缺钾胁迫下谷子转录组分析及相关基因挖掘

doi:10.13304/j.nykjdb.2023.0900

中国农业科技导报 ›› 2024, Vol. 26 ›› Issue (6): 30-44.DOI: 10.13304/j.nykjdb.2023.0900

缺钾胁迫下谷子转录组分析及相关基因挖掘

鲁一薇¹(), 夏雪岩¹(), 赵宇¹, 崔纪菡¹, 刘猛¹, 黄玫红¹, 褚程¹, 刘建军², 李顺国¹()

^1.河北省农林科学院谷子研究所，国家谷子改良中心，农业农村部特色杂粮遗传改良与利用重点实验室（部省共建），河北省杂粮研究实验室，石家庄 050035
^2.河北省杂粮产业技术研究院，河北邯郸 056000

收稿日期:2023-12-07 接受日期:2024-03-04 出版日期:2024-06-15 发布日期:2024-06-12
通讯作者: 夏雪岩,李顺国
作者简介:鲁一薇E-mail： 1604425941@qq.com
夏雪岩E-mail： xyxia7808@126.com第一联系人：鲁一薇和夏雪岩为共同第一作者。
基金资助:
国家谷子高粱产业技术体系建设项目(CARS-06-14.5-A23);河北省农林科学院基本科研业务费包干制项目(HBNKY-BGZ-02);河北省现代农业产业技术体系建设项目(HBCT2024070101)

Transcriptome Profiling and Gene Mining of Millet Response to Potassium Deficiency Stress

Yiwei LU¹(), Xueyan XIA¹(), Yu ZHAO¹, Jihan CUI¹, Meng LIU¹, Meihong HUANG¹, Cheng CHU¹, Jianjun LIU², Shunguo LI¹()

^1.National Millet Improvement Center, Key Laboratory for Genetic Improvement and Utilization of Special Miscellaneous Grains of the Ministry of Agriculture and Rural Affairs (Jointly Established by the Ministry and Province), Hebei Miscellaneous Grain Research Laboratory, Institute of Millet, Hebei Academy of Agricultural and Forestry Sciences, Shijiazhuang 050035, China
^2.Hebei Industry and Technology Academy of Coarse Cereals, Hebei Handan 056000, China

Received:2023-12-07 Accepted:2024-03-04 Online:2024-06-15 Published:2024-06-12
Contact: Xueyan XIA,Shunguo LI

摘要/Abstract

摘要：

钾是作物生长不可缺少的元素，挖掘提高作物钾吸收能力的基因，增强作物对缺钾的耐受性具有重要的意义。以‘冀谷45’为供试材料，幼苗长至6叶期时进行缺钾胁迫7 d，通过农艺指标及转录组分析挖掘缺钾胁迫的相关基因。结果表明，缺钾胁迫影响谷子的生长发育，株高、叶宽、叶长、茎粗、苗期地上部干重及叶绿素含量低于CK，而根长、根表面积显著增加。转录组分析显示，缺钾胁迫后217个基因表达上调，38个基因表达下调。GO功能富集发现，差异表达基因主要富集于核糖体的结构成分、有机氮化合物代谢过程、蛋白质代谢过程、细胞蛋白质代谢过程、无膜细胞器等类别。KEGG功能富集结果表明，差异表达基因主要富集于核糖体、植物激素信号转导、谷胱甘肽代谢等19个通路。进而筛选出35个与激素表达显著相关的基因，19个氧化应激相关差异表达基因，10个转录因子家族及11个与转导信号相关的基因，1个与氨基酸及核苷酸的糖代谢相关的基因。候选基因Seita.9G193900属于CYP45084A亚家族，调控S型木质素合成的关键酶；Seita.5G365500属于WRKY家族，调控抗病响应基因PAD3等基因的表达；Seita.3G216900是RNA聚合酶Ⅱ转录亚基37E相关介质——HSPA1s辅助蛋白；Seita.4G286000为几丁内质内切酶基因CHIB，调控糖的合成与酵解。以上结果初步揭示了谷子缺钾胁迫相关基因的转录调控，为谷子耐缺钾相关基因的克隆与功能验证奠定基础。

