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作物学报 ›› 2022, Vol. 48 ›› Issue (12): 3029-3044.doi: 10.3724/SP.J.1006.2022.14237

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

割手密萜烯合成酶(TPS)基因家族分析及其在生物胁迫下的表达分析

林焕泰(), 张天杰, 史梦婷, 郭燕芳, 高三基, 王锦达()   

  1. 福建农林大学国家甘蔗工程技术研究中心, 福建福州 350002
  • 收稿日期:2021-12-15 接受日期:2022-03-25 出版日期:2022-12-12 网络出版日期:2022-04-19
  • 通讯作者: 王锦达
  • 作者简介:E-mail: 471250594@qq.com
  • 基金资助:
    国家重点研发计划项目(2018YFD201100);财政部和农业农村部国家现代农业产业技术体系建设专项(糖料, CARS-170302);福建省大学生创新训练计划项目(S202110389051);福建农林大学科技创新专项(CXZX2020084A)

Genome-wide analysis of terpene synthase (TPS) gene family and its expression under biological stress in Saccharum spontaneum

LIN Huan-Tai(), ZHANG Tian-Jie, SHI Meng-Ting, GUO Yan-Fang, GAO San-Ji, WANG Jin-Da()   

  1. National Engineering Research Center for Sugarcane, Fujian Agriculture and Forestry University, Fuzhou 350002, Fujian, China
  • Received:2021-12-15 Accepted:2022-03-25 Published:2022-12-12 Published online:2022-04-19
  • Contact: WANG Jin-Da
  • Supported by:
    National Key Research and Development Program of China(2018YFD201100);China Agricultural Systrm of MOF and MARA(Sugar, CARS-170302);Innovation Training Program for College Students of Fujian Province(S202110389051);Special Fund for Science and Technology Innovation of Fujian Agriculture and Forestry University(CXZX2020084A)

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摘要：

由萜烯合酶(TPS)合成的萜类化合物在植物生物和非生物胁迫中起重要作用。作为现代甘蔗栽培种的重要亲本材料, 割手密(Saccharum spontaneum)含有大量的抗逆基因。为调查割手密TPS基因家族的特征和功能, 通过使用HMMER搜索在割手密基因组中鉴定出39个TPS基因, 该基因含有2个保守域(PF01397和PF03936)的蛋白质, 系统进化树分析显示SsTPS蛋白可以分为TPS-a、b、e/f和g四个分支。SsTPS基因家族主要通过片段复制进行扩展, 共有12个SsTPS基因参与了片段复制事件。定量实时PCR显示, 在草地贪夜蛾(Spodoptera frugiperda)胁迫和甘蔗白条黄单胞菌(Xanthomonas albilineans)感染的割手密植株中, SsTPS基因的表达模式不同, 其中7个SsTPS基因的表达受到强烈的调节。值得注意的是, SsTPS15在草地贪夜蛾的胁迫中显著上调, 但被甘蔗白条黄单胞菌感染时表达下调, 而SsTPS26、SsTPS37和SsTPS39显示相反的结果。研究结果对于进一步了解萜烯合酶的生物学作用和基于割手密的甘蔗抗逆育种具有重要意义。

关键词: 割手密, 萜烯合酶基因, 生物胁迫, 表达分析, 抗性育种

Abstract:

Terpenoids produced by the action of terpene synthase (TPS) enzymes play important roles in plant biotic and abiotic stress. Saccharum spontaneum, an important parent material of modern sugarcane cultivars, contains a large number of stress resistance genes. To investigate the characteristics and functions of the TPS gene family in S. spontaneum, 39 SsTPS genes were identified in S. spontaneum genome that encoded proteins with two conserved domains (PF01397 and PF03936) by using an HMMER search. The SsTPS proteins were divided into TPS-a, b, e/f, and g subfamilies. The SsTPS gene family had mainly expanded through segmental duplications, and a total of 12 SsTPS genes involved in segmental duplication events. In addition, qRT-PCR showed that the expression patterns of some SsTPS genes differed in S. spontaneum between Spodoptera frugiperda-stressed and Xanthomonas albilineans-infected plants, whereas the relative expression levels of seven SsTPS genes were strongly up-regulated. Notably, SsTPS15 were up-regulated in response to Spodoptera frugiperda-stressed but were down-regulated by X. albilineans infection, while SsTPS26, SsTPS37, and SsTPS39 had the opposite results. These results will be of great significance to further understanding the biological roles of terpene synthases and to develop resistant breeding in S. spontaneum.

