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作物杂志,2025, 第2期: 29–39 doi: 10.16035/j.issn.1001-7283.2025.02.005

• 遗传育种·种质资源·生物技术 • 上一篇    下一篇

甘蓝型油菜LPAT家族基因鉴定与表达模式分析

李坤杰1,2(), 刘念1,2(), 丁磊1,2, 朱燕1,2, 蒙大庆1,2, 范其新1,2, 李迎春1,2, 陈军1,2   

  1. 1绵阳市农业科学研究院，621023，四川绵阳
    2厅市共建作物特色资源创新及四川省重点实验室，621023，四川绵阳
  • 收稿日期:2024-01-03 修回日期:2024-03-19 出版日期:2025-04-15 发布日期:2025-04-16
  • 通讯作者: 刘念，主要从事油菜育种研究，E-mail：937832892@qq.com
  • 作者简介:李坤杰，主要从事油菜分子生物学基础研究，E-mail：lkjcrop@foxmail.com
  • 基金资助:
    四川省重大科技专项(2022ZDZX0015);四川省创新团队项目(sccxtd-2023-03);国家油菜产业技术体系建设项目(CARS-12);重庆市技术创新与应用发展专项(cstc2021jscx-cylhX0003);四川省“十四五”育种攻关项目(2021YFYZ0018);四川省“天府菜油”行动奖补项目(2023TFRO06)

Identification and Expression Pattern Analysis of LPAT Family Genes in Brassica napus

Li Kunjie1,2(), Liu Nian1,2(), Ding Lei1,2, Zhu Yan1,2, Meng Daqing1,2, Fan Qixin1,2, Li Yingchun1,2, Chen Jun1,2   

  1. 1Mianyang Academy of Agricultural Sciences, Mianyang 621023, Sichuan, China
    2Crop Characteristic Resources Creation and Utilization Key Laboratory of Sichuan Province, Mianyang 621023, Sichuan, China
  • Received:2024-01-03 Revised:2024-03-19 Online:2025-04-15 Published:2025-04-16

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摘要：

溶血磷脂酰胆碱酰基转移酶（lysophosphatidylcholine acyltransferase，LPAT）在植物脂肪酸的积累和代谢中起着至关重要的作用，为了研究LPAT基因家族在甘蓝型油菜（Brassica napus）中的进化和表达模式，利用生物信息学方法，在甘蓝型油菜全基因组水平进行BnLPAT基因的鉴定，并对其理化性质、系统进化、基因结构、顺式作用元件和表达模式进行分析。结果表明，在甘蓝型油菜全基因组水平一共鉴定到44个LPAT基因，它们不均等分布在17条染色体上。根据系统发育关系分为LPAT1~LPAT5，共5个分支，相邻分支的成员具有相似的基因结构和保守基序构成。在推定的BnLPAT基因启动子区域一共鉴定到4类共42种顺式作用元件。此外，表达模式分析发现它们在不同组织部位均能检测到，进一步检测了12个BnLPAT基因在种子发育过程中以及4个基因在高、中、低含油量材料中的表达特征。

关键词: 溶血磷脂酰胆碱酰基转移酶（LPAT）, 甘蓝型油菜, 基因家族鉴定, 表达模式

Abstract:

Lysophosphatidylcholine acyltransferase (LPAT) plays a crucial role in the accumulation and metabolism of plant fatty acids. To investigate the evolution and expression pattern of the LPAT gene family in Brassica napus, we identified BnLPAT genes in the B.napus genome using bioinformatics methods. The physical and chemical properties, phylogeny, gene structure, cis-elements, and expression patterns of BnLPAT genes were analyzed. The results identified 44 LPAT genes in the B.napus genome, which were unevenly distributed on 17 chromosomes. Based on phylogenetic relationships, they were classified into five branches, LPAT1 to LPAT5, respectively. Members of adjacent branches had shared similar gene structures and conserved motifs. The 42 types of cis-elements were identified in the putative BnLPAT gene promoter regions, including four categories. Furthermore, expression pattern analysis revealed that BnLPAT genes were detected in different tissue parts. We further analyzed the expression characteristics of 12 BnLPAT genes during seed development and four genes in varieties with high, medium, and low oil contents.

