基于多平台基因表达数据的水稻干旱和盐胁迫相关基因预测

doi:10.3724/SP.J.1006.2021.02084

作物学报 ›› 2021, Vol. 47 ›› Issue (12): 2423-2439.doi: 10.3724/SP.J.1006.2021.02084

• 作物遗传育种·种质资源·分子遗传学 • 上一篇下一篇

基于多平台基因表达数据的水稻干旱和盐胁迫相关基因预测

刘亚文¹(), 张红燕¹^,²^,^*(), 曹丹², 李兰芝²

¹湖南农业大学信息与智能科学技术学院, 湖南长沙 410128
²湖南农业大学 / 湖南省农业大数据分析与决策工程技术研究中心, 湖南长沙 410128

收稿日期:2020-11-30 接受日期:2021-04-26 出版日期:2021-12-12 网络出版日期:2021-06-02
通讯作者: 张红燕
作者简介:E-mail: lyw20201022@163.com
基金资助:
湖南省教育厅科学研究重点项目(18A105);长沙市工业科技特派员项目(201845);湖南农业大学“双一流”建设项目(SYL2019075)

Prediction of drought and salt stress-related genes in rice based on multi-platform gene expression data

LIU Ya-Wen¹(), ZHANG Hong-Yan¹^,²^,^*(), CAO Dan², LI Lan-Zhi²

¹College of Information and Intelligence, Hunan Agricultural University, Changsha 410128, Hunan, China
²Hunan Engineering and Technology Research Centre for Agricultural Big Data Analysis and Decision-making, Hunan Agricultural University, Changsha 410128, Hunan, China

Received:2020-11-30 Accepted:2021-04-26 Published:2021-12-12 Published online:2021-06-02
Contact: ZHANG Hong-Yan
Supported by:
Key Scientific Research Project of Hunan Education Department(18A105);Special Commissioner Project of Changsha City for Industrial Science and Technology(201845);“Double First-class” Construction Project of Hunan Agricultural University(SYL2019075)

摘要/Abstract

摘要：

基于多平台基因表达数据挖掘水稻胁迫相关基因, 可增加关键基因预测的可靠性, 获得更具普适意义的结果。本研究从NCBI数据库中收集了与水稻非生物胁迫相关的94份affymetrix基因芯片数据和42份RNA-seq转录组数据。首先对同一类型同一胁迫相关的多个数据集以数据转换法融合, 得到干旱胁迫相关的affymetrix数据集D_affy和RNA-seq数据集D_rnaseq, 盐胁迫相关的affymetrix数据集S_affy和RNA-seq数据集S_rnaseq; 接着对4个数据集分别基于Pearson线性相关系数的经典WGCNA法和基于MIC非线性相关系数的改进WGCNA法进行基因共表达网络分析, 共获取胁迫相关的8个Hub基因集; 进一步, 对同一胁迫相关的Hub基因进行整合分析, 得到最终的水稻干旱胁迫相关Hub基因1936个、盐胁迫相关的Hub基因1504个。最后, 从预测性能、富集分析、文献报道、STRING在线互作分析和Cytoscape可视化分析等多角度解析Hub基因的生物学意义。结果显示: Hub基因整体预测性能较优, 且大多富集到了与干旱/盐胁迫相关的通路上, 其中有文献已报道的干旱胁迫响应基因31个和盐胁迫响应基因22个。此外, 通过对Hub基因的互作分析, 预测得到11个干旱胁迫候选基因和5个盐胁迫候选基因。本研究为“高维度、小样本”的农作物基因测序数据的有效分析提供了新思路, 实验结果为抗逆水稻品种研究提供了参考。

关键词: 水稻, 多平台, 干旱胁迫, 盐胁迫, WGCNA-MIC

Abstract:

