棉花学报 ›› 2022, Vol. 34 ›› Issue (2): 93-106.doi: 10.11963/cs20210018
徐婷婷1(),张弛1,冯震1,刘其宝1,李黎贝1,俞啸天1,张雅楠1,喻树迅1,2,*(
)
收稿日期:
2021-03-09
出版日期:
2022-03-15
发布日期:
2022-07-19
通讯作者:
喻树迅
E-mail:yushuxun@zafu.edu.cn
作者简介:
徐婷婷(1995―),女,硕士研究生, 基金资助:
Xu Tingting1(), Zhang Chi1, Feng Zhen1, Liu Qibao1, Li Libei1, Yu Xiaotian1, Zhang Yanan1, Yu Shuxun1, 2, *(
)
Received:
2021-03-09
Online:
2022-03-15
Published:
2022-07-19
Contact:
Yu Shuxun
E-mail:yushuxun@zafu.edu.cn
摘要:
【目的】 对肌醇-1-磷酸合酶(myo-inositol-1-phosphate synthase,MIPS)基因GhMIPS1A进行功能分析,探究其在陆地棉纤维发育及抗逆过程中的作用。【方法】 通过系统发育分析、基因结构分析、保守基序分析、荧光定量分析、烟草瞬时转化、拟南芥过表达试验、病毒诱导的基因沉默(virus-induced gene silencing,VIGS)技术等对GhMIPS1A基因进行研究。【结果】 从陆地棉TM-1中克隆获得GhMIPS1A基因,通过生物信息学分析发现该基因编码蛋白中存在4个高度保守的序列,分别为GWGGNNG、LWTANTERY、NGSPQNTFVPGL和SYNHLGNNDG。亚细胞定位结果显示,GhMIPS1A蛋白位于细胞膜。在拟南芥中过表达GhMIPS1A基因可促使拟南芥根长显著增加,肌醇含量提高1倍以上。空间表达模式分析表明,GhMIPS1A基因在根、茎、叶、纤维中优势表达,且在高衣分品种新陆早18号中的表达量显著高于其在低衣分品种德字棉531中的表达量;表达模式分析表明,GhMIPS1A基因在纤维发育早期阶段表达量较高。利用VIGS技术沉默TM-1的GhMIPS1A基因,3个株系的棉纤维密度显著降低,衣分分别降低4.48、4.93、3.95百分点,肌醇含量分别降低31.83%、32.90%、29.46%。在干旱处理或盐处理下,GhMIPS1A基因的表达量均表现出先升高后降低的趋势。【结论】 GhMIPS1A在棉纤维发育中发挥积极作用,并且能够响应干旱胁迫和盐胁迫,可以为培育优质高产、耐盐耐旱的棉花新品种提供基因资源和遗传基础。
徐婷婷, 张弛, 冯震, 刘其宝, 李黎贝, 俞啸天, 张雅楠, 喻树迅. 陆地棉基因GhMIPS1A的克隆及功能分析[J]. 棉花学报, 2022, 34(2): 93-106.
Xu Tingting, Zhang Chi, Feng Zhen, Liu Qibao, Li Libei, Yu Xiaotian, Zhang Yanan, Yu Shuxun. Cloning and functional analysis of GhMIPS1A gene in upland cotton (Gossypium hirsutum L.)[J]. Cotton Science, 2022, 34(2): 93-106.