Key words: Lysophosphatidylcholine acyltransferase (LPAT), Brassica napus, Identification of gene family, Expression pattern

表1

试验所用引物

基因名称Gene name 上游引物（5′-3′）Forward primer (5′-3′) 下游引物（5′-3′）Reverse primer (5′-3′)
BnLPAT1-A AGCGTTGGATGCATCTGGTTC GGTTATTGTCAGTTCCTTGA
BnLPAT1-C GCACAATATTAGAGAGTGTT GGTTATTGTCAGTTCCTTGA
BnLPAT4-A ATGGCAGCAGCAGTGATTGT GGTCGAACGAGGACATAGCA
BnLPAT4-C ATGGCAGCAGCAGCAGTGAT CAGAGGTCGAATGAGGACAT
BnLPAT12-A TGAGGCTGATCTTCCAGCATT AGCAGAACACTCTGCATCGCT
BnLPAT14-A GTATCATATATCAGTGGGCA ACTGTGCCAGATACACCCTT
BnLPAT14-C TGTATCATATGTCTGCTTCT ACTGTGCCAGATACACCCTT
BnLPAT19-A AGTATTTGCGGATAATGAGAC AACCTGACCTCTGAGCGAGA
BnLPAT19-C AAGTGTTTGCTGATAATGAG ACCTGATCTCTGAGCCAGAAT
BnLPAT20-A TCACATCCTCCATATTATAG TCAAGTTCGAAGAACGAGCA
BnLPAT20-C CATCCTCCTCCATATTATAGC AGAACGACCAAAGCAACATC
BnLAPT22-C TGTATCATATGTCTGCTTCT CCTCTCGTTGGACATAGACA
BnActin7 TGGGTTTGCTGGTGACGAT TGCCTAGGACGACCAACAATACT

表2

甘蓝型油菜BnLPAT蛋白理化性质和亚细胞定位

基因编号
Gene ID		 基因名称
Gene name		 氨基酸数目
Number of
amino acids		 分子质量
Molecular
weight (kDa)		 理论
等电点
pI		 不稳定指数
Instability
index		 脂肪系数
Aliphatic
coefficient		 水溶性平均值
Average water
solubility		 亚细胞定位
Subcellular
localization
BnA01g0003370.1 BnLPAT1-A 361 41.53 8.93 46.58 87.20 -0.170 叶绿体，内质网
BnA01g0033510.1 BnLPAT2-A 355 41.02 8.77 39.55 94.96 -0.019 内质网
BnA03g0114510.1 BnLPAT3-A 527 59.81 6.15 38.63 90.61 -0.050 线粒体
BnA03g0128320.1 BnLPAT4-A 390 43.77 9.24 50.09 97.26 0.067 内质网
BnA03g0133720.1 BnLPAT5-A 432 48.25 9.63 45.85 93.87 -0.224 内质网
BnA03g0144790.1 BnLPAT6-A 286 32.09 9.13 52.32 96.12 -0.041 叶绿体，内质网
BnA04g0162010.1 BnLPAT7-A 384 43.19 9.18 41.28 101.59 0.136 内质网
BnA05g0207140.1 BnLPAT8-A 440 48.80 6.24 48.98 78.91 -0.338 叶绿体
BnA05g0211550.1 BnLPAT9-A 376 43.89 9.05 37.63 93.56 0.043 内质网
BnA05g0213610.1 BnLPAT10-A 531 60.03 8.06 42.27 92.34 -0.076 线粒体
BnA07g0286470.1 BnLPAT11-A 399 44.75 6.13 53.03 87.42 -0.182 叶绿体，内质网
BnA07g0288480.1 BnLPAT12-A 380 43.26 8.25 44.55 98.76 0.144 内质网
BnA07g0302330.1 BnLPAT13-A 287 31.98 9.30 47.91 94.11 -0.036 叶绿体，内质网
BnA07g0303170.1 BnLPAT14-A 370 41.87 5.93 48.34 91.38 -0.176 内质网
BnA08g0305940.1 BnLPAT15-A 345 39.42 9.14 34.60 100.06 -0.014 内质网
BnA08g0312680.1 BnLPAT16-A 454 50.01 6.25 43.76 78.00 -0.331 叶绿体
BnA08g0327830.1 BnLPAT17-A 814 92.41 8.00 49.32 96.38 -0.008 －
BnA09g0362990.1 BnLPAT18-A 451 49.63 6.32 44.65 83.28 -0.202 叶绿体
BnA09g0374960.1 BnLPAT19-A 390 43.76 9.38 47.42 99.51 0.105 内质网
BnA09g0384080.1 BnLPAT20-A 358 39.41 9.67 43.09 87.37 0.037 叶绿体
BnA10g0406100.1 BnLPAT21-A 326 37.21 8.70 47.19 87.30 -0.132 叶绿体，内质网
BnC01g0428330.1 BnLPAT1-C 376 43.22 9.05 47.12 88.88 -0.099 内质网
BnC01g0463380.1 BnLPAT2-C 339 39.24 8.67 42.96 92.27 0.085 内质网
BnC02g0500200.1 BnLPAT13-C 285 31.86 9.24 49.19 96.46 -0.049 叶绿体，内质网
BnC03g0553040.1 BnLPAT18-C 385 42.78 5.86 46.51 78.29 -0.367 叶绿体
BnC03g0566190.1 BnLPAT3-C 534 60.30 5.85 39.86 90.54 -0.057 线粒体，细胞核
BnC03g0576540.1 BnLPAT5-C 432 48.17 9.74 43.64 93.63 -0.214 叶绿体，内质网
BnC03g0615450.1 BnLPAT6-C 286 31.79 9.39 44.60 94.44 -0.030 叶绿体，内质网
BnC03g0616110.1 BnLPAT15-C 333 37.86 8.40 31.42 96.31 0.069 内质网
BnC04g0635520.1 BnLPAT12-C 376 43.16 6.46 48.00 94.63 0.112 内质网
BnC04g0683780.1 BnLPAT10-C 541 61.22 6.68 43.10 90.61 -0.089 线粒体，细胞核
BnC05g0715370.1 BnLPAT8-C 451 49.88 6.33 50.53 77.43 -0.359 叶绿体
BnC06g0752240.1 BnLPAT4-C 391 43.66 9.48 45.11 99.77 0.111 内质网
BnC06g0756180.1 BnLPAT11-C 397 44.50 6.13 54.75 89.82 -0.167 内质网
BnC06g0775100.1 BnLPAT22-C 383 43.04 5.86 49.72 88.54 -0.161 内质网
BnC06g0775680.1 BnLPAT14-C 381 43.06 6.19 50.25 89.00 -0.175 内质网
BnC07g0780780.1 BnLPAT23-C 492 55.49 5.99 40.00 89.15 -0.073 内质网
BnC07g0830090.1 BnLPAT20-C 345 37.98 9.65 42.19 89.54 0.025 叶绿体
BnC08g0842420.1 BnLPAT16-C 456 50.22 6.14 44.19 79.58 -0.302 叶绿体
BnC08g0866950.1 BnLPAT19-C 390 43.73 9.38 46.43 99.51 0.106 内质网
BnC09g0920060.1 BnLPAT21-C 371 42.51 8.92 47.19 92.18 -0.031 内质网
BnUnng0943360.1 BnLPAT9-U 346 40.38 9.01 39.34 90.69 0.032 内质网
BnUnng0994230.1 BnLPAT12-U 380 43.26 8.74 45.79 96.71 0.116 细胞膜，内质网
BnUnng1007920.1 BnLPAT2-U 378 43.65 8.73 38.59 95.37 0.096 内质网