Mining stress-related genes based on multi-platform gene expression data in rice can increase the reliability of key genes prediction and obtain more universally meaningful results. In this study, 94 affymetrix microarray data and 42 RNA-seq transcriptome data related to rice abiotic stress were collected from NCBI databases. First, multiple datasets related to the same stress on the same type were fused by data conversion method to obtain the affymetrix data set D_affy and RNA-seq data set D_rnaseq related to drought stress, and the affymetrix data set S_affy and the RNA-seq data set S_rnaseq related to salt stress. Then, the four datasets were analyzed by the classical WGCNA method based on Pearson's linear correlation coefficient and the improved WGCNA method based on the MIC nonlinear correlation coefficient respectively, and the eight Hub gene sets related to stress were obtained. Further, the integration analysis of stress-related Hub genes yielded the final 1936 drought stress-related Hub genes and 1504 salt stress-related Hub genes. Finally, the biological significance of Hub gene was analyzed from multiple perspectives, including prediction performance, enrichment analysis, literature report, STRING online interaction analysis, and Cytoscape visualization analysis. The results revealed that the overall prediction performance of Hub genes was better, and most of them were enriched in the pathways related to drought/salt stress. Among them, there were 31 drought stress response genes and 22 salt stress response genes reported in the literatures. In addition, 11 drought stress candidate genes and 5 salt stress candidate genes were predicted using the interaction analysis of Hub genes. In conclusion, This study provides a new idea for the effective analysis of “high-dimensional, small-sample” crop gene sequencing data, and the experimental results provide a reference for the study of stress-resistant rice varieties.

Key words: rice, multi-platform, drought stress, salt stress, WGCNA-MIC

刘亚文, 张红燕, 曹丹, 李兰芝. 基于多平台基因表达数据的水稻干旱和盐胁迫相关基因预测[J]. 作物学报, 2021, 47(12): 2423-2439.

LIU Ya-Wen, ZHANG Hong-Yan, CAO Dan, LI Lan-Zhi. Prediction of drought and salt stress-related genes in rice based on multi-platform gene expression data[J]. Acta Agronomica Sinica, 2021, 47(12): 2423-2439.

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链接本文: https://zwxb.chinacrops.org/CN/10.3724/SP.J.1006.2021.02084

https://zwxb.chinacrops.org/CN/Y2021/V47/I12/2423

图/表 15

图1

附表1

附表2

来源于NCBI的RNA-seq数据(RNA-seq)"

