植物学报 ›› 2015, Vol. 50 ›› Issue (2): 217-226.doi: 10.3724/SP.J.1259.2015.00217

• 研究报告 • 上一篇    下一篇

香港红山茶个体内ITS多态性及物种鉴定的应用

范文1, 徐颖1, 许汀1, 徐晶1,2, 高继银3, 张文驹1,*   

  1. 1复旦大学生命科学学院生物多样性科学研究所, 生物多样性与生态工程教育部重点实验室, 上海 200433
    2中国科学院上海生命科学信息中心, 上海 200031
    3广东棕榈风景园林科学研究院, 中山 528416
  • 收稿日期:2014-03-14 接受日期:2014-05-09 出版日期:2015-03-01 发布日期:2015-04-10
  • 通讯作者: 张文驹
  • 作者简介:

    ? 共同第一作者

  • 基金资助:
    国家重点基础研究发展规划(No.2007CB411600)

Intragenomic Polymorphism of the Internal Transcribed Spacer Region of Ribosomal DNA in Camellia hongkongensis (Theaceae) and Species Identification

Wen Fan1, Ying Xu1, Ting Xu1, Jing Xu1, 2, Takahiro Yonezawa1, Jiyin Gao3, Wenju Zhang1, *   

  1. 1Ministry of Education Key Laboratory for Biodiversity Science and Ecological Engineering, Institute of Biodiversity Science, School of Life Sciences, Fudan University, Shanghai 200433, China
    2Shanghai Information Center for Life Sciences, Chinese Academy of Sciences, Shanghai 200031, China
    3Guangdong Palm Landscape Architecture Research Institute, Zhongshan 528416, China
  • Received:2014-03-14 Accepted:2014-05-09 Online:2015-03-01 Published:2015-04-10
  • Contact: Zhang Wenju
  • About author:

    ? These authors contributed equally to this paper

摘要:

核糖体DNA的内转录间隔区(ITS)一直被作为一种重要的分子标记, 却很难用于山茶物种中。通过对1个疑似香港红山茶(Camellia hongkongensis)的样本进行ITS区域的扩增、克隆和测序, 从中获得74种不同序列。研究结果表明, 其ITS区域具有高度的多态性, 其中76%的序列为假基因。系统发育分析显示, 超过半数的假基因源自同一祖先。这些假基因在经历多次基因重复后分化成至少5个谱系, 且每个谱系中的序列非常相似, 这表明一些假基因不但未被剔除, 反而通过快速复制事件幸存下来。由于山茶物种个体内ITS的高度多态, 使用这个区域区分山茶物种可能导致错误。然而, 通过比较香港红山茶中的1个种间特异性rDNA假基因, 确定该样本属于香港红山茶。

Abstract:

The internal transcribed spacer (ITS) region of ribosomal DNA has been selected as an important barcoding region in plants, but its use in Camellia species is difficult. In this study, we amplified, cloned and sequenced the ITS region of a Camellia plant from Vietnam that was similar to C. hongkongensis and obtained 74 sequences from an individual. The ITS region showed high polymorphism within the individual, and 76% of the sequences were pseudogenes. Phylogenetic analysis showed that more than half of the pseudogenes originated from a common ancestor. These older rDNA pseudogenes differentiated into at least 5 lineages after repeated gene duplication, and sequences in each lineage were similar to each other, which suggests that some pseudogenes had not been deleted but had recently undergone rapid duplication events. The high polymorphism of the ITS region within an individual in Camellia species would likely result in faulty identities when using this region as a barcode. However, by comparing the species-specific rDNA pseudogenes in C. hongkongensis, we identified the sample from Vietnam that belongs to C. hongkongensis.