图1

拟南芥、白菜、甘蓝和甘蓝型油菜LPAT成员系统发育进化树 绿色、蓝色、紫色和黄色圆点代表分别来自拟南芥、白菜、甘蓝和甘蓝型油菜的成员。

图2

BnLPAT基因染色体定位与复制关系 不同颜色代表不同的染色体，棕色线段两端的基因代表具有片段复制关系。

图3

BnLPATs保守基序和基因结构 (a) 44个BnLAPT临近加入法聚类树；(b) BnLPAT保守基序，共10个基序被检测，不同的基序用不同颜色的矩形表示；(c) BnLPAT基因结构，编码序列用灰色矩形表示，上、下游非编码区用蓝色矩形表示，内含子序列用线段表示。

图4

BnLPAT蛋白的序列比对 (a) BnLPAT蛋白的ClustalW法多序列比对；(b) 2个保守结构域的LOGO组成。

图5

BnLPAT基因顺式作用元件分析 (a) 44个BnLAPT临近加入法聚类树；(b) 42种顺式作用元件按照相应类型分为4类，热图中的数字代表着元件出现的次数；(c) 22种顺式作用元件在BnLPAT基因上游2000 bp区域出现的位置，不同的顺式作用元件用不同颜色的矩形表示。

图6

BnLPAT基因表达模式 (a) 不同组织部位的BnLPAT基因转录组表达谱；(b) 12个BnLPAT基因在种子发育过程中的表达模式，*：0.01 < P < 0.05；**：P < 0.01。下同。