样品编号 Library name	样品描述 Library description	样品类型 Library type	单/双端 Library layout	水稻品种 Rice genotype
SRR3647331	1-leaf_34_day_Salt	RNAseq	Single	Nipponbare
SRR3647329	1-leaf_34_day_Salt	RNAseq	Single	Nipponbare
SRR3647327	1-leaf_34_day_Salt	RNAseq	Single	Nipponbare
SRR3647330	1-leaf_34_day_Control	RNAseq	Single	Nipponbare
SRR3647328	1-leaf_34_day_Control	RNAseq	Single	Nipponbare
SRR3647326	1-leaf_34_day_Control	RNAseq	Single	Nipponbare
ERR266228	1-Seedling_shoots_2_weeks_1h_Control	RNAseq	Single	Nipponbare
ERR266233	1-Seedling_shoots_2_weeks_1h_Control	RNAseq	Single	Nipponbare
ERR266230	1-Seedling_shoots_2_weeks_1h_Control	RNAseq	Single	Nipponbare
ERR266225	1-Seedling_shoots_2_weeks_24h_Control	RNAseq	Single	Nipponbare
ERR266234	1-Seedling_shoots_2_weeks_24h_Control	RNAseq	Single	Nipponbare
ERR266232	1-Seedling_shoots_2_weeks_24h_Control	RNAseq	Single	Nipponbare
ERR266229	1-Seedling_shoots_2_weeks_5h_Control	RNAseq	Single	Nipponbare
ERR266223	1-Seedling_shoots_2_weeks_5h_Control	RNAseq	Single	Nipponbare
ERR266222	1-Seedling_shoots_2_weeks_5h_Control	RNAseq	Single	Nipponbare
ERR266237	1-Seedling_shoots_2_weeks_1h_Salt	RNAseq	Single	Nipponbare
ERR266236	1-Seedling_shoots_2_weeks_1h_Salt	RNAseq	Single	Nipponbare
ERR266235	1-Seedling_shoots_2_weeks_1h_Salt	RNAseq	Single	Nipponbare
ERR266238	1-Seedling_shoots_2_weeks_24h_Salt	RNAseq	Single	Nipponbare
ERR266226	1-Seedling_shoots_2_weeks_24h_Salt	RNAseq	Single	Nipponbare
ERR266231	1-Seedling_shoots_2_weeks_24h_Salt	RNAseq	Single	Nipponbare
ERR266227	1-Seedling_shoots_2_weeks_5h_Salt	RNAseq	Single	Nipponbare
ERR266224	1-Seedling_shoots_2_weeks_5h_Salt	RNAseq	Single	Nipponbare
ERR266221	1-Seedling_shoots_2_weeks_5h_Salt	RNAseq	Single	Nipponbare
SRR7054183	2-Inflorescence_Control	RNAseq	Paired	Nipponbare
SRR7054182	2-Inflorescence_Control	RNAseq	Paired	Nipponbare
SRR7054181	2-Inflorescence_Control	RNAseq	Paired	Nipponbare
SRR7054180	2-Inflorescence_Control	RNAseq	Paired	Nipponbare
SRR7054179	2-Inflorescence_Drought	RNAseq	Paired	Nipponbare
SRR7054178	2-Inflorescence_Drought	RNAseq	Paired	Nipponbare
SRR7054177	2-Inflorescence_Drought	RNAseq	Paired	Nipponbare
SRR7054176	2-Inflorescence_Drought	RNAseq	Paired	Nipponbare
SRR3051752	Drought stress_rep1	RNAseq	Paired	Nipponbare
SRR3051753	Drought stress_rep2	RNAseq	Paired	Nipponbare
SRR3051754	Drought stress_rep3	RNAseq	Paired	Nipponbare
SRR3051755	Well-water_rep1	RNAseq	Paired	Nipponbare
SRR3051756	Well-water_rep2	RNAseq	Paired	Nipponbare
SRR3051757	Well-water_rep3	RNAseq	Paired	Nipponbare
SRR3051740	Well-water_rep1	RNAseq	Paired	Nipponbare
SRR3051741	Well-water_rep2	RNAseq	Paired	Nipponbare
SRR3051742	Well-water_rep3	RNAseq	Paired	Nipponbare
SRR3051743	Drought stress_rep1	RNAseq	Paired	Nipponbare
SRR3051744	Drought stress_rep2	RNAseq	Paired	Nipponbare
SRR3051745	Drought stress_rep3	RNAseq	Paired	Nipponbare

附表2

图2

表1

图3

图4

附图1

附图2

表2

表3

Hub基因的GO富集部分分析结果"