关键词: 谷子, 缺钾, 差异表达基因, 转录组

Abstract:

Potassium is an essential element for crop growth. It is of great significance to explore genes that improve crop potassium absorption capacity and enhance crop tolerance to potassium deficiency. In this study， ‘Jigu 45’ was used as material and its seedlings were subjected to potassium deficiency stress for 7 d at the 6-leaf stage. By analyzing agronomic indicators and transcriptome， the relevant genes and expression regulation of potassium deficiency stress were analyzed and explored. The results showed that potassium deficiency stress affected the growth and development of foxtail millet plants： the plant height， leaf width， leaf length， stem width， aboveground dry weight and chlorophyll content were lower then those of CK， while the root length and root surface area significantly increased. Transcriptome analysis showed that 217 genes were upregulated and 38 genes were downregulated after potassium deficiency stress. GO functional enrichment revealed that differentially expressed genes were mainly enriched in the structural components of ribosomes， organic nitrogen compound metabolism processes， protein metabolism processes， cellular protein metabolism processes， and membraneless organelles etc. The KEGG functional enrichment results determined that differentially expressed genes were mainly enriched in 19 pathways， including ribosomes， plant hormone signal transduction and glutathione metabolism etc. Subsequently， 35 genes significantly correlated with hormone expression， 19 differentially expressed genes related to oxidative stress， 10 transcription factor families， and 11 genes related to transduction signals were screened， as well as 1 gene related to sugar metabolism of amino acids and nucleotides. The candidate gene Seita.9G193900 belonged to the CYP45084A subfamily and was a key enzyme regulating S-type lignin synthesis； Seita.5G36500 belonged to WRKY family， which regulated the expression of disease response genes such as PAD3； Seita.3G216900 was an RNA polymeraseⅡtranscription subunit 37E related mediator， HSPA1s helper protein； Seita.4G286000 was CHB gene which coded chitin endonuclease that regulated sugar synthesis and fermentation. Above results revealed the transcriptional regulation of genes related to potassium deficiency stress in foxtail millet， and laid a foundation for the cloning and functional verification of genes related to potassium tolerance in foxtail millet.

Key words: millet, potassium deficiency, differentially expressed genes, transcriptome

中图分类号:

S515

鲁一薇, 夏雪岩, 赵宇, 崔纪菡, 刘猛, 黄玫红, 褚程, 刘建军, 李顺国. 缺钾胁迫下谷子转录组分析及相关基因挖掘[J]. 中国农业科技导报, 2024, 26(6): 30-44.

Yiwei LU, Xueyan XIA, Yu ZHAO, Jihan CUI, Meng LIU, Meihong HUANG, Cheng CHU, Jianjun LIU, Shunguo LI. Transcriptome Profiling and Gene Mining of Millet Response to Potassium Deficiency Stress[J]. Journal of Agricultural Science and Technology, 2024, 26(6): 30-44.

图/表 16

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成分Component	对照 Control	缺钾 Potassium deficiency
1.0 mol·L^-1 Ca（NO₃）₂·4H₂O	5	5
1.0 mol·L^-1 KNO₃	5	—
1 mol·L^-1 NaH₂PO₄	—	1
1.0 mol·L^-1 NaNO₃	—	5
1.0 mol·L^-1 MgSO₄·7H₂O	2	2
1.0 mol·L^-1 KH₂PO₄	1	—
0.5% Fe-EDTA	1	1
微量元素母液 Trace element mother liquor	1	1
ddH₂O	985	985
总体积 Total volume	1 000	1 000

