Comparative Transcriptome Analysis of Different Developmental Stages of Flowers and Fruits in Loquat Under Low Temperature Stress

doi:10.16420/j.issn.0513-353x.2021- 0219

Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (9): 1680-1694.doi: 10.16420/j.issn.0513-353x.2021- 0219

• Research Papers • Previous Articles Next Articles

Comparative Transcriptome Analysis of Different Developmental Stages of Flowers and Fruits in Loquat Under Low Temperature Stress

XU Hongxia¹, ZHOU Huifen², LI Xiaoying¹, JIANG Luhua³, CHEN Junwei¹^,^*()

¹Institute of Horticulture,Zhejiang Academy of Agricultural Sciences,Hangzhou 310021,China
²Zhejaing Agricultural Technology Extension Center,Hangzhou 310020,China
³Fruit Tree Technology Extension Station of Chun’an Agricultural Bureau,Chun’an,Zhejiang 311700,China

Received:2021-05-24 Revised:2021-08-25 Online:2021-09-25 Published:2021-09-30
Contact: CHEN Junwei E-mail:chenjunwei@zaas.ac.cn

Abstract

Abstract:

The study compared the low temperature response mechanism of loquat（Eriobotrya japonica Lindl.）flowers and fruits at different developmental stages. Flower buds,fully open flowers and young fruits were used as the experimental materials,treated at-3 ℃ for 12 h,and samples without low temperature treatment were used as controls. The relevant physiological and biochemical indexes,as well as transcriptome were determined both in control and treated samples. Low temperature stress led to the destruction of cell membranes,the generation rate of superoxide anion,the content of malondialdehyde and proline,and the activity of antioxidant enzymes were dramatically increased. In general,the low temperature resistance ability was flower bud > flower > young fruit. Transcriptome sequencing analysis obtained a total of 6 987 differentially expressed genes（DEG）. A large number of metabolic pathways related to low temperature stress were found,including glycolysis/gluconeogenesis,glyoxylic acid and dicarboxylic acid metabolism and phosphoinositol metabolism pathways in carbohydrate metabolism,tryptophan metabolism and tyrosine metabolism pathways in amino acid metabolism,glycerol phospholipid metabolism,α-linolenic acid metabolism and glyceride metabolism pathways in ester metabolism,isoquinoline alkaloid biosynthesis pathway in secondary metabolism,as well as thiometabolism pathway in energy metabolism,were significantly enriched in all comparison groups of low temperature and control at three flower and fruit development stages,which indicated that these pathways were important metabolic pathways in response to low temperature stress in loquat. The phenylpropanoid biosynthesis pathway was significantly enriched in both flower bud and flower comparison groups,and more amino acid metabolism pathways were enriched in young fruit comparison group after low temperature treatment. In addition,53 AP2-EREBP genes,14 WRKY genes and 15 NAC genes were screened from the differentially expressed genes related to low temperature treatment,and 12 low temperature response-related transcription factor genes were analyzed by qRT-PCR,which further confirmed the accuracy of transcriptome data.

Key words: Eriobotrya japonica, flower, fruit, development stage, low temperature, transcriptome, differentially expressed gene, transcription factor

CLC Number:

S667.3

XU Hongxia, ZHOU Huifen, LI Xiaoying, JIANG Luhua, CHEN Junwei. Comparative Transcriptome Analysis of Different Developmental Stages of Flowers and Fruits in Loquat Under Low Temperature Stress[J]. Acta Horticulturae Sinica, 2021, 48(9): 1680-1694.