胁迫 Stress	GO条目 GO term	基因数目 Number of genes	基因本体 Ontology	描述 Description	P值 P-value
干旱 Drought	GO:0010033	12	BP	对有机物的响应 Response to organic substance	9.00E-06
	GO:0009719	12	BP	内源性刺激响应 Response to endogenous stimulus	9.00E-06
	GO:0009725	12	BP	激素刺激响应 Response to hormone stimulus	9.00E-06
	GO:0006721	8	BP	萜类化合物代谢过程 Terpenoid metabolic process	2.80E-05
	GO:0007165	37	BP	信号传导Signal transduction	2.20E-05
	GO:0009628	15	BP	对非生物刺激的响应 Response to abiotic stimulus	0.00054
	GO:0006970	5	BP	渗透胁迫响应 Response to osmotic stress	0.0036
	GO:0009415	5	BP	对水的响应Response to water	0.006
	GO:0004872	22	MF	受体活性 Receptor activity	9.40E-13
	GO:0004713	21	MF	蛋白质酪氨酸激酶活性 Protein tyrosine kinase activity	9.00E-12
	GO:0004722	15	MF	蛋白丝氨酸/苏氨酸磷酸酶活性 Protein serine/threonine phosphatase activity	2.70E-05
	GO:0008135	12	MF	翻译因子活性, 核酸结合 Translation factor activity, Nucleic acid binding	0.0099
	GO:0044424	1120	CC	细胞内成分 Intracellular part	0
	GO:0005737	1011	CC	细胞质Cytoplasm	0
	GO:0016020	278	CC	薄膜 Membrane	3.10E-23
盐 Salt	GO:0070887	6	BP	细胞对化学刺激的响应 Cellular response to chemical stimulus	2.00E-06
	GO:0007275	13	BP	多细胞有机体的发育 Multicellular organismal development	5.30E-08
	GO:0043436	52	BP	草酸代谢过程 Oxoacid metabolic process	4.80E-06
	GO:0019752	52	BP	羧酸代谢过程 Carboxylic acid metabolic process	4.80E-06
	GO:0006082	52	BP	有机酸代谢过程 Organic acid metabolic process	5.10E-06
	GO:0010033	11	BP	对有机物的响应 Response to organic substance	4.70E-06
	GO:0009719	11	BP	内源性刺激响应 Response to endogenous stimulus	4.70E-06
	GO:0009416	6	BP	光刺激响应 Response to light stimulus	0.024
	GO:0004872	14	MF	受体活性 Receptor activity	5.70E-08
	GO:0008135	18	MF	翻译因子活性, 核酸结合 Translation factor activity, nucleic acid binding	1.00E-06
	GO:0003743	11	MF	翻译起始因子活性 Translation initiation factor activity	3.00E-05
	GO:0016874	32	MF	连接酶活性 Ligase activity	0.00064
	GO:0016020	225	CC	薄膜 Membrane	4.60E-21
	GO:0009507	20	CC	叶绿体 Chloroplast	4.30E-15
	GO:0005886	18	CC	细胞质膜 Plasma membrane	1.20E-09

表3

表4

图5

图6

表5

候选基因在STRING中的注释"