表1
本研究中使用的引物"
引物名称Primer name | 引物序列Primer sequence | 用途Purpose |
---|---|---|
GhMIPS1A-F | TGGGGCAAAAGAGAAATAA | 基因克隆 |
GhMIPS1A-R | AAGGGAAAATGGTCCTAAAG | Gene cloning |
GhMIPS1A-XbaI-F | CTCTAGATGGGGCAAAAGAGAAATAA | 构建表达载体 |
GhMIPS1A-SacI-R | CGAGCTCAAGGGAAAATGGTCCTAAAG | Construction of expression vector |
GhMIPS1A-qF | GCAAAAGCAACTGAGACCCTAC | 检测GhMIPS1A的表达 |
GhMIPS1A-qR | ACCTCTCAGTGTTTGCAGTCC | qRT-PCR of GhMIPS1A |
GhHistone3-F | TCAAGACTGATTTGCGTTTCCA | 检测GhHistone3的表达 |
GhHistone3-R | GCGCAAAGGTTGGTGTCTTC | qRT-PCR of GhHistone 3 |
VIGS-F | GGACTAGTGCAAAAGCAACTGAGACCCTAC | VIGS载体构建 |
VIGS-R | TTGGCGCGCCACCTCTCAGTGTTTGCAGTCC | Construction of VIGS vector |
pCAMBIA3301-GFP-F | CGAGCTCGTGGGGCAAAAGAGAAATAA | 亚细胞定位 |
pCAMBIA3301-GFP-R | GGACTAGTCTTGTATTCCAAAATCATGTTGTT | Subcellular localization |
[1] | Hallman M, Epstein B L. Role of myo-inositol in the synthesis of phosphatidylglycerol and phosphatidylinositol in the lung[J/OL]. Biochemical and Biophysical Research Communications, 1980, 92(4): 1151-1159[2021-03-01]. https://doi.org/10.1016/0006-291x(80)90407-6. |
[2] | Toker A. The synthesis and cellular role of phosphatidylinositol 4, 5-bisphosphate[J/OL]. Current Opinion in Cell Biology, 1998, 10(2): 254-261[2021-03-01]. https://doi.org/10.1016/s0955-0674(98)80148-8. |
[3] | Otto J C, Kelly P, Chiou S T, et al. Alterations in an inositol phosphate code through synergistic activation of a G protein and inositol phosphate kinases[J/OL]. Proceedings of the National Academy of Sciences of the United States of America, 2007, 104(40): 15653-15658[2021-03-01]. https://doi.org/10.1073/pnas.0705729104. |
[4] | Raboy V. Myo-inositol-1, 2, 3, 4, 5, 6-hexakisphosphate[J/OL]. Phytochemistry, 2003, 64(6): 1033-1043[2021-03-01]. https://doi.org/10.1016/s0031-9422(03)00446-1. |
[5] | 张梦, 谢益民, 杨海涛, 等. 肌醇在植物体内的代谢概述——肌醇作为细胞壁木聚糖和果胶前体物的代谢途径[J]. 林产化学与工业, 2013, 33(5): 106-114. |
Zhang Meng, Xie Yimin, Yang Haitao, et al. Myo-inositol metamolism as the precursor of xylan and pectin in plant[J]. Chemistry and Industry of Forest Products, 2013, 33(5): 106-114. | |
[6] | Majumder A L, Johnson M D, Henry S A. 1L-myo-inositol-1-phosphate synthase[J/OL]. Biochimica Biophysica Acta, 1997, 1348(1/2): 245-256[2021-03-01]. https://doi.org/10.1016/s0014-5793(03)00974-8. |
[7] | Latrasse D, Jegu T, Meng P H, et al. Dual function of MIPS1 as a metabolic enzyme and transcriptional regulator[J/OL]. Nucleic Acids Research, 2013, 41(5): 2907-2917[2021-03-01]. https://doi.org/10.1093/nar/gks1458. |
[8] | Bachhawat N, Mande S C. Complex evolution of the inositol-1-phosphate synthase gene among archaea and eubacterial[J/OL]. Trends in Genetics, 2000, 16(3): 111-113[2021-03-01]. https://doi.org/10.1016/s0168-9525(99)01966-6. |
[9] | Zhai H, Wang F B, Si Z Z, et al. A myo-inositol-1-phosphate synthase gene, IbMIPS1, enhances salt and drought tolerances and stem nematode resistance in transgenic sweet potato[J/OL]. Plant Biotechnology Journal, 2016, 14(2): 592-602[2021-03-01]. https://doi.org/10.1111/pbi.12402. |
[10] | Tan J L, Wang C Y, Xiang B, et al. Hydrogen peroxide and nitric oxide mediated cold and dehydration-induced myo-inositol phosphate synthase that confers multiple resistances to abiotic stresses[J/OL]. Plant Cell & Environment, 2013, 36(2): 2907-2917[2021-03-01]. https://doi.org/10.1111/j.1365-3040.2012.02573.x. |