图7

4个BnLPAT基因在不同品种的表达模式

[1] 施文华, 严茂林, 刘昌勇, 等. 我国油料进口贸易的结构特征及对策分析. 中国油脂, 2023, 48(8):1-8.
[2] 何微, 李俊, 王晓梅, 等. 全球油菜产业现状与我国油菜产业问题、对策. 中国油脂, 2022, 47(2):1-7.
[3] 王汉中. 中国油菜品种改良的中长期发展战略. 中国油料作物学报, 2004, 26(3):99-102.
[4] Harwood J L, Guschina I A. Regulation of lipid synthesis in oil crops. FEBS Letters, 2013, 587(13):2079-2081.
doi: 10.1016/j.febslet.2013.05.018 pmid: 23684640
[5] Gibellini F, Smith T K. The Kennedy pathway—De novo synthesis of phosphatidylethanolamine and phosphatidylcholine. IUBMB Life, 2010, 62(6):414-428.
doi: 10.1002/iub.337 pmid: 20503434
[6] Xu C, Shanklin J. Triacylglycerol metabolism, function, and accumulation in plant vegetative tissues. Annual Review of Plant Biology, 2016,67:179-206.
[7] Körbes A P, Kulcheski R, Margis R, et al. Molecular evolution of the lysophosphatidic acid acyltransferase (LPAAT) gene family. Molecular Phylogenetics and Evolution, 2016,96:55-69.
[8] Leung D W. The structure and functions of human lysophosphatidic acid acyltransferases. Frontiers in Bioscience, 2001,6:944-953.
[9] Maisonneuve S, Bessoule J J, Lessire R, et al. Expression of rapeseed microsomal lysophosphatidic acid acyltransferase isozymes enhances seed oil content in Arabidopsis. Plant Physiology, 2010, 152(2):670-684.
doi: 10.1104/pp.109.148247 pmid: 19965969
[10] 张军平, 江木兰, 龚阳敏, 等. 三角褐指藻LPAAT基因在酵母中表达对脂肪酸的影响. 中国油料作物学报, 2012, 34(5):483-488.
[11] Kim H U, Huang A H. Plastid lysophosphatidyl acyltransferase is essential for embryo development in Arabidopsis. Plant Physiology, 2004, 134(3):1206-1216.
[12] Wang N H, Ma J J, Pei W F, et al. A genome-wide analysis of the lysophosphatidate acyltransferase (LPAAT) gene family in cotton: organization, expression, sequence variation, and association with seed oil content and fiber quality. BMC Genomics, 2017, 18(1):218.
doi: 10.1186/s12864-017-3594-9 pmid: 28249560
[13] 陈四龙. 花生油脂合成相关基因的鉴定与功能研究. 北京: 中国农业科学院, 2013.
[14] 周雅莉, 黄旭升, 郝月茹, 等. 紫苏溶血磷脂酸酰基转移酶基因的克隆与功能分析. 生物工程学报, 2022, 38(8):3014-3028.
[15] Song J M, Guan Z, Hu J, et al. Eight high-quality genomes reveal pan-genome architecture and ecotype differentiation of Brassica napus. Nature Plants, 2020, 6(1):34-45.
[16] Lian Z, Nguyen C D, Liu L, et al. Application of developmental regulators to improve in planta or in vitro transformation in plants. Plant Biotechnology Journal, 2022, 20(8):1622-1635.
[17] Larkin M A, Blackshields G, Brown N P, et al. Clustal W and Clustal X version 2.0. Bioinformatics, 2007, 23(21):2947-2948.
doi: 10.1093/bioinformatics/btm404 pmid: 17846036
[18] Kumar S, Stecher G, Tamura K. MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Molecular Biology and Evolution, 2016, 33(7):1870-1874.
doi: 10.1093/molbev/msw054 pmid: 27004904
[19] Gu Z L, Cavalcanti A, Chen F C, et al. Extent of gene duplication in the genomes of Drosophila, nematode, and yeast. Molecular Biology and Evolution, 2002, 19(3):256-262.
[20] Chen C C, Chen H, Zhang Y, et al. TBtools: an integrative toolkit developed for interactive analyses of big biological data. Molecular Plant, 2020, 13(8):1194-1202.
doi: S1674-2052(20)30187-8 pmid: 32585190
[21] Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2-∆∆CT method. Methods, 2001, 25(4):402-408.
doi: 10.1006/meth.2001.1262 pmid: 11846609
[22] Mishra P, Singh U, Pandey C M, et al. Application of student's t-test, analysis of variance, and covariance. Annals of Cardiac Anaesthesia, 2019, 22(4):407-411.