胁迫 Stress	候选基因 Candidate gene	STRING中名称 Name in STRING	注释 Annotation
干旱Drought	Os01g0733200	HSF11	热应激转录因子C-1b; 与热休克启动子元件(HSE)的DNA特异性结合的转录调控因子。 Heat stress transcription factor C-1b; transcriptional regulator that specifically binds DNA of heat shock promoter elements (HSE).
	Os07g0178600	HSF5	热应激转录因子A-2b; 转录调控因子, 可特异性结合热休克启动子元件(HSE)的DNA; 属于HSF家族, A类亚科。 Heat stress transcription factor A-2b; transcriptional regulator that specifically binds DNA of heat shock promoter elements (HSE); belongs to the HSF family. Class A subfamily.
	Os09g0526600	HSFB2C	热应激转录因子B-2c; 转录调控因子, 可特异性结合热休克启动子元件(HSE)的DNA; 属于HSF家族, B类亚科。 Heat stress transcription factor B-2c; transcriptional regulator that specifically binds DNA of heat shock promoter elements (HSE); belongs to the HSF family. Class B subfamily.
	Os01g0583100	OS01T0583100-01	可能的蛋白质磷酸酶2C 6; 属于PP2C家族。 Probable protein phosphatase 2C 6; belongs to the PP2C family.
	Os03g0231700	OS03T0231700-02	Os03g0231700蛋白; 角鲨烯单加氧酶, 假定表达; cDNA克隆: J033045D18, 完整插入序列。 Os03g0231700 protein; Squalene monooxygenase, putative, expressed; cDNA clone: J033045D18, full insert sequence.
	Os03g0376100	OS03T0376100-01	Os03g0376100蛋白。 Os03g0376100 protein.
胁迫 Stress	候选基因 Candidate gene	STRING中名称 Name in STRING	注释 Annotation
	Os04g0107900	OS04T0107900-02	Os04g0107900蛋白。 Os04g0107900 protein.
	Os01g0840100	OsJ_04024	cDNA克隆: J100050G20, 完整插入序列; 70 kD热激蛋白; Os01g0840100蛋白; 假定的HSP70; 未表征的蛋白质。 cDNA, clone: J100050G20, full insert sequence; 70 kD heat shock protein; Os01g0840100 protein; Putative HSP70; uncharacterized protein.
	Os03g0277300	OsJ_10337	热休克同源70 kD蛋白, 假定表达。 Heat shock cognate 70 kD protein, putative, expressed.
	Os05g0460000	OsJ_18811	Os05g0460000蛋白; 推定的hsp70; cDNA克隆: J090096I11, 完整插入序列; 属于热休克蛋白70家族。 Os05g0460000 protein; Putative hsp70; cDNA, clone: J090096I11, full insert sequence; Belongs to the heat shock protein 70 family.
	Os06g0110200	OsJ_19858	cDNA克隆: 002-135-D09, 完整插入序列; Os06g0110200蛋白; 假定的未表征蛋白OSJNBa0004I20.22; 假定的未表征蛋白P0514G12.46。 cDNA clone: 002-135-D09, full insert sequence; Os06g0110200 protein; putative uncharacterized protein OSJNBa0004I20.22; putative uncharacterized protein P0514G12.46.
盐 Salt	Os06g0727200	CATB	过氧化氢酶同工酶B; 几乎发生在所有有氧呼吸的生物中, 并保护细胞免受过氧化氢的毒性作用。 Catalase isozyme B; occurs in almost all aerobically respiring organisms and serves to protect cells from the toxic effects of hydrogen peroxide.
	Os12g0502300	CYCA2-1	Cyclin-A2-1; 属于细胞周期蛋白家族。Cyclin AB亚家族。 Cyclin-A2-1; belongs to the cyclin family. Cyclin AB subfamily.
	Os03g0821100	OsJ_13143	热休克同源70 kD蛋白2, 假定表达; 热休克蛋白同源物70; Os03g0821100蛋白; cDNA克隆: J023030D03, 完整插入序列。 Heat shock cognate 70 kD protein 2, putative, expressed; heat shock protein cognate 70; Os03g0821100 protein; cDNA clone:J023030D03, full insert sequence.
	Os10g0491801	OsJ_31995	假定泛素/核糖体蛋白S27a融合蛋白;泛素融合蛋白, 假定表达。 Putative ubiquitin/ribosomal protein S27a fusion protein; ubiquitin fusion protein, putative, expressed.
	Os02g0775200	RFC3	复制因子C亚基3; 可能参与DNA复制, 从而调节细胞增殖。 Replication factor C subunit 3; may be involved in DNA replication and thus regulate cell proliferation.

表5

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胁迫 Stress	GEO关联 GEO accession	测序平台 Platform	样本数(对照组/胁迫组) Samples (control/stress)
干旱 Drought	GSE6901	Affymetrix Rice Genome Array (GPL2025)	N = 6 (3/3 drought)
干旱 Drought	GSE21651	Affymetrix Rice Genome Array (GPL2025)	N = 8 (4/4 drought)
	GSE23211	Affymetrix Rice Genome Array (GPL2025)	N = 12 (6/6 drought)
	GSE26280	Affymetrix Rice Genome Array (GPL2025)	N = 36 (18/18 drought)
盐 Salt	GSE6901	Affymetrix Rice Genome Array (GPL2025)	N = 6 (3/3 salt)
盐 Salt	GSE14403	Affymetrix Rice Genome Array (GPL2025)	N = 18 (9/9 salt)
	GSE16108	Affymetrix Rice Genome Array (GPL2025)	N = 8 (4/4 salt)

数据集 Data set	基因数 No. of genes	总样本数 Total samples	对照组样本数 Control samples	胁迫组样本数 Stress samples
干旱芯片数据 Drought stress-related affymetrix dataset (D_affy)	27,344	62	31	31
盐芯片数据 Salt stress-related affymetrix dataset (S_affy)	27,344	32	16	16
干旱RNA-seq数据 Drought stress-related RNA-seq dataset (D_rnaseq)	29,828	20	10	10
盐RNA-seq数据 Salt stress-related RNA-seq dataset (S_rnaseq)	28,425	22	11	11