[11] | Murphy A M, Otto B, Brearley C A, et al. A role for inositol hexakisphosphate in the maintenance of basal resistance to plant pathogens[J/OL]. The Plant Journal, 2008, 56(4): 638-652[2021-03-01]. https://doi.org10.1111/j.1365-313X.2008.03629.x. |
[12] | Donahue J L, Alford S R, Torabinejad J, et al. The Arabidopsis myo-inositol-1-phosphate synthase1 gene is required for myo-inositol synthesis and suppression of cell death[J/OL]. The Plant Cell, 2010, 22(3): 888-903[2021-03-01]. https://doi.org/10.1105/tpc.109.071779. |
[13] | Keller R, Brearley C A, Rethewey R N, et al. Reduced inositol content and altered morphology in transgenic potato plants inhibited for 1-D-myo-inositol 3-phosphate synthase[J/OL]. The Plant Journal, 1998, 16(3): 403-410[2021-03-01]. https://doi.org/10.1034/j.1399-5618.2002.00108.x. |
[14] | Hu L Y, Zhou J, Ren G J, et al. Myo-inositol mediates reactive oxygen species-induced programmed cell death via salicylic acid-dependent and ethylene-dependent pathways in apple[J/OL]. Horticulture Research, 2020, 7: 138-145[2021-03-01]. https://doi.org/10.1038/s41438-020-00357-2. |
[15] | Majee M S, Maitra K G, Dastidar, et al. A novel salt-tolerant L-myo-inositol-1-phosphate synthase from Porteresia coarctata (Roxb.) tateoka, a halophytic wild rice: molecular cloning, bacterial overexpression, characterization, and functional introgression into tobacco-conferring salt tolerance phenotype[J/OL]. The Journal of Biological Chemistry, 2004, 279(27): 28539-28552[2021-03-01]. https://doi.org/10.1074/jbc.M310138200. |
[16] | 杨楠. 杨树肌醇代谢关键酶基因的表达及功能研究[D]. 烟台: 鲁东大学, 2017. |
Yang Nan. Expression and function of inositol metabolism key enzyme gene in poplar[D]. Yantai: Ludong University, 2017. | |
[17] | 郝广龙. 玉米肌醇-1-磷酸合成酶基因 ZmMIPS 的克隆及其抗旱功能鉴定[D]. 杨凌:西北农林科技大学, 2019. |
Hao Guanglong.. Cloning of maize myo-inositol-1-phosphate synthase gene (ZmMIPS) and functional characterization of its drought tolerance[D]. Yangling: Northwest A&F University, 2019. | |
[18] | Qin Y M, Zhu Y X. How cotton fibers elongate: a tale of linear cell-growth mode[J/OL]. Current Opinion in Plant Biology, 2011, 14(1): 106-111[2021-03-01]. https://doi.org/10.1016/j.pbi.2010.09.010. |
[19] | Wendel J F, Flagel L E, Adams K L. Jeans, genes, and genomes: cotton as a model for studying polyploidy[M/OL]. SoltisP S, SoltisD. Polyploidy and genome evolution. Berlin: Springer, 2012, 181-207[2021-03-01]. https://doi.org/10.1007/978-3-642-31442-1. |
[20] | 喻树迅, 范术丽, 王寒涛, 等. 中国棉花高产育种研究进展[J/OL]. 中国农业科学, 2016, 49(18): 3465-3476[2021-03-01]. https://doi.org/10.3864/j.issn.0578-1752.2016.18.001. |
Yu Shuxun, Fan Shuli, Wang Hantao, et al. Progresses in research on cotton high yield breeding in China[J/OL]. Scientia Agricultura Sinica, 2016, 49(18): 3465-3476[2021-03-01]. https://doi.org/10.3864/j.issn.0578-1752.2016.18.001. | |
[21] | Shen X L, Guo W Z, Lu Q X, et al. Genetic mapping of quantitative trait loci for fiber quality and yield trait by RIL approach in upland cotton[J/OL]. Euphytica, 2007, 155(3): 371-380 [2021-03-01]https://doi.org/10.1007/s10681-006-9338-6. |
