopsis[J]. Plant Physiology,2012,158(4):1944-1954.
[19]谭太龙,冯韬,罗海燕,等.甘蓝型油菜磷脂二酰甘油酰基转移酶(BnPDAT1)cDNA的克隆和功能鉴定[J].作物学报,2016,42(5):658-666.