茉莉花萜类合成酶基因的转录组鉴定及响应外源激素的表达研究

doi:10.13560/j.cnki.biotech.bull.1985.2021-0716

摘要/Abstract

摘要：

探究萜类合成酶（Terpenoid synthase,TPS）在茉莉花萜类化合物合成中的作用及外源激素处理下的表达模式,为深入研究茉莉花TPS基因家族的功能及在外源激素胁迫中的作用奠定理论依据。以茉莉花转录组数据为基础,利用生物信息学方法对茉莉花TPS家族进行鉴定及分析,采用实时荧光定量分析茉莉花TPS家族在茉莉花不同组织中及不同激素处理下的表达模式。结果显示,茉莉花TPS家族含有8个同时具有Terpene_synthase和Terpene_synthase_C结构域的家族成员,分别命名为JsTPS1-JsTPS8。茉莉花TPS成员的蛋白长度为163-844个氨基酸;分子量为19 103.6-96 887.4 kD,主要定位于细胞质和叶绿体中。系统进化树显示茉莉花TPS基因家族可以分为4个亚家族（a、b、e/f、g）;qRT-PCR结果显示,JsTPS1、JsTPS2、JsTPS3、JsTPS5、JsTPS7基因在花瓣中高表达;经IAA、GA、SA和MeJA处理后JsTPS1、JsTPS2、JsTPS3、JsTPS5、JsTPS6和JsTPS7的表达量受到不同程度的诱导表达。MeJA处理可显著诱导6个茉莉花TPS基因的上调表达,相对表达量最大值出现在4-9 h;4种激素的诱导作用由大到小分别为：MeJA>IAA>GA>SA。茉莉花含有8个TPS家族成员,茉莉花TPS基因主要在花瓣中高表达且与茉莉花香气形成关系密切,能够被IAA和MeJA诱导上调表达,GA和SA可能会抑制部分茉莉花TPS基因的表达。

关键词: 茉莉花, 萜类合成酶, 生物信息学, 表达分析

Abstract:

The objective of this work is to explore the role of terpenoid synthase（TPS）in the synthesis of terpenoid in jasmine and its expression under exogenous hormones treatments,which will lay a theoretical basis for further study of the function of J. sambac TPS（JsTPS）family and its role in exogenous hormones stress. Based on the transcriptome data of jasmine in this study,bioinformatics was applied to identify and analyze the JsTPS family. Real-time fluorescence quantitative detection was to analyze the expressions of JsTPS family in different tissues and exogenous hormones treatments. The result showed that there were 8 TPS genes of having both Terpene_synthase and Terpene_synthase_C conserved domain in JsTPS family,named JsTPS1 - JsTPS8 respectively. The protein lengths of JsTPS genes ranged from 163 to 844 amino acids. The molecular weights of JsTPS genes ranged from 19 103.6 to 96 887.4 kD. And they were mainly located in cytoplasm and chloroplast. The JsTPS genes were divided into 4 subfamilies（a,b,e/f,and g）based on phylogenetic relationships. The analysis of qRT-PCR showed that the expressions of gene JsTPS1,JsTPS2,JsTPS3,JsTPS5,JsTPS6,and JsTPS7 were high in petals;also the expressions of the JsTPS1,JsTPS2,JsTPS3,JsTPS5,JsTPS6 and JsTPS7 were induced by IAA,GA,SA and MeJA. The MeJA significantly induced the expressions of 6 JsTPS genes,and the relative expression reached the maximum at 4-9 h. The induction from high to low was MeJA > IAA > GA > SA. J. sambac contained 8 TPS family members which were mainly expressed in petals and closely related to the formation of aroma in J. sambac. They were up-regulated under IAA and MeJA inductions. GA and SA treatments may inhibit the expressions of partial JsTPS genes.

Key words: Jasminum sambac, terpenoid synthase, bioinformatics, expression analysis

洪雅萍, 陈雪津, 王鹏杰, 谷梦雅, 高婷, 叶乃兴. 茉莉花萜类合成酶基因的转录组鉴定及响应外源激素的表达研究[J]. 生物技术通报, 2022, 38(3): 41-49.

HONG Ya-ping, CHEN Xue-jin, WANG Peng-jie, GU Meng-ya, GAO Ting, YE Nai-xing. Transcriptome Identification of Terpenoid Synthase Genes in Jasminum sambac and Their Expressions Responding to Exogenous Hormones[J]. Biotechnology Bulletin, 2022, 38(3): 41-49.