[22] | Ma Rendi, Wang Yangsong, Wang Fei, et al. A cotton (Gossy-pium hirsutum) myo-inositol-1-phosphate synthase (GhMIPS1D) gene promotes root cell elongation in Arabidopsis[J/OL]. International Journal of Molecular Sciences, 2019, 20(5): 1224[2021-03-01]. https://doi.org/10.3390/ijms20051224. |
[23] | 黄毅, 张玉龙. 保护地生产条件下的土壤退化问题及其防治对策[J/OL]. 土壤通报, 2004, 35(2): 212-216[2021-03-01]. https://doi.org/10.19336/j.cnki.trtb.2004.02.027. |
Huang Yi, Zhang Yulong. The soil degradation problem in green-house and control counter-measures[J/OL]. Chinese Journal of Soil Science, 2004, 35(2): 212-216[2021-03-01]. https://doi.org/10.19336/j.cnki.trtb.2004.02.027. | |
[24] | Ullah A, Nisar M, Ali H, et al. Drought tolerance improvement in plants: an endophytic bacterial approach[J/OL]. Applied Microbiology and Biotechnology, 2019, 103(18): 7385-7397[2021-03-01]. https://doi.org/10.1007/s00253-019-10045-4. |
[25] | Zhang H, Li Y Y, Zhu J K. Developing naturally stress-resistant crops for a sustainable agriculture[J/OL]. Nature Plants, 2018, 4(12): 989-996[2021-03-01]. https://doi.org/10.1038/s41477-018-0309-4. |
[26] | Loewus F A. Inositol and plant cell wall polysaccharide bioge-nesis[J/OL]. Cell Biochemistry and Function, 2006, 39: 21-45[2021-03-01]. https://doi.org/10.1007/0-387-27600-9_2. |
[27] | 宿俊吉. 陆地棉早熟与产量纤维品质性状的全基因组关联分析及候选基因筛选[D]. 杨凌: 西北农林科技大学, 2017. |
Su Junji. identification of candidate genes and genome-wide association studies of traits related to early maturity, yield and fiber quality in upland cotton (Gossypium hirsutum L.)[D]. Yangling: Northwest A&F University, 2017. | |
[28] | Gu Z H, Huang C J, Li F F, et al. A versatile system for functional analysis of genes and microRNAs in cotton[J/OL]. Plant Biotechnology Journal, 2014, 12(5): 638-649[2021-03-01]. https://doi.org/10.1111/pbi.12169. |
[29] | 郭亚宁. NAC转录因子在陆地棉叶片衰老中的作用[D]. 杨凌: 西北农林科技大学, 2017. |
Guo Yaning. The function of NAC transcription factors in leaf senescence of upland cotton[D]. Yangling: Northwest A&F University, 2017. | |
[30] | 赵凤利. 棉花叶片衰老相关基因GhNAC12的功能分析[D]. 北京: 中国农业科学院, 2014. |
Zhao Fengli. Functional analysis of GhNAC12, a leaf sene-scence-related gene in cotton (Gossypium hirsutum L.)[D]. Beijing: Chinese Academy of Agricultural Sciences, 2014. | |
[31] | 吕淑霞. 基础生物化学实验指导[M]. 北京: 中国农业出版社, 2003. |
Lü Shuxia. Basic biological chemistry experiment guidance[M]. Beijing: China Agriculture Press, 2003. | |
[32] | 李合生. 植物生理生化实验原理和技术[M]. 北京: 高等教育出版社, 2000: 164-260. |
Li Hesheng. Principles and techniques of plant physiology and biochemistry experiments[M]. Beijing: Higher Education Press, 2000: 164-260. | |
[33] | Abid G, Silue S, Muhovski Y, et al. Role of myo-inositol phosphate synthase and sucrose synthase genes in plant seed development[J/OL]. Gene, 2009, 439(1/2): 1-10[2021-03-01]. https://doi.org/10.1016/j.gene.2009.03.007. |
[34] | Cui M, Liang D, Wu S, et al. Isolation and developmental expression analysis of L-myo-inositol-1-phosphate synthase in four Actinidia species[J/OL]. Plant Physiology and Biochemistry, 2013, 73: 351-358[2021-03-01]. https://doi.org/10.1016/j.plaphy.2013.10.015. |