Figures/Tables 11

References 43

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基因名称 Gene name	基因 ID Gene ID	引物序列（5′-3′） Primer sequence
Ejactin	c158550.graph_c1	F：GGATTTGCTGGTGATGATGC;	R：CCGTGCTCAATGGGATACTT
ERF37	c106174.graph_c0	F：GCTGGGTGAGGAGATAACGG	R：AAGATGCGAAGACGGTGGTG
ERF73	c123742.graph_c0	F：GCTTTCCTGAGGCGGACGAT	R：AATTGAGCTTGGAGGCGATT
ERF35	c125424.graph_c0	F：GGGGAGAGCATAGCGATACA	R：GCTTACGTCATTTTCTGGGC
ERF105	c126492.graph_c0	F：TCGCTCGTACTTGTCACACC	R：GCTGACCATCAAAGGCAACT
RAV2	c126935.graph_c1	F：GAGGATGATCGGCTGGGAGT	R：AGCTGCTGCAAGGCACTACA
ERF60	c129543.graph_c0	F：TGGCCAATAAGAATCACCAA	R：CCAAAAAGGATCTGACGAGG
ERF33	c46214.graph_c0	F：GCCTGTTCTTTGATTTTGGT	R：TCATGAGCTGATCTTCCCTC
WRKY33	c142932.graph_c0	F：CCACTGAAAACCAATCGTAC	R：CTTAGACTTCTTATCGCAAC
NAC72	c51471.graph_c0	F：ATGTTGGGGTTGGATAAGAA	R：CGGAATAGAAGAAGCAGAAGAG
NAC74	c136256.graph_c0	F：AGTCCATAGTATGCGCCACC	R：TGACTGCGGTTTTCTTTTGA
NAC29	c130635.graph_c0	F：GAAACGGGAACATTGGTAGT	R：AATGAATGAGATGCTGGTGT
NAC91	c136368.graph_c0	F：ATCATCGCTAAATTCATTCC	R：ACTTTACCACCTCTACCAAC

基因名称 Gene name	基因 ID Gene ID	引物序列（5′-3′） Primer sequence
Ejactin	c158550.graph_c1	F：GGATTTGCTGGTGATGATGC;	R：CCGTGCTCAATGGGATACTT
ERF37	c106174.graph_c0	F：GCTGGGTGAGGAGATAACGG	R：AAGATGCGAAGACGGTGGTG
ERF73	c123742.graph_c0	F：GCTTTCCTGAGGCGGACGAT	R：AATTGAGCTTGGAGGCGATT
ERF35	c125424.graph_c0	F：GGGGAGAGCATAGCGATACA	R：GCTTACGTCATTTTCTGGGC
ERF105	c126492.graph_c0	F：TCGCTCGTACTTGTCACACC	R：GCTGACCATCAAAGGCAACT
RAV2	c126935.graph_c1	F：GAGGATGATCGGCTGGGAGT	R：AGCTGCTGCAAGGCACTACA
ERF60	c129543.graph_c0	F：TGGCCAATAAGAATCACCAA	R：CCAAAAAGGATCTGACGAGG
ERF33	c46214.graph_c0	F：GCCTGTTCTTTGATTTTGGT	R：TCATGAGCTGATCTTCCCTC
WRKY33	c142932.graph_c0	F：CCACTGAAAACCAATCGTAC	R：CTTAGACTTCTTATCGCAAC
NAC72	c51471.graph_c0	F：ATGTTGGGGTTGGATAAGAA	R：CGGAATAGAAGAAGCAGAAGAG
NAC74	c136256.graph_c0	F：AGTCCATAGTATGCGCCACC	R：TGACTGCGGTTTTCTTTTGA
NAC29	c130635.graph_c0	F：GAAACGGGAACATTGGTAGT	R：AATGAATGAGATGCTGGTGT
NAC91	c136368.graph_c0	F：ATCATCGCTAAATTCATTCC	R：ACTTTACCACCTCTACCAAC

数据库 Database	注释数量 Number of annotated unigene	比例/% Percentage of annotated unigene
Nr	38 093	97.80
Swiss-Prot	22 119	56.79
KEGG	14 813	38.03
eggNOG	35 024	89.92
KOG	20 989	53.89
GO	25 436	65.30
Pfam	27 156	69.72

数据库 Database	注释数量 Number of annotated unigene	比例/% Percentage of annotated unigene
Nr	38 093	97.80
Swiss-Prot	22 119	56.79
KEGG	14 813	38.03
eggNOG	35 024	89.92
KOG	20 989	53.89
GO	25 436	65.30
Pfam	27 156	69.72

