东亚特有珍稀蕨类植物岩穴蕨(碗蕨科)高通量转录组测序及分析

doi:10.17520/biods.2016231

生物多样性 ›› 2016, Vol. 24 ›› Issue (12): 1325-1334. DOI: 10.17520/biods.2016231

东亚特有珍稀蕨类植物岩穴蕨(碗蕨科)高通量转录组测序及分析

刘莉^1,³, 舒江平^1,³, 韦宏金¹, 张锐¹, 沈慧^1,^*(), 严岳鸿^1,²

1 上海辰山植物园, 中国科学院上海辰山植物科学研究中心, 上海 201602
2 华东野生濒危资源植物保育中心, 上海 201602
3 上海师范大学生命与环境科学学院, 上海 200234

收稿日期:2016-08-23 接受日期:2016-11-09 出版日期:2016-12-20 发布日期:2017-01-10
通讯作者: 沈慧
作者简介:* 通讯作者 E-mail: shenhui@sibs.ac.cn
基金资助:
上海市绿化和市容管理局科学技术项目(G142433)和中国科学院战略生物资源科技支撑体系运行专项(ZSZY-001)

De novo transcriptome analysis of the rare fern Monachosorum maximowiczii (Dennstaedtiaceae) endemic to East Asia

Li Liu^1,³, Jiangping Shu^1,³, Hongjin Wei¹, Rui Zhang¹, Hui Shen^1,^*(), Yuehong Yan^1,²

1 Shanghai Chenshan Botanical Garden, Shanghai Chenshan Plant Science Research Center, Chinese Academy of Sciences, Shanghai 201602
2 National Conservation Center for Endangered Useful Plants in East China, Shanghai 201602
3 College of Life and Environmental Sciences, Shanghai Normal University, Shanghai 200234

Received:2016-08-23 Accepted:2016-11-09 Online:2016-12-20 Published:2017-01-10
Contact: Hui Shen

摘要/Abstract

摘要：

岩穴蕨(Monachosorum maximowiczii)隶属于碗蕨科稀子蕨属, 是东亚中高海拔地区所特有的珍稀濒危植物。为了在分子水平对该物种有进一步的认识, 本文首次利用二代高通量测序技术(RNA-seq)对岩穴蕨进行了转录组测序分析。通过Illumina Hiseq 2500测序平台, 共计获得4.95 Gb原始数据(raw data), 经过滤后得到4.83 Gb有效数据(clean reads), 并进行从头组装得到了101,448条unigene。其中, 54,106条unigene预测到完整的开放阅读框。我们利用目前已知的51个植物基因组数据, 对岩穴蕨的unigene进行了详尽的功能注释, 并通过GO、COG、KEGG注释进一步了解了这些编码基因的作用方式、特征以及所参与的代谢通路。同时, 转录因子分析结果也为岩穴蕨的环境适应机制研究提供了初步线索。

关键词: 蕨类植物, 转录组学, 高通量测序, 生物多样性, 适应性

Abstract

Monachosorum maximowiczii, which belongs to the family Dennstaedtiaceae, is a unique fern rarely distributed in the middle-high altitudinal areas in East Asia. In order to identify the molecular basis of this species at the transcriptome level, a second generation sequencing technology (RNA-seq) was employed here for the first time. Using Illumina Hiseq 2500 platform, a total of 4.95 Gb raw data were obtained. After filtration, 4.83 Gb clean reads were kept and 101,448 unigene were generated using de novo assembly. Among them, 54,106 unigene were predicted to have a complete open reading frame. The functional annotation of the unigene obtained here was carried out using the Phytozome v11.0 databases. The gene function pattern and participated pathway were further explored based on GO, COG and KEGG annotation. Meanwhile, transcription factors prediction and analysis will provide preliminary insights into mechanisms of Monachosorum maximowiczii for environmental adaptation.

Key words: ferns, transcriptomics, high-throughput sequencing, biodiversity, adaptation

刘莉, 舒江平, 韦宏金, 张锐, 沈慧, 严岳鸿 (2016) 东亚特有珍稀蕨类植物岩穴蕨(碗蕨科)高通量转录组测序及分析. 生物多样性, 24, 1325-1334. DOI: 10.17520/biods.2016231.

Li Liu, Jiangping Shu, Hongjin Wei, Rui Zhang, Hui Shen, Yuehong Yan (2016) De novo transcriptome analysis of the rare fern Monachosorum maximowiczii (Dennstaedtiaceae) endemic to East Asia. Biodiversity Science, 24, 1325-1334. DOI: 10.17520/biods.2016231.

