石栗叶绿体基因组研究

doi:10.11983/CBB22026

摘要/Abstract

摘要： 石栗(Aleurites moluccana)是大戟科石栗属的常绿阔叶乔木, 具有能源、药用和观赏价值。为填补石栗叶绿体基因组研究的空白, 通过二代高通量全基因组测序, 组装和注释了石栗叶绿体基因组, 并进行基因组特征和系统发育分析。结果显示, 石栗叶绿体基因组为典型的四段式结构, 总长度为163 298 bp, LSC、SSC及IR的长度分别为91 301、18 501和26 748 bp。石栗叶绿体基因组共有131个基因, 包括8个rRNA基因, 37个tRNA基因, 86个蛋白质编码基因。研究发现145个SSR位点, 检测到重复单元有单核苷酸、二核苷酸、三核苷酸和四核苷酸, 数目分别为80、53、10和2个。共线性分析结果表明, 石栗叶绿体基因组存在基因倒位和重排现象。利用最大似然法和贝叶斯法构建了系统发育树, 显示石栗与油桐(Vernicia fordii)和东京桐(Deutzianthus tonkinensis)亲缘关系较近, 并形成姐妹群。利用化石时间进行定年分析, 表明石栗属、油桐属和东京桐属的分化时间为25.94 Ma (95% HPD: 24.71-63.32 Ma)。该研究丰富了石栗基因组信息, 可为石栗种质资源的开发利用提供基础遗传数据, 同时为石栗属物种鉴定及系统发育研究提供参考。

关键词: 石栗, 叶绿体基因组, 系统发育

Abstract: Aleurites moluccana is an evergreen broad-leaved tree of the genus Aleurites in the family Euphorbiaceae, with energy, medicinal and ornamental values. To fill the gap in study of the chloroplast genome of A. moluccana, we assembled and annotated the chloroplast genome of A. moluccana by next-generation high-throughput whole genome sequencing, and performed genomic characterization and phylogenetic analysis. The results showed that the chloroplast genome of A. moluccana exhibited typical quadripartite and circular structures with a total length of 163 298 bp, the length of LSC, SSC, and IR was 91 301, 18 501, and 26 748 bp, respectively. It contains 131 genes, including 8 rRNA genes, 37 tRNA genes and 86 protein coding genes. A total of 145 SSR loci were found, with mononucleotide, dinucleotide, trinucleotide and tetranucleotide repeat units, and the numbers detected were 80, 53, 10, and 2, respectively. The results of collinearity analysis showed that the chloroplast genome of A. moluccana has the phenomenon of gene inversion and rearrangement. Phylogenetic trees were constructed using the maximum likelihood and Bayesian methods. It was found that A. moluccana was closely related to Vernicia fordii and Deutzianthus tonkinensis, and formed a sister group. The results of the dating analysis using fossil time showed that the differentiation time of the Aleurites, Vernicia and Deutzianthus was 25.94 Ma (95% HPD: 24.71-63.32 Ma). This study enriched the genomic information of A. moluccana and provided basic genetic data for the development and utilization of A. moluccana germplasm resources, as well as a reference for species identification and phylogenetic study of the Aleurites.

Key words: Aleurites moluccana, chloroplast genome, phylogeny

包金波, 丁志杰, 苗浩宇, 李雪丽, 任书贤, 焦若岩, 李浩, 邓茜茜, 李英姿, 田新民. 石栗叶绿体基因组研究. 植物学报, 2023, 58(2): 248-260.

Jinbo Bao, Zhijie Ding, Haoyu Miao, Xueli Li, Shuxian Ren, Ruoyan Jiao, Hao Li, Qianqian Deng, Yingzi Li, Xinmin Tian. Analysis of Chloroplast Genomes of Aleurites moluccana. Chinese Bulletin of Botany, 2023, 58(2): 248-260.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB22026

https://www.chinbullbotany.com/CN/Y2023/V58/I2/248

图/表 12

图1 石栗自然种群 (A) 花序; (B) 枝条; (C) 幼枝和幼叶; (D) 果实

Figure 1 Natural population of Aleurites moluccana (A) Inflorescence; (B) Branches; (C) Young branches and leaves; (D) Fruit

