高温胁迫下陆地棉转录组差异性分析

doi:10.6048/j.issn.1001-4330.2023.11.003

摘要/Abstract

摘要：

【目的】 研究陆地棉高温胁迫下关键基因的响应,比较基因表达差异,为揭示陆地棉高温胁迫响应的分子机制提供参考。【方法】 利用RNA-Seq对正常温度(26℃)和高温胁迫(42℃)的陆地棉品系YZ1进行转录组测序和数据分析。【结果】 在WN4/WH4,WN12/WH12,WN24/WH24,WN48/WH48共鉴定出383个共有上调基因和234个共有下调基因。将差异基因按GO功能分类,主要富集到上调差异基因和下调差异基因20个类别;差异基因显著富集到类黄酮生物合成途径、苯丙烷代谢、昼夜节律-植物、内质网加工等通路;鉴定出39个转录因子,主要是WRKY和MYB两大家族;Gohir.A08G104100是高温胁迫主要的基因,对植物的耐热性至关重要,Gohir.D08G033300是质体类异戊二烯生物合成的限制酶,对叶绿体发育至关重要。2个差异基因在整个时期表达量差异倍数较为显著。【结论】 高温胁迫和正常温度下在不同时间段,陆地棉的基因转录表达差异。

关键词: RNA-Seq; 陆地棉; 高温胁迫; 差异表达; 代谢通路

Abstract:

【Objective】 To explore the response of key genes to heat stress and compare the differences of gene expression.【Methods】 RNA-Seq was used to sequence and analyze the transcriptome of YZ1 unde²r normal(26℃)and high temperature stress(42℃).【Results】 Gohir.A08G10410A total of 383 up-regulated genes and 234 down regulated genes were identified in WN4/WH4, WN12/WH12, WN24/WH24 and WN48/WH48. According to GO function, the differential genes were mainly enriched into 20 categories, namely, up-regulated differential genes and down-regulated differential genes. According to KEGG function analysis, the differential genes were significantly enriched in flavonoid biosynthesis pathway, phenylalanine metabolism, circadian rhythm-plant and endoplasmic reticulum processing and other pathways. 39 transcription factors were identified, mainly belonging to WRKY and MYB families; Combined with gene annotation, it was found that the expression levels of these two genes were significantly different in the whole period. Gohir.A08G104100 was the main gene of heat stress, which was crucial to the heat tolerance of plants, while Gohir. D08G033300 was a limiting enzyme for plastid isoprene biosynthesis and was critical for chloroplast development.【Conclusion】 Through comparative transcriptome analysis, the difference of gene transcription expression between upland cotton under high temperature stress and normal temperature is compared.

Key words: RNA-Seq; Gossypium hirsutum; heat stress; differential expression; metabolic pathway

中图分类号:

S562

王辉, 董永梅, 郭伟锋, 曹新川, 郭金成, 谢宗铭, 何良荣. 高温胁迫下陆地棉转录组差异性分析[J]. 新疆农业科学, 2023, 60(11): 2618-2626.

WANG Hui, DONG Yongmei, GUO Weifeng, CAO Xinchuan, GUO Jincheng, XIE Zongming, HE Liangrong. Analysis of the difference of transcription groups of upland cotton under heat stress[J]. Xinjiang Agricultural Sciences, 2023, 60(11): 2618-2626.