成分Component	对照 Control	缺钾 Potassium deficiency
1.0 mol·L^-1 Ca（NO₃）₂·4H₂O	5	5
1.0 mol·L^-1 KNO₃	5	—
1 mol·L^-1 NaH₂PO₄	—	1
1.0 mol·L^-1 NaNO₃	—	5
1.0 mol·L^-1 MgSO₄·7H₂O	2	2
1.0 mol·L^-1 KH₂PO₄	1	—
0.5% Fe-EDTA	1	1
微量元素母液 Trace element mother liquor	1	1
ddH₂O	985	985
总体积 Total volume	1 000	1 000

引物名称 Primer name	引物序列 Primer sequence （5'-3 '）
Seita.9G193900-F2	ATGTTTGGCGGGACGGAGAC
Seita.9G193900-R2	CCGATGGCCCAGACGTTGA
Seita.5G299700-F	GCGGGAGGACTTCGTGATGC
Seita.5G299700-R	CGGTGACGGGAATGCTGGA
Seita.5G365500-F1	CGGCCCTCCTCACCTCCAGTAT
Seita.5G365500-R1	CGCGTCCATGCCTTGTTGC
Seita.3G216900-F	ACCCGTCCGTGCAGAGTGA
Seita.3G216900-R	GGCAGTGGGCTCGTTGATT
Seita.7G222200-F1	GGTGGTGGAGGTCTTCGTC
Seita.7G222200-R1	TGTTGCCACTGTGCTTGATG
Seita.4G286000-F2	GACGCCACCATCGCCTTCA
Seita.4G286000-R2	GTTGGACCCGTAGCTGACCC
Seita.2G285600-F	TGCACGTCCACTGCCACATC
Seita.2G285600-R	CACGAACGCCTTCAGAACCAC
SiActin-F	GGCAAACAGGGAGAAGATGA
SiActin-R	GAGGTTGTCGGTAAGGTCACG

引物名称 Primer name	引物序列 Primer sequence （5'-3 '）
Seita.9G193900-F2	ATGTTTGGCGGGACGGAGAC
Seita.9G193900-R2	CCGATGGCCCAGACGTTGA
Seita.5G299700-F	GCGGGAGGACTTCGTGATGC
Seita.5G299700-R	CGGTGACGGGAATGCTGGA
Seita.5G365500-F1	CGGCCCTCCTCACCTCCAGTAT
Seita.5G365500-R1	CGCGTCCATGCCTTGTTGC
Seita.3G216900-F	ACCCGTCCGTGCAGAGTGA
Seita.3G216900-R	GGCAGTGGGCTCGTTGATT
Seita.7G222200-F1	GGTGGTGGAGGTCTTCGTC
Seita.7G222200-R1	TGTTGCCACTGTGCTTGATG
Seita.4G286000-F2	GACGCCACCATCGCCTTCA
Seita.4G286000-R2	GTTGGACCCGTAGCTGACCC
Seita.2G285600-F	TGCACGTCCACTGCCACATC
Seita.2G285600-R	CACGAACGCCTTCAGAACCAC
SiActin-F	GGCAAACAGGGAGAAGATGA
SiActin-R	GAGGTTGTCGGTAAGGTCACG

性状 Trait	对照 Control	缺钾 Potassium deficiency
株高 Plant height /cm	12.43	9.36^*
倒一叶叶宽 Top 1 leaf width /cm	0.80	0.67
倒一叶叶长 Top 1 leaf length /cm	20.75	17.75^*
倒二叶叶宽 Top 2 leaf width /cm	0.80	0.67
倒二叶叶长 Top 2 leaf length/ cm	19.63	19.50
倒三叶叶宽 Top 3 leaf width/cm	0.80	0.60
倒三叶叶长 Top 3 leaf length /cm	18.05	16.55^*
茎宽 Stem width /cm	3.31	2.51
苗期地上部干重 Seedling stage aboveground dry weight/g	0.07	0.06
苗期地下部干重 Seedling stage underground dry weight /g	0.01	0.02