图2 石栗叶绿体基因组图谱标注在大环外部的基因按照顺时针方向转录, 标注在大环内部的基因按照逆时针方向转录。不同颜色代表基因功能不同。内环阴影部分代表石栗叶绿体基因组的GC组成。LSC: 大单拷贝区; SSC: 小单拷贝区; IRA: 反向重复区A; IRB: 反向重复区B。* 含有内含子的基因。

Figure 2 The chloroplast genome map of Aleurites moluccana Genes on the outside of the large circle are transcribed clockwise and those on the inside are transcribed counterclockwise. The genes are color-coded based on their function. The dashed area on the inside represents the GC composition of the A. moluccana chloroplast genome. LSC: Large single-copy region; SSC: Small single-copy region; IRA: Inverted repeat region A; IRB: Inverted repeat region B. * for genes containing introns.

表1 石栗叶绿体基因组功能基因注释

Table 1 Annotation of functional genes in the chloroplast genome of Aleurites moluccana

Categories of genes	Group of genes	Name of genes
Genes for photosynthesis	Subunits of photosystem I	psaA, psaB, psaC, psaI, psaJ
	Subunits of photosystem II	psbA, psbB, psbC, psbD, psbE, psbF, psbH, psbI, psbJ, psbK, psbL, psbM, psbN, psbT, psbZ
	Subunits of ATP synthase	atpA, atpB, atpE, atpF, atpH, atpI
	Subunits of cytochrome	petA, petB, petD, petG, petL, petN
	ATP-dependent protease subunits P gene	clpP
	Large subunits of Rubisco	rbcL
	Subunits of NADH dehydrogenase	ndhA, ndhB, ndhC, ndhD, ndhE, ndhF, ndhG, ndhH, ndhI, ndhJ, ndhK
Self replication	Small subunit of ribosome	rps2, rps3, rps4, rps7, rps8, rps11, rps12, rps14, rps15, rps16, rps18, rps19
	Large subunit of ribosome	rpl2, rpl14, rpl16, rpl20, rpl2, rpl23, rpl32, rpl33, rpl36, rpl22
	DNA dependent RNA polymerase	rpoA, rpoB, rpoC1, rpoC2
	Ribosomal RNA genes	rrn5, rrn4.5, rrn16, rrn23
	Transfer RNA genes	trnN-GUU, trnR-ACG, trnH-GUG, trnL-CAU, trnA-UGC, trnL-GAU, trnV-GAC, trnL- CAA, trnL-CAU, trnP-UGG, trnW-CCA, trnM-CAU, trnV-UAC, trnF-GAA, trnL-UAA, trnT-UGU, trnS-GGA, trnfM-CAU, trnG-GCC, trnS-UGA, trnT-GGU, trnE-UUC, trnY-GUA, trnD-GUC, trnC-GCA, trnR-UCU, trnS-GCU, trnQ-UUG, trnK-UUU, trnL-CAA, trnV-GAC, trnL-GAU, trnR-UGC, trnL-UAG, trnR-ACG, trnN-GUU
Other genes	Maturase	matK
	Envelop membrane protein	cemA
	Translation initiation factor IF-1	infA
	C-type cytochrome synthesis gene	ccsA
Unknown function	Conserved open reading frames	ycf1, ycf2, ycf3, ycf4, ycf15

表2 石栗叶绿体基因组简单重复序列(SSR)信息

Table 2 Information of simple sequence repeat (SSR) identified in the chloroplast genome of Aleurites moluccana

SSR repeat type (number of copies)	SSR repeat sequence	Number of copies
SSR repeat type (number of copies)	SSR repeat sequence	5	6	7	8	9	10	11	12	13	14	15	16-30	Total
Mononucleotide (80)	A/T						24	17	7	14		3	3	68
Mononucleotide (80)	C/G						7	3		2				12
Dinucleotide (53)	AT/TA/AC/CA/AG/GA		18	7		6	4	6		2			2	45
Dinucleotide (53)	CG/GC		4	2		1	1							8
Trinucleotide (10)	CGG/AAG	2	2	1										5
	TGT/TTA	1		2										3
	ATT/AAT	1	1											2
Tetranucleotide (2)	TACA/TGGT	1			1									2