[35] | Basak P, Maitra-Majee S, Das J K, et al. An evolutionary analysis identifies a conserved pentapeptide stretch containing the two essential lysine residues for rice L-myo-inositol 1-phosphate synthase catalytic activity[J/OL]. PLoS One, 2017, 12(9): 73-78[2021-03-01]. https://doi.org/10.1371/journal.pone.0185351. |
[36] | Pang C Y, Wang H, Pang Y, et al. Comparative proteomics indicates that biosynthesis of pectic precursors is important for cotton fiber and Arabidopsis root hair elongation[J/OL]. Molecu-lar & Cellular Protemomics, 2010, 9(9): 2019-2033[2021-03-01]. https://doi.org/10.1074/mcp.M110.000349. |
[37] | Li H B, Qin Y M, Pang Y, et al. A cotton ascorbate peroxidase is involved in hydrogen peroxide homeostasis during fiber cell development[J/OL]. New Phytologist, 2007, 175(3): 462-471[2021-03-01]. https://doi.org/10.1111/j.1469-8137.2007.02120.x. |
[38] | Kusuda H, Koga W, Kusano M, et al. Ectopic expression of myo-inositol 3-phosphate synthase induces a wide range of metabolic changes and confers salt tolerance in rice[J/OL]. Plant Science, 2015, 232: 49-56[2021-03-01]. https://doi.org/10.1016/j.plantsci.2014.12.009. |
[39] | Liu G J, Xiao G H, Liu N J, et al. Targeted lipidomics studies reveal that linolenic acid promotes cotton fiber elongation by activating phosphatidylinositol and phosphatidylinositol monophosphate biosynthesis[J/OL]. Molecular Plant, 2015, 8(6): 911-921[2021-03-01]. https://doi.org/10.1016/j.molp.2015.02.010. |
[40] | Han S J, Yu B J, Wang Y, et al. Role of plant autophagy in stress response[J/OL]. Protein & Cell, 2011, 2(10): 784-791[2021-03-01]. https://doi.org/10.1007/s13238-011-1104-4. |
[41] | 刘楚楚, 吴玉香. 棉花RCI2基因家族的鉴定及表达分析[J]. 山西农业大学学报 (自然科学版), 2021, 41(4): 41-49. |
Liu Chuchu, Wu Yuxiang. Identification and expression analysis of RCI2 gene family in cotton[J]. Journal of Shanxi Agricultural University (Natural Science Edition), 2021, 41(4): 41-49. |
[1] | 杨炳磊, 许好标, 李黎贝, 冯震, 刘林, 喻树迅. 陆地棉株高相关基因GhGA20ox6的克隆及功能初探[J]. 棉花学报, 2022, 34(4): 275-285. |
[2] | 晁毛妮, 董洁, 胡根海, 黄玲, 张金宝, 付远志, 王清连. 陆地棉AGPase基因家族的鉴定及表达分析[J]. 棉花学报, 2022, 34(4): 299-312. |
[3] | 赵曾强,张析,李潇玲,张薇. GhEIN3基因对棉花枯萎病胁迫响应的功能分析[J]. 棉花学报, 2022, 34(3): 173-186. |
[4] | 吴健锋,樊志浩,武连杰,胡晓旺,韩知里,高巍,龙璐. 陆地棉衰老相关基因GhSAG101的克隆及抗病功能分析[J]. 棉花学报, 2022, 34(3): 187-197. |
[5] | 田一波,潘奥,陈劲,周仲华,袁小玲,刘志. 陆地棉ACX基因家族的鉴定与功能分析[J]. 棉花学报, 2022, 34(3): 215-226. |
[6] | 张素君, 李兴河, 王海涛, 唐丽媛, 蔡肖, 刘存敬, 张香云, 张建宏. 陆地棉主要育种性状SSR关联位点的验证及优异材料鉴定(2022年11月8日更新)[J]. 棉花学报, 2022, 34(2): 120-136. |
[7] | 陈琴,李多露,赵杰银,高文举,陈全家,曲延英. 陆地棉UDPGP基因家族的鉴定及抗旱性分析[J]. 棉花学报, 2022, 34(1): 12-22. |
[8] | 上官小霞,曹俊峰,杨琴莉,吴霞. 棉花纤维发育的分子机理研究进展[J]. 棉花学报, 2022, 34(1): 33-47. |
[9] | 贺浪,张华崇,司宁,简桂良. 陆地棉GhBZR1基因的克隆及功能研究[J]. 棉花学报, 2021, 33(6): 435-447. |
[10] | 李丹,赵存鹏,赵丽英,刘旭,刘素恩,王凯辉,王兆晓,耿军义,郭宝生. 棉花类表皮特异性分泌糖蛋白基因GhA01EP1的克隆和功能分析[J]. 棉花学报, 2021, 33(6): 448-458. |
[11] | 王雪慧,陈丽锦,赵若林,程海亮,张友平,王巧连,吕丽敏,宋国立,左东云. 陆地棉纤维起始期优势表达基因GhCRPK1的克隆及功能研究[J]. 棉花学报, 2021, 33(6): 459-468. |
[12] | 姜辉,郑锦秀,王永翠,张超,王秀丽,陈莹,高明伟,王家宝,柴启超,赵军胜. 陆地棉L-D1等位基因特异性分子标记的开发及应用[J]. 棉花学报, 2021, 33(5): 412-421. |
[13] | 卞英杰,王寒涛,魏恒玲,张蒙,李弈,喻树迅. 陆地棉叶片发育相关基因GhRH39克隆与功能分析[J]. 棉花学报, 2021, 33(4): 319-327. |
[14] | 张岚,程琦,梁士辰,邓雨潇,潘玉欣. 棉花UGPase基因鉴定与生物信息学分析[J]. 棉花学报, 2021, 33(4): 337-346. |
[15] | 程成,李斌,王雅丽,赵楠,苏莹,聂虎帅,华金平. 转FBP7::iaaM基因陆地棉育种应用初报[J]. 棉花学报, 2021, 33(4): 368-376. |
|