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链接本文: https://www.biodiversity-science.net/CN/10.17520/biods.2016231

https://www.biodiversity-science.net/CN/Y2016/V24/I12/1325

图/表 9

参考文献 27

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物种名 Species	测序部位 Description	测序平台 Platform	数据总量 Total size (Mb)	单基因簇或转录本数目 No. of unigene or transcript	文献 References
欧洲蕨 Pteridium aquilinum	配子体 Gametophyte	Roche 454	254	56,256	Der et al, 2011
水蕨 Ceratopteris richardii	孢子 Spore	Roche 454	266	15,730	Bushart et al, 2013
鸟巢蕨 Asplenium nidus	叶片 Leaf	Illumina HiSeq 2000	5,910	42,907	贾新平等, 2014
海金沙 Lygodium japonicum	原叶体、营养叶、孢子体、根状茎 Pro- thalli, trophophylls, sporophylls, rhizomes	Roche 454 Illumina HiSeq 2000	28 268	18,999 381,814	Aya et al, 2015
巨木贼 Equisetum giganteum	茎、叶、孢子囊 Stem, leaf, strobili	Illumina RNA-Seq	8,800	34,282	Vannesta et al, 2015

物种名 Species	测序部位 Description	测序平台 Platform	数据总量 Total size (Mb)	单基因簇或转录本数目 No. of unigene or transcript	文献 References
欧洲蕨 Pteridium aquilinum	配子体 Gametophyte	Roche 454	254	56,256	Der et al, 2011
水蕨 Ceratopteris richardii	孢子 Spore	Roche 454	266	15,730	Bushart et al, 2013
鸟巢蕨 Asplenium nidus	叶片 Leaf	Illumina HiSeq 2000	5,910	42,907	贾新平等, 2014
海金沙 Lygodium japonicum	原叶体、营养叶、孢子体、根状茎 Pro- thalli, trophophylls, sporophylls, rhizomes	Roche 454 Illumina HiSeq 2000	28 268	18,999 381,814	Aya et al, 2015
巨木贼 Equisetum giganteum	茎、叶、孢子囊 Stem, leaf, strobili	Illumina RNA-Seq	8,800	34,282	Vannesta et al, 2015

	原始数据 Raw data	有效数据 Clean data
序列数目 Number of sequences	49,027,722	48,497,004
全长 Total length (Gb)	4.95	4.83
Q20值 Q20%	96.54%	98.61%
Q30值 Q30%	91.54%	93.58%
GC值 GC%	47.82%	47.71%

	原始数据 Raw data	有效数据 Clean data
序列数目 Number of sequences	49,027,722	48,497,004
全长 Total length (Gb)	4.95	4.83
Q20值 Q20%	96.54%	98.61%
Q30值 Q30%	91.54%	93.58%
GC值 GC%	47.82%	47.71%

	转录本 Transcript	单基因簇 Unigene
序列数目 Sequence number	107,197	101,448
平均长度 Mean length (bp)	955	900
最大长度 Max. length (bp)	14,804	14,804
最小长度 Min. length (bp)	201	201
N50值 N50	1,907	1,817
GC含量 GC content	0.44	0.44

东亚特有珍稀蕨类植物岩穴蕨(碗蕨科)高通量转录组测序及分析

De novo transcriptome analysis of the rare fern Monachosorum maximowiczii (Dennstaedtiaceae) endemic to East Asia

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参考文献 27

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三大类 Three categories	GO条目 GO term	Unigene 数目 No. of unigene
生物学过程 Biological process	生物过程调控 Regulation of biological process	1,368
	细胞组织部分或生物合成 Cellular component organization or biogenesis	561
	定位活性 Establishment of localization	1,559
	免疫系统进程 Immune system process	9
	单一的生物过程 Single-organism process	5,268
	定位 Localization	1,576
	细胞活动 Locomotion	8
	生物粘附 Biological adhesion	2
	刺激反应 Response to stimulus	994
	细胞杀伤 Cell killing	1
	繁殖进程 Reproductive process	24
	生物调节 Biological regulation	1,448
	代谢进程 Metabolic process	9,458
	信号传导 Signaling	393
	生长 Growth	16
	细胞进程 Cellular process	8,525
	发育进程 Developmental process	37
	有机体进程 Multi-organism process	32
	繁殖 Reproduction	12
	生物过程的负调控 Negative regulation of biological process	48
	多细胞进程 Multicellular organismal process	54
	生物过程的正调控 Positive regulation of biological process	35
细胞组分 Cellular component	细胞器部分 Organelle part	864
	细胞 Cell	2,598
	高分子复合物 Macromolecular complex	1,399
	膜结构 Membrane	2,329
	胞外区域 Extracellular region	21
	膜部分 Membrane part	1,361
	病毒体 Virion	1
	膜关闭内腔 Membrane-enclosed lumen	150
	细胞部分 Cell part	2,598
	细胞器 Organelle	1,859
	胞外基质部分 Extracellular region part	1
	胞外基质 Extracellular matrix	1
	病毒体部分 Virion part	1
分子功能 Molecular function	结合蛋白转录活性 Protein binding transcription factor activity	60
	鸟嘌呤核苷酸交换因子活性 Guanyl-nucleotide exchange factor activity	33
	抗氧化活性 Antioxidant activity	114
	受体活性 Receptor activity	105
	结构分子活性 Structural molecule activity	294
	分子转导活性 Molecular transducer activity	116
	酶调节活性 Enzyme regulator activity	118
	营养库活性 Nutrient reservoir activity	7
	金属伴侣蛋白活性 Metallochaperone activity	4
	催化活性 Catalytic activity	8,722
	转运活性 Transporter activity	936
	结合活性 Binding	9,666
	电子载体活性 Electron carrier activity	119
	核苷酸结合转录因子活性 Nucleic acid binding transcription factor activity	350