缺钾胁迫下谷子转录组分析及相关基因挖掘

Transcriptome Profiling and Gene Mining of Millet Response to Potassium Deficiency Stress

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参考文献 36

相关文章 15

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Metrics

样品 Sample		原始读数 Raw reads	过滤读数 Clean reads	碱基含量 GC content/%	多个映射 Multiple mapped	唯一映射 Uniquely mapped	映射比率 Mapping Ratio/%
对照 Control	CK-1	46 868 802	46 106 644	49.45	2 097 694	40 095 522	86.97
	CK-2	44 338 374	43 789 738	49.98	2 111 597	38 217 463	87.28
	CK-3	45 196 350	44 462 108	50.03	2 855 499	37 291 700	83.88
缺钾Potassium deficiency	QK-1	45 814 604	44 963 384	49.28	3 083 637	38 025 162	84.58
	QK-2	42 927 104	42 284 578	50.06	2 059 902	37 037 265	87.60
	QK-3	51 831 874	51 061 668	56.00	2 097 363	45 595 390	89.30

途径 Pathway	基因数量 Gene number
核糖体Ribosome	74
植物激素信号转导Plant hormone signal transduction	35
谷胱甘肽代谢Glutathione metabolism	28
信号通路-植物MAPK signaling pathway-plant MAPK	27
丙氨酸天冬氨酸和谷氨酸代谢Alanine， aspartate and glutamate metabolism	15
丝氨酸甘氨酸和苏氨酸代谢Glycine， serine and threonine metabolism	13
真核生物的核糖体合成Ribosome biogenesis in eukaryotes	13
精氨酸生物合成Arginine biosynthesis	10
苯丙氨酸、酪氨酸、色氨酸生物合成Phenylalanine， tyrosine and tryptophan biosynthesis	10
酪氨酸代谢Tyrosine metabolism	10
苯丙胺酸代谢Phenylalanine metabolism	9
异喹啉类生物碱的生物合成Isoquinoline alkaloid biosynthesis	8
氮代谢Nitrogen metabolism	8
玉米素生物合成Zeatin biosynthesis	8
泛醌和其他萜醌的合成Ubiquinone and other terpenoid-quinone biosynthesis	8
脂肪酸的降解Fatty acid degradation	8
精氨酸和脯氨酸代谢Arginine and proline metabolism	8
莨菪烷哌啶和吡啶碱的生物合成Tropane， piperidine and pyridine alkaloid biosynthesis	7
β-氨基丙酸代谢β-alanine metabolism	7

激素种类 Hormone kind		基因名称 Gene name	log₂（FC）	描述 Description
赤霉素 Gibberellin	Seita.3G246300	GID1B	1.11	赤霉素受体 Gibberellin receptor
生长素 Auxin	Seita.7G230900	AUX	2.20	生长素诱导的蛋白 Auxin induced like-protein
	Seita.1G252000	AUX	2.21	生长素调节的相关蛋白 Auxin-regulated protein-related
	Seita.4G241400	ARF	7.17	生长素反应蛋白 Auxin responsive protein （auxin_inducible）
	Seita.1G345400	ARF	2.36	生长素反应蛋白 Auxin responsive protein （auxin_inducible）
	Seita.1G333600	ARF	1.59	生长素反应蛋白 Auxin responsive protein （auxin_inducible）
	Seita.9G194600	ARF	1.93	生长素反应蛋白 Auxin responsive protein （auxin_inducible）
	Seita.2G375300	GH3.2	4.78	吲哚-3-乙酸酰胺合成酶 Indole-3-acetic acid-amido synthetase
	Seita.1G075600	ARF	2.50	生长素反应蛋白 Auxin responsive protein （auxin_inducible）
	Seita.7G182000	AUX	2.95	生长素调节的相关蛋白Auxin-regulated protein-related