图/表 8

参考文献 31

[1]	David B. Lobell, Wolfram Schlenker, Justin Costa-Roberts. Climate trends and global crop production since 1980[J]. Science, 2011, 333(6042):616-620. DOI PMID
[2]	Peng S, Huang J, Sheehy J E. Rich yields decline with higher night temperature from global warming[J]. Pro Natl Acad Sci USA, 2004, 101(27):9971-9975. DOI URL
[3]	王秀琴, 段维. 新疆莫索湾高温日数统计特征[J]. 干旱气象, 2014, 32(2):220-225. DOI
	WANG Xiuqin, DUAN Wei. Statistical characteristics of high-temperature days in Mosuowan of Xinjiang[J]. Journal of Arid Meteorology, 2014, 32(2) : 220-225.
[4]	黄帅, 江静. 中国持续性高温事件的时空分析[J]. 南京大学学报(自然科学版), 2012, 48(6):689-700.
	HUANG Shuai, JIANG Jing. The spatial-temporal analysis of persistent high-temperature events in China[J]. Journal of Nanjing University (Natural Sciences), 2012, 48(6) : 689-700.
[5]	尹波. 高温胁迫下番茄Solexa转录组测序及LeCOR413-TM1基因克隆和功能分析[D]. 山东农业大学, 2014.
	Yin Bo. Transcriptional analysis of tomato under high temperature stress via Solexa sequencing and cloning and functional analysis of LeCOR413-TM1 gene[D]. Shandong Agricultural University, 2014.
[6]	徐洪国. 葡萄耐热性评价及不同耐热性葡萄转录组研究[D]. 北京: 中国农业大学, 2014.
	Xu Hongguo. Evaluation of grape heat tolerance and transcriptome of different heat tolerance of grape[D]. Beijing: China Agricultural University, 2014.
[7]	Elizabeth R. Waters, Garrett J. Lee,Elizabeth Vierling. Evolution, structure and function of the small heat shock proteins in plants[J]. Journal of Experimental Botany, 1996, 47(296): 325-338. DOI URL
[8]	Timperio A M, Egidi M G, Zolla L. Proteomics applied on plant abiotic stresses: role of heat shock proteins (HSP)[J]. Journal of Proteomics, 2008, 71: 391-411. DOI PMID
[9]	Kimpel J A, Key J L. Heat shock in plants[J]. Trends in Biochemical Sciences, 1985, 10: 353-357. DOI URL
[10]	Van Montfort R L M, Basha E, Friedrich K L, et al. Crystal structure and assembly of a eukaryotic small heat shock protein[J]. Nature Structural Biology, 2001, 8: 1025-1030. DOI PMID
[11]	Arasakesary S. J., Manonmani S, Pushpam R, et al. New temperature sensitive genic male sterile lines with better outcrossing ability for production of two-line hybrid in rice[J]. Rice Science, 2015, 22(1):49-52. DOI
[12]	Raafat EN, Saber S, Yonnelle DM, et al. Microsatellite-aided screening for fertility restoration genes (Rf) facilitates hybrid improvement[J]. Rice Science, 2016, 23(3):160-164. DOI
[13]	Frey FP, Urbany C, Hüttel B, et al. Genome-wide expression profiling and phenotypic evaluation of European maize inbreds at seedling stage in response to heat stress[J]. BMC Genomics, 2015, 16:123. DOI PMID
[14]	Qin D, Wu H, Peng H, et al. Heat stress-responsive transcriptome analysis in heat susceptible and tolerant wheat (Triticum aestivum L.) by using wheat genome array[J]. BMC Genomics, 2008, 9:432. DOI
[15]	Lee YH, Kim KS, Jang YS, et al. Global gene expression responses to waterlogging in leaves of rape seedlings[J]. Plant Cell Rep, 2014, 33(2):289-299. DOI URL
[16]	Aulakh SS, Veilleux RE, Dickerman AW, et al. Characterization and RNA-seq analysis of underperformer, an activation-tagged potato mutant[J]. Plant Mol Biol, 2014, 84(6):635-658. DOI PMID