图3 石栗叶绿体基因组重复序列长度及类型 (A) 重复序列长度; (B) 重复序列类型

Figure 3 The length and type of repeat sequences in the chloroplast genome of Aleurites moluccana (A) The length of repeat sequences; (B) The type of repeat sequences

图4 石栗叶绿体基因组不同氨基酸的使用频率

Figure 4 Usage frequency of different amino acid in the chloroplast genome of Aleurites moluccana

表3 石栗叶绿体基因组密码子使用偏好性

Table 3 Codon usage bias in the chloroplast genome of Aleurites moluccana

Amino acid	Codon	No. of codon	RSCU	Amino acid	Codon	No. of codon	RSCU
Ala	GCU	431	1.20	Cys	UGU	698	1.20
	GCC	306	0.86	Cys	UGC	468	0.80
	GCA	427	1.19	Glu	GAA	1416	1.40
	GCG	267	0.75	Glu	GAG	611	0.60
Asp	GAU	1073	1.41	Gly	GGU	534	0.93
Asp	GAC	448	0.59		GGC	382	0.67
Phe	UUU	2457	1.26		GGA	803	1.40
Phe	UUC	1452	0.74		GGG	572	1.00
His	CAU	874	1.36	Ile	AUU	1927	1.19
His	CAC	412	0.64		AUC	1184	0.73
Lys	AAA	2578	1.38		AUA	1728	1.07
Lys	AAG	1162	0.62	Leu	UUA	1299	1.46
Trp	UGG	748	1.00		UUG	1125	1.26
Asn	AAU	2122	1.43		CUU	1021	1.15
Asn	AAC	843	0.57		CUC	577	0.65
Arg	CGU	341	0.58		CUA	844	0.95
	CGC	246	0.42		CUG	442	0.50
	CGA	590	1.00	Pro	CCU	568	1.02
	CGG	412	0.70		CCC	556	1.00
	AGA	1253	2.12		CCA	699	1.25
	AGG	702	1.19		CCG	408	0.73
Thr	ACU	683	1.18	Met	AUG	887	1.00
Thr	ACC	550	0.95	Gln	CAA	1041	1.38
	ACA	696	1.20		CAG	463	0.62
	ACG	383	0.66	Ser	UCU	1070	1.36
Val	GUU	737	1.34		UCC	840	1.07
	GUC	402	0.73		UCA	1002	1.27
	GUA	667	1.21		UCG	565	0.72
	GUG	393	0.71		AGU	720	0.92
Tyr	UAU	1842	1.43		AGC	521	0.66
Tyr	UAC	737	0.57

图5 石栗叶绿体基因组共线性分析 LSC、SSC、IRA和IRB同图2。

Figure 5 Collinear analysis of Aleurites moluccana chloroplast genome LSC, SSC, IRA, and IRB are the same as in Figure 2.

图6 石栗叶绿体基因组反向重复区的收缩与扩张 LSC、SSC、IRA和IRB同图2。JLB: LSC与IRB的边界; JSB: SSC与IRB的边界; JSA: SSC与IRA的边界; JLA: LSC与IRA的边界

Figure 6 Contraction and expansion of inverted repeat region in the chloroplast genome of Aleurites moluccana LSC, SSC, IRA, and IRB are the same as in Figure 2. JLB: Boundary between LSC and IRB; JSB: Boundary between SSC and IRB; JSA: Boundary between SSC and IRA; JLA: Boundary between LSC and IRA

图7 基于蛋白编码序列构建的石栗与其它19个物种贝叶斯系统发育树分支上的数值为后验概率。

Figure 7 Bayesian phylogenetic tree of Aleurites moluccana and other 19 species based on protein coding sequence The values on the branch are posteriori probability.

图8 基于蛋白编码序列构建的石栗与其它19个物种最大似然(ML)系统发育树分支上的数值为后验概率。

Figure 8 Maximum likelihood (ML) phylogenetic tree of Aleurites moluccana and other 19 species based on protein coding sequence The values on the branch are posteriori probability.

图9 基于宽松分子钟模型构建的石栗与其它19个物种系统发育时间树分支上的数值为分化时间(单位: 百万年前)。

Figure 9 Phylogenetic dating tree of Aleurites moluccana and other 19 species based on relax molecular clock model The values on the branch are the differentiation time (unit: million years ago).

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