Key words: Saccharum spontaneum, terpene synthase gene, biotic stresses, expression analysis, resistance breeding

表1

用于检测SsTPS基因表达水平的引物序列"

基因名称
Gene name		 正向引物
Forward primer (5'-3')		 反向引物
Reverse primer (5'-3')
SsTPS5 AGTACCGCATGCCCTACCTA CACCGCTTCTTGCTGAAACC
SsTPS6 AGTACCGCATGCCCTACCTA CGCTGCAAGTGTTCCTCAAC
SsTPS8 ATGATGTTCAGGGGCTGCAA CATGTATGCACGCTTGGGTG
SsTPS10 AGGGGAAAGTGCGTCAGATG AGTGTGGATACGGCTTAGCG
SsTPS15 AAAAGACCGGACCTTCCACC CATGTATGCACGCTTGGGTG
SsTPS18 ACCAAGGAGGCATTCGAGTG CGCGTTCTTCCTCCCTTTCT
SsTPS26 GTCCCATGCACTCCATCACA GAGGTGGCAACGGTTCCTAA
SsTPS27 AATGGGCTCTCACCTTTCCG AAGCTTGTTCACGCTCATGC
SsTPS35 TGTTCCTAGGCCATGCAAGG TTTTGGACTCCGAGTGGCTC
SsTPS36 TGCTGCTGCGCAATGATTTT GGTCGCTCTTCTGACGTTGA
SsTPS37 GAGCCAAGCCTTGCAGAGTA ATGGGCAGGCTCTGTAGGTA
SsTPS39 GAAATACCTGTGGTGCGGGA CCACAACATAGACAAGCGCG
GAPDH CACGGCCACTGGAAGCA TCCTCAGGGTTCCTGATGCC

表2

割手密SsTPS家族成员基本信息和理化性质"