[17]	Blanco-Ulate B, Cantu D. Tomato transcriptome and mutant analyses suggest a role for plant stress hormones in the interaction between fruit and Botrytis cinerea[J]. Front Plant Science, 2013, 4:142.
[18]	Kim D, Langmead B, Salzberg S L. HISAT: a fast spliced aligner with low memory requirements[J]. Nature Methods, 2015, 12:357. DOI PMID
[19]	Liao Y, Smyth G K, Shi W. FeatureCounts: an efficient general-purpose program for assigning sequence reads to genomic features[J]. Bioinformatics, 2014, 30:923-930. DOI URL
[20]	Love M I, Huber W, Anders S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2[J]. Genome Biol, 2014, 15:550. DOI URL
[21]	Goldstein L D, Cao Y, Pau G, et al. Prediction and Quantification of Splice Events from RNA-Seq Data.[J]. Plos One, 2016, 11(5):e0156132. DOI URL
[22]	Schulze S K, Kanwar R, et al. SERE: Single-parameter quality control and sample comparison for RNA-Seq[J]. BMC genomics, 2012, 13(1): 524. DOI
[23]	Wahid A, Farooq M, Hussain I, et al. Responses and management of heat stress in plants[J]. In: Ahmad P, Prasad MNV, editors. Environmental adaptations and stress tolerance of plants in the era of climate change. New York: Springer, 2012, p. 135-157.
[24]	李川, 乔江方, 朱卫红, 等. 玉米自交系响应花粒期高温胁迫差异表达基因的分析[J]. 华北农学报, 2019, 34(1):1-11. DOI
	LI Chuan, QIAO Jiangfang, ZHU Weihong, et al. Differential expressed of high temperature stress in anthesis stage related genes of maize inbred lines[J]. Acta agriculture boreali-sinica, 2019, 34(1):1-11.
[25]	翟秀明, 唐敏, 李解, 等. 基于RNA-Seq技术的茶树响应高温胁迫转录组差异性分析[J]. 分子植物育种, 2020, 18(17):5629-5637.
	ZHAI Xiuming, TANG Min, LI Jie, et al. Difference analysis of heat stress-responsive transcriptome of Camellia sinensis based on RNA-Seq technology[J]. Molecular plant breeding, 2020, 18(17):5629-5637.
[26]	许小芳. 高温胁迫下玉米叶片的转录组和蛋白质组分析[D]. 郑州: 河南农业大学, 2019.
	Xu Xiaofang. Transcriptome and proteome analysis of maize leaf under high temperature stress[D]. Zhenzhou: Henan Agricultural University, 2019.
[27]	赵贝贝, 叶蕴灵, 王莉, 等. 银杏类黄酮响应非生物胁迫研究进展[J]. 扬州大学学报(农业与生命科学版), 2018, 39(3):106-112.
	ZHAO Beibei, YE Yunling, WANG Li, et al. The research progress on response to abiotic stress by flavonoid in Ginkgo biloba[J]. Journal of Yangzhou university (Agriculture and life science edition), 2018, 39(3):106-112.
[28]	李鑫雨, 何媛, 代娅, 等. 多年生黑麦草温度胁迫差异表达转录因子的比较转录组分析[J]. 西北植物学报, 2020, 40(5):773-784.
	LI Xinyu, HE Yuan, DAI Ya, et al. Comparative transcriptome analysis reveals differentially expressed transcription factors associated with temperature stresses in Lolium perenne[J]. Acta botanica boreali-occidentalia sinica, 2020, 40(5):773-784.
[29]	姚娜, 刘秀明, 董园园, 等. 转录组的测序方法及应用研究概述[J]. 北方园艺, 2017(12):192-198.
	YAO Na, LIU Xiuming, DONG Yuanyuan, et al. Advances in application and seguencing methods of transcriptome[J]. Northern Horticulture, 2017(12):192-198.
[30]	Wang Z, Gerstein M, Snyder M. RNA-Seq: a revolutionary tool for transcriptomics[J]. Nature reviews genetics, 2009, 10(1):57. DOI PMID
[31]	潘琪. 番茄SYTA的功能及其与Fd Ⅰ互作研究[D]. 重庆: 西南大学, 2018.
	Pan Qi. Function analysis of Solanum lycopersicum SYTA and analyzing the interaction between Solanum lycopersicum SYTA and Nicotiana benthamiana Ferredoxin I[D]. Changqing: Southwest University, 2018.