GO分类 GO classification	GO序列号 GO accession	注释功能（词条） Term	差异基因数 DEGs number	-3 ℃ vs 25 ℃
GO分类 GO classification	GO序列号 GO accession	注释功能（词条） Term	差异基因数 DEGs number	花蕾 Flower bud	花 Flower	幼果 Young fruit
生物学过程 Biological process	GO:0008152	代谢过程Metabolic process	2 193	298	781	1 239
	GO:0009987	细胞过程Cellular process	2 127	248	661	1 075
	GO:0044699	单一生物过程Single-organism process	1 863	230	585	1 033
	GO:0051179	定位Localization	790	108	276	546
	GO:0065007	生物调控Biological regulation	616	142	270	427
	GO:0071840	细胞组分或生物合成 Cellular component organization or biogenesis	465	40	132	293
	GO:0050896	应激反应Response to stimulus	457	94	193	279
	GO:0023052	信号Signaling	122	24	47	77
	GO:0051704	多生物过程Multi-organism process	114	11	39	73
	GO:0032502	发育过程Developmental process	94	25	35	46
	GO:0040007	生长Growth	75	2	16	41
	GO:0022414	生殖过程Reproductive process	59	15	25	44
	GO:0000003	再生Reproduction	55	0	26	46
	GO:0032501	多细胞生物过程Multicellular organismal process	48	17	22	17
	GO:0022610	生物粘附Biological adhesion	17	0	3	6
	GO:0002376	免疫系统过程Immune system process	7	0	2	0
	GO:0048511	节律过程Rhythmic process	3	0	1	1
细胞组分 Cellular component	GO:0005623	细胞Cell	1 808	283	718	1 229
	GO:0044464	细胞部分Cell part	1 796	280	710	1 222
	GO:0043226	细胞器Organelle	1 260	201	485	858
	GO:0016020	膜Membrane	1 642	294	623	1 024
	GO:0044425	膜部分Membrane part	1 293	234	495	805
	GO:0044422	细胞器部分Organelle part	621	67	218	418
	GO:0032991	大分子复合物Macromolecular complex	552	42	208	361
	GO:0031974	膜封闭的管腔Membrane-enclosed lumen	160	11	40	112
	GO:0005576	细胞外区域Extracellular region	87	18	45	61
	GO:0030054	细胞连接Cell junction	15	4	6	5
	GO:0055044	共质体Symplast	14	4	5	5
	GO:0019012	病毒Virion	4	0	1	4
	GO:0044423	病毒部分Virion part	4	0	1	4
	GO:0009295	内核Nucleoid	11	0	5	6
	GO:0044421	细胞外区域Extracellular region part	28	3	21	26
分子功能	GO:0003824	催化活性Catalytic activity	1 949	308	725	1 222
Molecular	GO:0005488	联结Binding	1 670	321	688	1 024
function	GO:0005215	转运活动Transporter activity	402	53	144	277
	GO:0005198	结构分子活动Structural molecule activity	178	9	102	85
	GO:0001071	核酸联结转录因子活性 Nucleic acid binding transcription factor activity	109	40	64	68
	GO:0016209	抗氧化活性Antioxidant activity	40	10	21	21
	GO:0009055	电子载体活动Electron carrier activity	35	5	13	20
	GO:0060089	分子传感器活动Molecular transducer activity	36	5	7	8
	GO:0004871	信号传导活动Signal transducer activity	36	10	18	23
	GO:0000988	转录因子活性,蛋白质结合 Transcription factor activity,protein binding	16	4	3	12
	GO:0045735	营养库活性Nutrient reservoir activity	5	1	3	2
	GO:0031386	蛋白质标签Protein tag	5	0	0	2
	GO:0016530	金属伴侣活性Metallochaperone activity	1	1	1	1
	GO:0045182	翻译调控活性Translation regulator activity	2	0	0	2

Comparative Transcriptome Analysis of Different Developmental Stages of Flowers and Fruits in Loquat Under Low Temperature Stress

