转录组学分析意大利蜜蜂脑部哺育行为相关基因

doi:10.3864/j.issn.0578-1752.2020.19.021

Abstract

Abstract:

【Objective】The nursing behavior of Apis mellifera ligustica plays an important role in maintaining colony stability and production of royal jelly (RJ). In this study, samples (3-day-old worker bees, 10-day-old nurses/foragers, 21-day-old nurses/foragers) were obtained by constructing artificial colony, which can eliminate the influence of age on nursing behavior. Moreover, genes closely related to the nursing behavior were strictly screened out in the brain of these samples, which could reveal the molecular network of brain regulation of nursing behavior.【Method】The 3-day-old worker bees, 10-day-old nurses and foragers, 21-day-old nurses and foragers were obtained by constructing artificial bee colony. And then the head of different group’s worker bees was dissected to obtain brain tissues of these samples. RNA-seq was used to analyze the transcriptome sequencing of the 5 groups (3-day-old worker bees, 10-day-old nurses, 10-day-old foragers, 21-day-old nurses, 21-day-old foragers) of brain samples. Differentially expressed genes (DEGs) which are closely related to the nursing behavior in the nurses’ brain were screened out. GO and KEGG enrichment analysis were carried out for these genes. qPCR was used to verify the expression patterns of 4 randomly selected DEGs.【Result】RNA-seq analysis screened out 32 DEGs that were closely related to the nursing behavior of nurses. These genes were significantly up-regulated in the brain of 10-day-old nurses than 3-day-old worker bees, 10-day-old foragers and 21-day-old foragers, and the expression level in the brain of 21-day-old nurses was significantly higher than that in 21-day-old foragers. GO enrichment analysis showed that the up-regulated DEGs were mainly involved in oxidoreductase activity, odor binding, transmembrane transport and other functional items. KEGG enrichment results showed that the up-regulated DEGs were mainly involved in protein metabolism (ribosome), energy metabolism (oxidative phosphorylation, carbon metabolism, the citrate cycle (TCA cycle), starch and sucrose metabolism, nitrogen metabolism and other glycan degradation pathway), signal transduction (Toll and Imd signaling pathways, phototransduction, sphingolipid metabolism), digestive function (lysosome). Of them, only sphingolipid metabolism and other glycan degradation pathway were notably enriched. qPCR results showed that the expression patterns of 3 up-regulated DEGs (LOC409709, LOC551813, LOC409708) and 1 down-regulated DEG (LOC551165) were consistent with the sequencing data.【Conclusion】To obtain gene expression profiles of brains from age-matched nurses and foragers, the brains of 5 groups (3-day-old worker bees, 10-day-old nurses, 10-day-old foragers, 21-day-old nurses, and 21-day-old foragers) were comprehensively analyzed using transcriptome sequencing, revealing that the 32 up-regulated DEGs were associated with brain nursing behavior. These genes were mainly involved in signal transduction and energy metabolism, which can affect the nursing behavior.

Key words: Apis mellifera ligustica, nurses, brain, nursing behavior, royal jelly, RNA-seq

GAO Yan,ZHU YaNan,LI QiuFang,SU SongKun,NIE HongYi. Transcriptomic Analysis of Genes Related to Nursing Behavior in the Brains of Apis mellifera ligustica[J].Scientia Agricultura Sinica, 2020, 53(19): 4092-4102.

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样品 Sample	原始读数 Number of raw reads	有效读数 Number of clean reads	有效匹配数（匹配率） Clean map (Map rate)	单一匹配数（匹配率）Unique map (Map rate)	多匹配数（匹配率）Multiple map (Map rate)	99.9%的碱基正确率99.9% base accuracy (%)
3 d_1	67515798	66708824	56766708 (85.10%)	55951632 (83.87%)	815076 (1.22%)	88.25
3 d_2	83920302	81832028	70899666 (86.64%)	69821133 (85.32%)	1078533 (1.32%)	90.59
3 d_3	81546254	80442702	69502321 (86.40%)	68471536 (85.12%)	1030785 (1.28%)	89.66
10 dN_1	57288572	56348822	49321933 (87.53%)	48015717 (85.21%)	1306216 (2.32%)	90.51
10 dN_2	83787622	82964416	63408429 (76.43%)	60124557 (72.47%)	3283872 (3.96%)	90.66
10 dN_3	72837334	71663646	56922446 (79.43%)	56028890 (78.18%)	893556 (1.25%)	91.00
10 dF_1	67610626	66671128	55057317 (82.58%)	54208585 (81.31%)	848732 (1.27%)	90.37
10 dF_2	81258562	80246018	63725456 (79.41%)	62620480 (78.04%)	1104976 (1.38%)	90.06
10 dF_3	66021926	65365544	52522944 (80.35%)	51744723 (79.16%)	778221 (1.19%)	91.00
21 dN_1	71882296	71020506	56181047 (79.11%)	54846870 (77.23%)	8223024 (13.18%)	92.19
21 dN_2	72018192	70803290	60295752 (85.16%)	58523770 (82.66%)	11236986 (15.51%)	92.33
21 dN_3	72458810	71460376	59684702 (83.52%)	57902404 (81.03%)	8742852 (13.63%)	92.50
21 dF_1	72168956	70890254	59385891 (83.77%)	58485427 (82.50%)	900464 (1.27%)	91.94
21 dF_2	69418794	68335020	55559809 (81.31%)	54623413 (79.93%)	936396 (1.37%)	88.98
21 dF_3	81670548	80692702	67432186 (83.57%)	66368993 (82.25%)	1063193 (1.32%)	91.86