碱基质量值分值 Quality phred	不正确的碱基识别 Incorrect base identification	碱基正确识别率 Correct base recognition rate(%)
Q10	1/10	90
Q20	1/100	99
Q30	1/1000	99.90
Q40	1/10000	99.99

碱基质量值分值 Quality phred	不正确的碱基识别 Incorrect base identification	碱基正确识别率 Correct base recognition rate(%)
Q10	1/10	90
Q20	1/100	99
Q30	1/1000	99.90
Q40	1/10000	99.99

基因序号Gene_id	基因名称Gene name	描述Description	调控Regulation
Gohir.A11G135500	HSP15.7	15.7 k Da heat shock protein, peroxisomal	down
Gohir.A05G091600	HSP17.3-B	17.3 k Da class I heat shock protein	down
Gohir.D05G092500	HSP17.3-B	17.3 k Da class I heat shock protein	down
Gohir.D05G139900	HSP17.4B	17.4 k Da class ⅡI heat shock protein	down
Gohir.D06G084500	HSP17.6	17.6 k Da class Ⅱ heat shock protein	down
Gohir.D07G106100	HSP18.5-C	18.5 k Da class I heat shock protein	down
Gohir.A08G104100	HSP22	Small heat shock protein, chloro-plastic	down
Gohir.D05G127000	HSP26.5	26.5 k Da heat shock protein, mitochondrial	down
Gohir.D05G096900	HSP70	Heat shock cognate 70 k Da protein	down
Gohir.A13G234100	HSP70	Heat shock 70 k Da protein	down
Gohir.D13G239700	HSP70	Heat shock 70 k Da protein	down
Gohir.D09G208200	HSP70-7	Heat shock 70 k Da protein 7, chloro-plastic	down
Gohir.D09G212600	HSP70-7	Heat shock 70 k Da protein 7, chloro-plastic	down
Gohir.A12G254100	HSP83A	Heat shock protein 83	down
Gohir.D08G135400	HSP83A	Heat shock protein 83	down
Gohir.D03G148100	HSP83A	Heat shock protein 83	down
Gohir.D12G256400	HSP83A	Heat shock protein 83	down

基因序号Gene_id	基因名称Gene name	描述Description	调控Regulation
Gohir.A11G135500	HSP15.7	15.7 k Da heat shock protein, peroxisomal	down
Gohir.A05G091600	HSP17.3-B	17.3 k Da class I heat shock protein	down
Gohir.D05G092500	HSP17.3-B	17.3 k Da class I heat shock protein	down
Gohir.D05G139900	HSP17.4B	17.4 k Da class ⅡI heat shock protein	down
Gohir.D06G084500	HSP17.6	17.6 k Da class Ⅱ heat shock protein	down
Gohir.D07G106100	HSP18.5-C	18.5 k Da class I heat shock protein	down
Gohir.A08G104100	HSP22	Small heat shock protein, chloro-plastic	down
Gohir.D05G127000	HSP26.5	26.5 k Da heat shock protein, mitochondrial	down
Gohir.D05G096900	HSP70	Heat shock cognate 70 k Da protein	down
Gohir.A13G234100	HSP70	Heat shock 70 k Da protein	down
Gohir.D13G239700	HSP70	Heat shock 70 k Da protein	down
Gohir.D09G208200	HSP70-7	Heat shock 70 k Da protein 7, chloro-plastic	down
Gohir.D09G212600	HSP70-7	Heat shock 70 k Da protein 7, chloro-plastic	down
Gohir.A12G254100	HSP83A	Heat shock protein 83	down
Gohir.D08G135400	HSP83A	Heat shock protein 83	down
Gohir.D03G148100	HSP83A	Heat shock protein 83	down
Gohir.D12G256400	HSP83A	Heat shock protein 83	down

记录号 Accession	项目 Term	输入基因数目 Input Gene Number	所有基因数目 All Gene Number	P值 P-value	Q值 Q-value
ko00941	Flavonoid biosynthesis	20 (14.60%)	81 (0.57%)	8.40E-25	2.90E-23
ko01110	Biosynthesis of secondary metabolites	62 (45.26%)	2,573 (18.15%)	5.40E-14	1.20E-12
ko00360	Phenylalanine metabolism	10 (7.30%)	104 (0.73%)	4.80E-09	5.50E-08
ko04915	Estrogen signaling pathway	9 (6.57%)	89 (0.63%)	1.40E-08	1.30E-07
ko01100	Metabolic pathways	76 (55.47%)	4,904 (34.59%)	1.70E-07	1.30E-06
ko04612	Antigen processing and presentation	10 (7.30%)	175 (1.23%)	1.00E-06	7.00E-06
ko04940	Type I diabetes mellitus	4 (2.92%)	24 (0.17%)	2.90E-06	1.80E-05
ko05134	Legionellosis	8 (5.84%)	145 (1.02%)	1.10E-05	6.00E-05
ko04621	NOD-like receptor signaling pathway	4 (2.92%)	32 (0.23%)	1.30E-05	6.30E-05
ko04141	Protein processing in endoplasmic reticulum	17 (12.41%)	593 (4.18%)	1.60E-05	7.30E-05
ko00072	Synthesis and degradation of ketone bodies	3 (2.19%)	17 (0.12%)	1.80E-05	7.30E-05
ko04712	Circadian rhythm-plant	7 (5.11%)	120 (0.85%)	2.10E-05	8.00E-05