胁迫 Stress	基因编号RAP_locus	基因符号 Gene symbol	胁迫 Stress	基因编号 RAP_locus	基因符号 Gene symbol
干旱 Drought	Os05g0455500	OsP5CS; OsP5CS1; OsALDH18B1	干旱 Drought	Os03g0286900	OsRCI2-5
干旱 Drought	Os02g0766700	OsbZIP23	干旱 Drought	Os02g0149800	OsPP18
	Os08g0112700	OsMADS26		Os03g0267000	OsHSP18.0-CI; OsMSR3; OsSHSP1
	Os06g0130100	OsSIK1		Os05g0475400	OsAMTR1
	Os09g0552300	OsRPK1		Os08g0408500	OsERF48; OsDRAP1
	Os01g0867300	OsABF1; OsbZIP12		Os04g0676700	OsMYB6
	Os03g0125100	DSM2		Os12g0597500	OsUAH
	Os03g0745000	OsHsfA2a		Os06g0316000	Os2H16
	Os05g0542500	OsLEA3; OsLEA3-1		Os06g0612800	OsiSAP8
	Os02g0671100	MAIF1		Os04g0572400	OsDREB1E
	Os06g0211200	OsAREB1; OsbZIP46; OsABF2; ABL1		Os11g0126900	OsNAC10; ONAC122
	Os05g0437700	EDT1; OsbZIP40		Os11g0707600	OsGL1-11
	Os05g0569300	OsbZIP45		Os03g0805100	SQS
	Os08g0196700	OsNF-YA7; OsHAP2A		Os05g0213500	OsPYL/RCAR5; OsPYL5; OsPYL11
	Os03g0230300	OsSRO1c; BOC1		Os06g0598800	WSL1
	Os04g0541700	Oshox22
盐Salt	Os03g0348900	OsSRFP1; SDEL2	盐Salt	Os02g0121300	OsCYP2; LRT2
	Os04g0652400	OsSULTR3; 3;lpa		Os03g0272300	OsSDIR1
	Os03g0329900	OsPHR1		Os07g0129200	OsPR1a; OsSCP
	Os07g0187700	OsPHF1		Os05g0437700	EDT1; OsbZIP40
	Os02g0678200	OsSPX-MFS2; OsPSS2		Os08g0557000	OsPIMT1
	Os02g0325600	NIGT1		Os06g0693700	OsSIDP366
	Os01g0755700	NBIP1		Os04g0676700	OsMYB6
	Os10g0545700	OsACR2.1		Os01g0612700	OsLOL2; OsLOL5
	Os01g0869900	OsSAPK4; OSPDK		Os01g0948400	OsP5CR
	Os03g0719900	OsPTR8; OsNPF8.5		Os03g0319300	OsCam1-1
	Os09g0434500	OsBIERF1		Os05g0584200	OsLEA5

基于多平台基因表达数据的水稻干旱和盐胁迫相关基因预测

Prediction of drought and salt stress-related genes in rice based on multi-platform gene expression data

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胁迫 Stress	Hub基因集 Hub gene set	基因数目 Number of genes	数据集 Data set	平均精度 Average accuracy (%)
干旱Drought	D_affy_P	220	D_affy	100
	D_affy_MIC	104	D_affy	100
	D_rnaseq_P	738	D_rnaseq	90.0
	D_rnaseq_MIC	1634	D_rnaseq	91.0
	D_meta_hub	1936	D_affy	100
	D_meta_hub	1936	D_rnaseq	96.0
盐Salt	S_affy_P	470	S_affy	100
	S_affy_MIC	293	S_affy	100
	S_rnaseq_P	684	S_rnaseq	81.0
	S_rnaseq_MIC	331	S_rnaseq	84.6
	S_meta_hub	1504	S_affy	100
	S_meta_hub	1504	S_rnaseq	84.6

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