[23] Taylor J S, Raes J. Duplication and divergence: the evolution of new genes and old ideas. Annual Review of Genetics, 2004,38:615-643.
[24] Agarwal A K, Sukumaran S, Bartz R, et al. Functional characterization of human 1-acylglycerol-3-phosphate-O- acyltransferase isoform 9: cloning, tissue distribution, gene structure, and enzymatic activity. The Journal of Endocrinology, 2007, 193(3):445-457.
[25] Wang P D, Xiong X J, Zhang X B, et al. A review of erucic acid production in brassicaceae oilseeds: progress and prospects for the genetic engineering of high and low-erucic acid rapeseeds (Brassica napus). Frontiers in Plant Science, 2022,13:899076.
[26] Bradley R M, Duncan R E. The lysophosphatidic acid acyltransferases (acylglycerophosphate acyltransferases) family: one reaction, five enzymes, many roles. Current Opinion in Lipidology, 2018, 29(2):110-115.
doi: 10.1097/MOL.0000000000000492 pmid: 29373329
[27] Rao S S, Hildebrand D. Changes in oil content of transgenic soybeans expressing the yeast SLC1 gene. Lipids, 2009, 44(10):945-951.
[28] 郝月茹. 紫苏溶血磷脂酸酰基转移酶基因(PfLPATs)的鉴定及功能分析. 太原:山西农业大学, 2023.
[29] Bin Y, Wakao S, Fan J L, et al. Loss of plastidic lysophosphatidic acid acyltransferase causes embryo-lethality in Arabidopsis. Plant and Cell Physiology, 2004, 45(5):503-510.
pmid: 15169931
[30] Moore R C, Purugganan M D. The early stages of duplicate gene evolution. Proceedings of the National Academy of Sciences of the United States of America, 2003, 100(26):15682-15687.
[31] Kong H Z, Landherr L L, Frohlich M W, et al. Patterns of gene duplication in the plant SKP1 gene family in angiosperms: evidence for multiple mechanisms of rapid gene birth. The Plant Journal, 2007, 50(5):873-885.
[32] Cannon S B, Mitra A, Baumgarten A, et al. The roles of segmental and tandem gene duplication in the evolution of large gene families in Arabidopsis thaliana. BMC Plant Biology, 2004,4:10.
[33] Kim H U, Li Y, Huang A H. Ubiquitous and endoplasmic reticulum-located lysophosphatidyl acyltransferase, LPAT2, is essential for female but not male gametophyte development in Arabidopsis. The Plant Cell, 2005, 17(4):1073-1089.
[34] Kim H U, Vijayan P, Carlsson A S, et al. A mutation in the LPAT1 gene suppresses the sensitivity of fab 1 plants to low temperature. Plant Physiology, 2010, 153(3):1135-1143.
[35] Hishikawa D, Shindou H, Kobayashi S, et al. Discovery of a lysophospholipid acyltransferase family essential for membrane asymmetry and diversity. Proceedings of the National Academy of Sciences of the United States of America, 2008, 105(8):2830-2835.
[36] Schmidt J A, Brown W J. Lysophosphatidic acid acyltransferase 3 regulates Golgi complex structure and function. The Journal of Cell Biology, 2009, 186(2):211-218.
[37] 王芹. 甘蓝型油菜种子发育进程中油脂积累差异及关键基因表达. 重庆:西南大学, 2022.
[1] 邹小云, 官梅, 官春云. 甘蓝型油菜氮素高效吸收的植株形态与生理特性研究[J]. 作物杂志, 2022, (5): 97–103
[2] 林晓阳, 杜德志, 柳海东, 李钧. 特早熟甘蓝型春油菜恢复系核心种质构建[J]. 作物杂志, 2022, (1): 31–37
[3] 蔡东芳,张书芬,王建平,曹金华,文雁成,张书法,何俊平,赵磊,王东国,朱家成. 利用SNP芯片解析油菜杂交种丰油10号的遗传基础[J]. 作物杂志, 2019, (3): 80–85
[4] 何俊平,张书芬,王建平,蔡东芳,曹金华,文雁成,胡坤,赵磊,王东国,朱家成. 甘蓝型双低油菜杂交种丰油10号纯度的SSR鉴定[J]. 作物杂志, 2019, (1): 75–80
[5] 蔡东芳,王建平,张书芬,何俊平,曹金华,文雁成,赵磊,王东国,朱家成. 河南省不同轮作区双低杂交油菜氮磷钾肥的效应研究[J]. 作物杂志, 2018, (6): 130–137
[6] 张振乾, 肖钢, 官春云. 甘蓝型油菜近等基因系构建及应用研究进展[J]. 作物杂志, 2013, (5): 9–13
[7] 王敏, 曲存民, 刘晓兰, 等. 温度胁迫下甘蓝型油菜苗期生理生化指标的研究[J]. 作物杂志, 2013, (2): 53–59
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