基因识别码 Gene identification code	差异倍数 log₂ （FC）	描述 Description
Seita.5G155100	3.76	过氧化物酶Peroxidase
Seita.7G180100	2.09	细胞色相关B561 Cytochrome B561-related
Seita.9G173500	5.14	细胞色素P450 85A1相关Cytochrome P450 85A1-related
Seita.5G064800	2.68	细胞色素P450家族成员Cytochrome P450 family member
Seita.4G077800	1.93	相关的NADH氧化还原酶NADH oxidoreductase-related
Seita.3G298400	4.71	细胞色素P450 CYP2亚家族Cytochrome P450 CYP2 subfamily
Seita.9G545600	3.39	细胞色素P450 CYP4/CYP19/CYP26亚家族Cytochrome P450 CYP4/CYP19/CYP26 subfamilies
Seita.9G342600	2.54	氧化还原酶Oxidoreductase
Seita.3G260700	3.27	细胞色素P450 CYP4/CYP19/CYP26亚家族Cytochrome P450 CYP4/CYP19/CYP26 subfamilies
Seita.9G081400	4.02	过氧化物酶/乳过氧化物酶Peroxidase / Lactoperoxidase
Seita.5G235200	1.23	细胞色素P450家族成员Cytochrome P450 family member
Seita.2G004800	2.65	过氧化物酶35相关Peroxidase 35-related
Seita.9G199900	1.09	细胞色素B561相关Cytochrome B561-related
Seita.1G117500	3.12	过氧化氢酶Catalase
Seita.1G042800	2.82	细胞色素P450家族成员Cytochrome P450 family member
Seita.3G226600	1.85	氧化还原酶Oxidoreductase
Seita.9G193900	4.77	细胞色素P450 84A1相关Cytochrome P450 84A1-related
Seita.5G010100	1.87	细胞色素P450 CYP2亚家族Cytochrome P450 CYP2 subfamily
Seita.9G444200	1.69	L-抗坏血酸过氧化物酶L-ascorbate peroxidase

转录因子基因 Transcription factor gene	基因识别码 Gene identification code	差异倍数 log₂ （FC）
ERF	Seita.5G280700	2.19
WRKY	Seita.2G433600	4.98
	Seita.5G365500	3.31
NAC	Seita.3G225400	4.91
	Seita.5G391200	1.15

基因识别码 Gene identification code	差异倍数 log₂（FC）	描述 Description
Seita.3G216900	1.87	RNA聚合酶Ⅱ转录亚基37E相关介质 Mediator of RNA polymeraseⅡtranacription subunit 37E-realated
Seita.9G536800	-2.03	Armadillo重复含驱动蛋白样蛋白2 Armadillo repeat-containing kinesin-like protein 2
Seita.2G212100	-1.79	SF71-磷脂酶D SF71-phospholipase D
Seita.1G047800	1.07	SF245-ADP-核糖基化因子GTPASE激活蛋白AGD12相关 SF245-ADP-ribosylation factor GTPASE-activating protein AGD12-related
Seita.2G044300	1.66	小核核糖核蛋白相关蛋白B和N Small nuclear ribonucleoprotein-associated protein B and N
Seita.3G065200	1.49	小核核糖核蛋白SM D2 Small nuclear ribonucleoprotein SM D2
Seita.5G376100	2.21	RNA聚合酶Ⅱ转录亚基37E相关介质 Mediator of RNA polymeraseⅡtranacription subunit 37E-realated
Seita.9G367700	1.05	SF9-小核核糖核蛋白 SF9-Small nuclear ribonucleoprotein
Seita.2G402800	1.37	RNA结合蛋白RBM8/Tsunagi（RRM超家族） RNA-binding protein RBM8/Tsunagi （RRM superfamily）
Seita.5G092600	2.88	SF141&小分子热休克蛋白HSP20家族 SF141-Small heat-shock protein HSP20 family
Seita.J011800	1.27	SF17-BCR相关蛋白 SF17-BCR-associated protein，BAP
Seita.4G286000	6.66	SF34-碱性内切酶 B SF34-basic endochitinase B

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