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Figures/Tables 11

References 43

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COG 分类 COG classification	分类名称 Classification name	-3 ℃ vs 25 ℃
COG 分类 COG classification	分类名称 Classification name	花蕾 Flower bud	花 Flower	幼果 Young fruit
A	RNA 加工和修饰RNA processing and modification	0	0	1
B	染色质结构与动态Chromatin structure and dynamics	0	0	1
C	能量产生与转换Energy production and conversion	41	81	132
D	细胞周期控制,细胞分裂,染色体分割 Cell cycle control,cell division,chromosome partitioning	7	7	13
E	氨基酸转运与代谢Amino acid transport and metabolism	27	72	184
F	核苷酸转运与代谢Nucleotide transport and metabolism	7	19	56
G	碳水化合物转运与代谢Carbohydrate transport and metabolism	120	150	274
H	辅酶转运与代谢Coenzyme transport and metabolism	26	36	73
I	脂质转运与代谢Lipid transport and metabolism	66	82	156
J	翻译,核糖体结构与生物合成Translation,ribosomal structure and biogenesis	33	89	156
K	转录Transcription	26	25	33
L	复制,重组与修复Replication,recombination and repair	15	16	35
M	细胞壁/膜/包膜生物合成Cell wall/membrane/envelope biogenesis	36	43	73
N	细胞运动Cell motility	2	1	4
O	翻译后修饰,蛋白质折叠,分子伴侣 Posttranslational modification,protein turnover,chaperones	59	71	141
P	无机离子转运与代谢Inorganic ion transport and metabolism	46	80	122
Q	次生代谢物生物合成,转运与代谢 Secondary metabolites biosynthesis,transport and catabolism	42	69	123
R	一般功能预测General function prediction only	85	126	261
S	功能未知Function unknown	29	40	48
T	信号转导机制Signal transduction mechanisms	67	70	101
U	细胞内运输,分泌和囊泡运输 Intracellular trafficking,secretion,and vesicular transport	0	1	8
V	防御机制Defense mechanisms	10	22	47
W	细胞外结构Extracellular structures	0	0	1
X	移动元件：原噬菌体,转座子Mobilome：prophages,transposons	7	9	7
Y	细胞核结构Nuclear structure	0	0	0
Z	细胞骨架Cytoskeleton	2	3	2

Unigene 编号 Unigene ID	转录组log₂值 RNA-seq log₂ FC			qRT-PCR相对表达量 qRT-PCR relative expression			表达 Express	Nr注释 Nr_ annotation	来源 Source
Unigene 编号 Unigene ID	花蕾 Flower bud	花 Flower	幼果 Young fruit	花蕾 Flower bud	花 Flower	幼果 Young fruit	表达 Express	Nr注释 Nr_ annotation	来源 Source
c106174.graph_c0	1.88	1.90	1.56	2.86 ± 0.32	3.45 ± 0.46	2.26 ± 0.34	上调Up	ERF37	枇杷Eriobotrya japonica
c123742.graph_c0	3.34	2.37	1.39	1.59 ± 0.23	4.78 ± 0.35	2.56 ± 0.21	上调Up	ERF73	苹果Malus × domestica
c125424.graph_c0	3.29	3.67	3.91	6.57 ± 0.58	4.23 ± 0.52	3.12 ± 0.29	上调Up	ERF35	枇杷Eriobotrya japonica
c126935.graph_c1	2.85	2.16	1.42	1.23 ± 0.15	1.85 ± 0.20	1.36 ± 0.16	上调Up	ERF105	苹果Malus × domestica
c129543.graph_c0	2.75	2.63	1.57	1.28 ± 0.12	2.35 ± 0.24	3.42 ± 0.21	上调Up	RAV2	枇杷Eriobotrya japonica
c46214.graph_c0	1.55	3.52	4.65	0.79 ± 0.12	5.48 ± 0.35	2.59 ± 0.38	上调Up	ERF60	苹果Malus × domestica
c126492.graph_c0	-1.90	-2.04	-2.54	-1.02 ± 0.11	-3.05± 0.36	-2.13± 0.22	下调Down	ERF33	苹果Malus × domestica
c142932.graph_c0	2.13	2.87	1.42	1.36 ± 0.21	1.25 ± 0.15	1.49 ± 0.23	上调Up	WRKY33	苹果Malus × domestica
c51471.graph_c0	4.92	4.59	4.00	6.89 ± 0.71	7.68 ± 0.68	6.21 ± 0.52	上调Up	NAC 72	苹果Malus × domestica
c130635.graph_c	1.92	2.95	1.78	2.06 ± 0.16	3.46 ± 0.42	2.76 ± 0.31	上调Up	NAC74	梨Pyrus bretschneideri
c136256.graph_c0	3.65	2.69	2.57	4.69 ± 0.53	3.76 ± 0.47	2.63 ± 0.18	上调Up	NAC29	苹果Malus × domestica
c136368.graph_c0	1.22	1.01	1.00	0.69 ± 0.10	1.38 ± 0.20	1.54 ± 0.23	上调Up	NAC91	苹果Malus × domestica

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