COG功能分类 COG function classifications	Unigene数目 No. of unigene	百分比 Percentage (%)
RNA加工与修饰 RNA processing and modification	576	4.76
染色质结构和活力 Chromatin structure and dynamics	168	1.39
能量生成与转换 Energy production and conversion	475	3.92
细胞周期控制、细胞分裂、染色体分区 Cell cycle control, cell division, chromosome partitioning	288	2.38
氨基酸转运与代谢 Amino acid transport and metabolism	318	2.63
核苷酸转运与代谢 Nucleotide transport and metabolism	181	1.49
碳水化合物转运与代谢 Carbohydrate transport and metabolism	546	4.51
辅酶转运与代谢 Coenzyme transport and metabolism	154	1.27
脂类转运与代谢 Lipid transport and metabolism	424	3.50
翻译、核糖体结构和生物发生 Translation, ribosomal structure and biogenesis	562	4.64
转录 Transcription	669	5.52
复制、重组和修复 Replication, recombination and repair	403	3.33
细胞壁膜生物合成 Cell wall membrane biogenesis	66	0.54
细胞运动 Cell motility	2	0.02
翻译后修饰、蛋白质折叠、分子伴侣 Posttranslational modification, protein turnover, chaperones	1,209	9.98
无机离子转运与代谢 Inorganic ion transport and metabolism	315	2.60
次生代谢物合成、转运与代谢 Secondary metabolites biosynthesis, transport and catabolism	441	3.64
一般功能预测 General function prediction only	1,380	11.39
未知功能 Function unknown	932	7.69
信号转导机制 Signal transduction mechanisms	1,933	15.96
细胞内转运、分泌和小泡运输 Intracellular trafficking, secretion, and vesicular transport	610	5.04
防卫机制 Defense mechanisms	114	0.94
胞外结构 Extracellular structures	56	0.46
核结构 Nuclear structure	32	0.26
细胞骨架 Cytoskeleton	259	2.14

类别 Categories	通路 Pathways	Unigene数目 No. of unigene	通路数 No. of pathways
代谢途径 Metabolism etabolism	碳水化合物代谢 Carbohydrate metabolism	1,511	15
	脂类物质代谢 Lipid metabolism	813	14
	辅助因子和维生素代谢 Metabolism of cofactors and vitamins	371	11
	能量代谢 Energy metabolism	477	6
	核苷酸代谢 Nucleotide metabolism	446	2
	氨基酸代谢 Amino acid metabolism	831	14
	多酮类和萜类化合物的代谢 Metabolism of terpenoids and polyketides	224	8
	其他次生代谢产物的生物合成 Biosynthesis of other secondary metabolites	314	8
	其他氨基酸代谢 Metabolism of other amino acids	270	5
	多糖生物合成与代谢 Glycan biosynthesis and metabolism	166	7
	全球与总体图 Global and overview maps 小计 Sub-total	4,956 10,379 (73.41%)	6 96 (75%)
遗传信息加工 Genetic information processing	折叠、分类和降解 Folding, sorting and degradation	769	7
	翻译 Translation	876	5
	转录 Transcription	413	3
	复制与修复 Replication and repair 小计 Sub-total	493 2,551 (18.04%)	6 21 (16.41%)
环境信息处理 Environmental information processing	信号转导 Signal transduction 膜运输 Membrane transport 小计 Sub-total	350 36 386 (2.73%)	2 1 3 (2.34%)
细胞过程 Cellular processes	运输与代谢 Transport and catabolism 小计 Sub-total	506 506 (3.58%)	4 4 (3.13%)
生物系统 Organismal systems	环境适应性 Environmental adaptation 小计 Sub-total	252 252 (1.78%)	2 2 (1.56%)
人类疾病 Human diseases	耐药性: 抗菌 Drug resistance: Antimicrobial	7	1
人类疾病 Human diseases	内分泌与代谢性疾病 Endocrine and metabolic diseases 小计 Sub-total	57 64 (0.45%)	1 2 (1.56%)

编码 ID	代谢通路名称 Pathway	unigene数目 No. of unigene	百分比 Percentage (%)
ko01100	代谢通路 Metabolic pathways	2,618	23.03
ko01110	次生代谢产物的生物合成 Biosynthesis of secondary metabolites	1,418	12.47
ko01200	碳代谢 Carbon metabolism	406	3.57
ko01230	氨基酸生物合成 Biosynthesis of amino acids	330	2.90
ko03040	剪接体 Spliceosome	297	2.61
ko03010	核糖体 Ribosome	292	2.57
ko00500	淀粉与蔗糖代谢 Starch and sucrose metabolism	263	2.31
ko04075	植物激素信号转导 Plant hormone signal transduction	255	2.24
ko00230	嘌呤代谢 Purine metabolism	243	2.14
ko04141	内质网中的蛋白质加工 Protein processing in endoplasmic reticulum	230	2.02

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