基因名称
Gene name		 基因编号
Gene ID		 氨基酸数
Number of amino acids		 等电点
pI		 分子量
Molecular weight (kD)		 负电荷残基
Asp+Glu		 正电荷残基Arg+Lys 不稳定系数Coefficient of instability 平均疏水性 GRAVY 脂肪系数 AI α-螺旋
α-helix(%)		 延伸链
Extended chain (%)		 无规则卷曲
Random curl (%)
SsTPS1 Sspon.01G0009730-1A 514 5.90 59,392.66 72 62 36.28 -0.306 88.29 57.39 11.67 30.93
SsTPS2 Sspon.01G0010010-1A 496 5.29 57,214.41 78 57 37.65 -0.288 94.92 54.03 14.72 31.25
SsTPS3 Sspon.01G0044880-1B 497 5.75 56,761.45 69 54 40.04 -0.312 83.84 49.90 17.51 32.60
SsTPS4 Sspon.04G0011840-1A 735 9.77 81,962.81 74 104 60.66 -0.460 75.17 43.67 8.03 48.30
SsTPS5 Sspon.04G0011840-2B 353 5.78 41,253.51 51 43 47.03 -0.333 87.00 64.31 3.68 32.01
SsTPS6 Sspon.04G0011840-3C 501 6.43 57,711.12 62 58 45.18 -0.311 87.96 55.09 6.99 37.92
SsTPS7 Sspon.05G0015000-1A 581 5.53 66,604.19 78 61 39.07 -0.216 91.02 51.29 12.22 36.49
SsTPS8 Sspon.06G0010490-1P 528 5.33 61,308.56 76 59 39.80 -0.173 98.26 52.65 12.12 35.23
SsTPS9 Sspon.06G0010500-1A 376 5.10 43,465.54 59 41 52.71 -0.328 87.95 52.39 11.17 36.44
SsTPS10 Sspon.06G0011520-1A 433 5.84 49,561.84 62 51 40.69 -0.242 83.44 55.43 9.70 34.87
SsTPS11 Sspon.06G0011680-1A 468 5.32 53,911.93 71 58 49.94 -0.247 97.67 60.47 7.69 31.84
SsTPS12 Sspon.06G0010490-1A 538 5.34 62,470.81 78 61 40.28 -0.201 96.43 50.74 12.64 36.62
SsTPS13 Sspon.06G0021690-1B 325 5.52 37,701.75 51 39 46.74 -0.380 89.14 55.38 12.62 32.00
SsTPS14 Sspon.06G0023410-1B 397 5.47 46,236.32 64 52 38.72 -0.254 96.93 50.13 12.85 37.03
SsTPS15 Sspon.06G0010490-2B 538 5.34 62,468.83 78 61 40.74 -0.192 97.16 50.93 12.64 36.43
SsTPS16 Sspon.06G0011680-2B 607 5.96 70,631.16 81 70 49.39 -0.222 94.23 50.74 13.67 35.58
SsTPS17 Sspon.06G0024130-1B 606 5.97 70,317.44 86 74 53.41 -0.363 83.83 55.61 13.53 30.86
SsTPS18 Sspon.06G0015570-2B 533 5.29 61,439.38 76 57 45.23 -0.220 83.49 50.66 14.82 34.52
SsTPS19 Sspon.06G0023520-2C 491 5.08 57,333.63 76 54 49.64 -0.302 84.83 46.84 13.65 39.51
SsTPS20 Sspon.06G0011660-2C 482 5.04 55,885.74 75 53 48.91 -0.274 86.72 47.51 18.67 33.82
SsTPS21 Sspon.06G0011680-1P 414 5.47 47,542.37 62 48 48.39 -0.301 93.16 53.86 10.63 35.51
SsTPS22 Sspon.06G0030650-1C 358 9.40 41,189.25 38 51 38.63 -0.391 79.55 46.37 13.69 39.94
SsTPS23 Sspon.06G0011680-3C 468 5.15 53,761.70 72 55 51.78 -0.219 98.93 58.55 9.83 31.62
SsTPS24 Sspon.06G0034360-1D 317 4.93 36,898.43 50 32 39.19 -0.134 91.29 48.26 17.98 33.75
SsTPS25 Sspon.06G0011680-2P 333 8.65 38,083.58 38 42 35.22 -0.328 83.75 38.44 21.32 40.24
SsTPS26 Sspon.06G0024130-2D 545 5.56 63,780.15 81 68 48.69 -0.286 90.11 53.39 13.58 33.03
SsTPS27 Sspon.06G0024130-1P 497 5.71 57,610.39 75 65 43.03 -0.181 93.00 51.71 17.30 30.99
SsTPS28 Sspon.06G0011680-4D 328 8.47 37,598.00 37 40 33.44 -0.324 82.65 42.38 17.38 40.24
SsTPS29 Sspon.07G0027750-1B 509 5.55 59,706.37 75 59 55.08 -0.414 90.84 52.65 9.23 38.11
SsTPS30 Sspon.08G0029890-1D 629 5.83 71,361.82 80 67 49.40 -0.160 86.38 46.26 13.67 40.06
SsTPS31 Sspon.01G0039100-1B 348 5.62 39,581.24 49 39 48.50 -0.288 84.74 47.14 12.93 39.66
SsTPS32 Sspon.01G0039100-2C 550 8.25 62,827.75 67 70 54.38 -0.376 82.91 50.36 9.45 40.18
SsTPS33 Sspon.04G0000830-1A 388 5.42 44,015.32 47 38 54.09 -0.088 93.84 55.41 7.47 37.11
SsTPS34 Sspon.05G0030980-1C 545 6.04 62,692.00 73 67 49.04 -0.309 84.02 41.47 12.29 46.24
SsTPS35 Sspon.05G0002480-3D 783 6.03 87,936.45 90 18 46.73 -0.168 89.13 47.51 11.11 41.38
SsTPS36 Sspon.04G0020000-1A 550 5.33 62,580.79 74 53 52.94 -0.116 92.62 49.27 16.55 34.18
SsTPS37 Sspon.04G0019960-2C 515 5.28 59,084.06 71 52 47.41 -0.090 96.37 53.59 14.17 32.23
SsTPS38 Sspon.04G0020000-4P 563 6.13 63,167.56 68 58 50.45 -0.145 89.68 52.75 11.90 35.35
SsTPS39 Sspon.04G0020000-3D 552 5.25 63,021.29 78 56 51.02 -0.123 95.27 54.53 11.78 33.70

图1

割手密(Ss)、高粱(Sb)、水稻(Os)中萜烯合酶基因家族系统进化树 TPS-a、b、e/f、g分别表示萜烯合酶基因家族不同分支。"

图2

割手密基因家族成员基因结构 A: SsTPS系统进化树。B: SsTPS内含子-外显子结构预测。C: SsTPS motif预测。D: Motif基序结构。"

图3

甘蔗萜烯合酶基因家族的染色体定位"

图4

甘蔗萜烯合酶基因家族的共线性分析"

图5

割手密TPS基因家族各成员启动子区顺式作用元件种类和数量 1~39分别表示割手密萜烯合酶基因家族SsTPS1~SsTPS39成员。"

图6

割手密萜烯合酶基因在不同组织中的表达热图"

图7

割手密中12个候选SsTPS基因在草地贪夜蛾胁迫下的相对表达水平 *、**、***分别表示在0.05、0.01和0.001水平差异显著。"

图8

割手密中12个SsTPS基因在X. albilinans感染下的表达分析 柱上不同小写字母表示处理间在0.05水平差异显著。"

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