图/表 7

表1 引物序列

Table 1 Primer sequences

基因名称 Gene name	上游引物 Forward primer（5'-3'）	下游引物 Reverse primer（5'-3'）
JsTPS1	TTGAAGTTGCTCGACGCTCT	GCATCTCCAAGGCTGCACTA
JsTPS2	AGCACGATGACCAGACAGTG	GCACCAGTCGAGCGTAATCT
JsTPS3	GATGATCGGAGGTGGCGATT	TGCAAAATGTGACGCTTCGG
JsTPS4	GGTGGTGGGAGAATCTTGGG	CAAGGAATGAAGCGCTTCCG
JsTPS5	GAGAACCTCCACCTCACTGC	AAGCCATTTTCGCAAGCTCG
JsTPS6	TACCACAGGAGCATGGCAAG	AATCGGCAACTCCCTGGAAG
JsTPS7	GCTCTACCCGCAACTCTTGT	TCGTCTGCAGGATACCTTGC
JsTPS8	GCTTGAGCTCTTTGAGGCCT	GTGGTTCAAATGCGACTCCA

表2 茉莉花JsTPS基因家族的序列特征

Table 2 Sequence characteristics of JsTPS genes in J. sambac

基因 Gene	转录组基因 Transcriptome gene	氨基酸数目 Number of amino acids/aa	分子量 MW/kD	等电点 pI	平均亲水性 GRAVY	亚细胞定位 Subcellular location	功能预测 Predicted functional
JsTPS1	c33849.graph_c0	552	64070.58	5.25	-0.191	叶绿体 Chloroplast	大根香叶烯D合酶 Germacrene D synthase
JsTPS2	c29553.graph_c0	593	69302.16	5.78	-0.371	细胞质 Cytoplasm	-
JsTPS3	c30319.graph_c0	600	69530.43	5.90	-0.351	叶绿体 Chloroplast	-
JsTPS4	c19904.graph_c0	584	66906.02	5.18	-0.326	叶绿体 Chloroplast	-
JsTPS5	c33314.graph_c0	540	62628.94	5.63	-0.245	细胞质 Cytoplasm	α-法尼烯合酶 α-farnesene synthase
JsTPS6	c32177.graph_c0	584	68311.93	5.55	-0.390	叶绿体 Chloroplast	-
JsTPS7	c27401.graph_c0	844	96887.40	6.09	-0.343	细胞质 Cytoplasm	（E,E）-香叶基芳樟醇合酶（E,E）-geranyllinalool synthase
JsTPS8	c44781.graph_c0	163	19103.06	5.64	-0.071	-	-

图1 茉莉花与其他植物TPS蛋白的系统进化分析 AtTPS：拟南芥;TPS：番茄;CsTPS：茶树;OeTPS：油橄榄;OfTPS：桂花;JsTPS：茉莉花

Fig. 1 Phylogenetic analysis of the TPS proteins in the J. sambac and other plants AtTPS：Arabidopsis thaliana. TPS：Solanum lycopersicum. CsTPS：Camellia sinensis. OeTPS：Olea europaea. OfTPS：Osmanthus fragrans. JsTPS：J. sambac

图2 茉莉花TPS中RRX8W、RXR、DDXXD及NSE/DTE保守序列的比对 A：纵坐标为氨基酸序列堆叠的高度,表示该位置处的序列保守性;横坐标为氨基酸序列排列的顺序,表示该位置的每个氨基酸的相对频率;B：利用DNAMAN7.0软件对茉莉花TPS中 RRX8W、RXR、DDXXD及NSE/DTE保守序列进行对齐

Fig. 2 Alignment of the RRX8W,RXR,DDXXD and NSE/DTE conserved sequence among J. sambac TPS A：The ordinate is the height of amino acid sequence stack,indicating the sequence conservation at this position. The abscissa is the sequence of amino acid sequences and represents the relative frequency of each amino acid at that position. B：Alignment of the RRX8W,RXR,DDXXD and NSE/DTE conserved sequence among JsTPS by using DNAMAN7.0

图3 茉莉花TPS家族保守基序分析 A：茉莉花TPS家族系统进化树;B：茉莉花TPS家族保守基序分布;C：基序logo分析

Fig. 3 Conservative motif analysis of TPS family in J. sambac A: Phylogenetic tree of TPS family in J. sambac. B: Conserved motif distribution of TPS family in J. samba. C: Motif logo analysis

图4 茉莉花JsTPS家族在不同组织的表达谱红色代表高表达,蓝色代表低表达

Fig. 4 Expression patterns of J. sambac JsTPS family in different tissues Red represents high expression,and bule represents low expression

图5 茉莉花JsTPS在不同激素处理下的表达模式不同小写字母代表同一基因在不同激素处理下存在显著差异（P<0.05）

Fig. 5 Expression analysis of JsTPS under different treat-ments of IAA,GA,SA and MeJA Different normal letters indicate significant differences among different treatments of IAA,GA,SA and MeJA at 0.05 level

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