基因数目 Gene number	10 dN vs 3 d显著性比较Significant comparison of 10 dN vs 3 d	10 dN vs 21 dF显著性比较Significant comparison of 10 dN vs 21 dF	10 dN vs 10 dF显著性比较Significant comparison of 10 dN vs 10 dF	21 dN vs 21 dF显著性比较Significant comparison of 21 dN vs 21 dF	10 dN vs 21 dN显著性比较 Significant comparison of 10 dN vs 21 dN
32	↑	↑	↑	↑	ns
6	↓	↓	↓	↓	ns
3	↑	↓	↓	↓	ns

基因登录号 Gene_ID	基因 Gene	基因描述 Gene description	平均FPKM值 Average FPKM value					log₂FC (10 dN vs 10 dF)	log₂FC (21 dN vs 21 dF)
基因登录号 Gene_ID	基因 Gene	基因描述 Gene description	3 d	10 dN	10 dF	21 dN	21 dF	log₂FC (10 dN vs 10 dF)	log₂FC (21 dN vs 21 dF)
551813	LOC551813	王浆主蛋白1 Major royal jelly protein 1	0.40	1564.78	2.27	510.71	9.64	9.44	5.71
725215	LOC725215	毒酸磷酸酶Acph-1样 Venom acid phosphatase Acph-1-like	0.31	29.97	0.66	23.80	0.73	5.52	5.01
411186	LOC411186	突触小泡糖蛋白2C Synaptic vesicle glycoprotein 2C	0.22	12.81	0.47	11.96	0.60	4.78	4.29
410149	LOC410149	羧肽酶Q样 Carboxypeptidase Q-like	1.59	31.09	1.82	26.29	2.81	4.10	3.21
409709	LOC409709	假定的葡萄糖基神经酰胺酶4 Putative glucosylceramidase 4	2.83	75.03	7.81	50.90	8.63	3.27	2.54
552478	Obp17	气味结合蛋白17 Odorant binding protein 17	1.77	6.43	0.73	4.05	0.80	3.14	2.33
551935	Obp21	气味结合蛋白21 Odorant binding protein 21	4.50	34.70	4.43	22.20	7.71	2.97	1.51
677673	Obp14	气味结合蛋白14 Odorant binding protein 14	1.00	7.70	1.01	3.32	0.69	2.92	2.24
409708	LOC409708	葡糖基神经酰胺酶样 Glucosylceramidase-like	13.84	132.92	22.42	113.84	27.93	2.57	2.01
726798	LOC726798	Sn1特异性二酰基甘油脂肪酶β Sn1-specific diacylglycerol lipase beta	0.53	2.67	0.49	3.46	0.85	2.45	2.01
408383	CYP6AQ1	细胞色素P450 6AQ1 Cytochrome P450 6AQ1	0.49	1.97	0.42	1.41	0.39	2.22	1.84

GO条目 GO term	基因 Gene
氧化还原酶活性 Oxidoreductase activity	CYP6AQ1/LOC726418/ LOC408734/LOC413590
气味结合 Odorant binding	Obp21/Obp14/Obp17
跨膜运输 Transmembrane transport	LOC411186/LOC725462
蛋白质水解 Proteolysis	LOC725215/LOC725154
翻译 Translation	LOC550715
脂质代谢 Lipid metabolism	LOC726798
DNA复制 DNA replication	LOC725238
细胞器组分Organelle part	LOC408444

Transcriptomic Analysis of Genes Related to Nursing Behavior in the Brains of Apis mellifera ligustica

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