表1

香港红山茶ITS序列特征"

Typea rDNA region Haptotypesb Genetic distance h±SD π Lengths
(bp)
GC contents ΔG (kcal∙mol-1)c
All ITS1-5.8S-ITS2 47 0.210 5±0.011 7 0.991±0.008 0.166 72 589-687 0.557
ITS1 0.297 4±0.023 0 0.958±0.015 0.242 21 221-291 0.550
5.8S 0.083 1±0.011 5 0.961±0.013 0.065 27 139-163 0.501
ITS2 0.240 1±0.020 1 0.962±0.012 0.171 55 198-233 0.610
F ITS1-5.8S-ITS2 10 0.010 3±0.001 9 0.978±0.054 0.009 74 674-687 0.686
ITS1 0.007 0±0.002 1 0.667±0.163 0.006 47 281-291 0.719
5.8S 0.008 4±0.003 6 0.667±0.163 0.008 32 163 0.543 -36.90, -47.42
ITS2 0.015 7±0.004 2 0.933±0.077 0.014 69 230-233 0.751
P ITS1-5.8S-ITS2 37 0.212 2±0.012 0 0.994±0.009 0.164 71 589-632 0.531
ITS1 0.283 5±0.023 0 0.964±0.015 0.225 14 221-258 0.515
5.8S 0.095 8±0.013 3 0.959±0.014 0.073 47 139-163 0.491
ITS2 0.248 3±0.020 9 0.953±0.016 0.176 47 198-223 0.581
Pa1d ITS1-5.8S-ITS2 5 0.003 6±0.001 6 1.000±0.126 0.003 58 614 0.523
ITS1 0.002 6±0.002 6 0.600±0.175 0.002 61 230 0.523
5.8S 0.004 9±0.003 5 0.700±0.218 0.004 91 163 0.461 -24.90, -36.00
ITS2 0.003 6±0.002 6 0.700±0.218 0.003 62 221 0.567
Pa2 ITS1-5.8S-ITS2 8 0.006 1±0.002 0 1.000±0.063 0.006 06 607 0.541
ITS1 0.005 8±0.003 0 0.857±0.108 0.005 77 223 0.535
5.8S 0.005 1±0.003 7 0.679±0.122 0.005 04 163 0.512 -30.22, -35.70
ITS2 0.007 2±0.003 7 0.750±0.139 0.007 11 221 0.568
Pa3 ITS1-5.8S-ITS2 5 0.005 8±0.002 1 1.000±0.126 0.005 77 589 0.529
ITS1 0.003 5±0.002 5 0.700±0.218 0.003 51 228 0.528
5.8S 0.009 9±0.005 0 0.900±0.161 0.009 82 163 0.490 -20.80, -23.62
ITS2 0.005 1±0.003 6 0.800±0.164 0.005 05 198 0.564
Pa4 ITS1-5.8S-ITS2 9 0.011 3±0.002 9 0.972±0.064 0.011 11 610-611 0.577
ITS1 0.006 7±0.003 6 0.833±0.098 0.006 70 224-225 0.584
5.8S 0.020 0±0.008 3 0.806±0.120 0.019 43 163 0.501 -23.10, -34.50
ITS2 0.009 6±0.004 6 0.694±0.147 0.009 47 223 0.624
Pb ITS1-5.8S-ITS2 9 0.151 4±0.009 5 1.000±0.052 0.121 31 590-632 0.582
ITS1 0.174 4±0.017 4 1.000±0.052 0.139 17 221-258 0.587
5.8S 0.088 8±0.013 5 0.917±0.092 0.076 94 139-163 0.499 -15.22, -42.12
ITS2 0.181 0±0.019 4 0.917±0.092 0.136 33 206-221 0.644

图1

疑似香港红山茶及外类群ITS序列的邻接树 总共53条香港红山茶序列, 其中47条来自本研究, 3条来自GenBank, 3条(C hon S256ppA, C hon S257L, C hon S257J)由邹稚华教授提供。外类群序列来自GenBank, 由14个山茶属物种组成(以种名的前3个字母指代)"

1 闵天禄 (2000). 世界山茶属的研究. 昆明: 云南科学技术出版社. pp. 4-308.
2 闵天禄, 张文驹 (1996). 山茶属植物的进化与分布. 云南植物研究 18, 1-13.
3 唐绍清, 施苏华, 钟杨, 王燕 (2004). 基于ITS序列探讨山茶属金花茶组的系统发育关系. 广西植物 24, 488-492.
4 徐颖, 徐晶, 高继银, 张文驹 (2011). 山茶属植物ITS的多态性——一个广泛逃离一致性进化的实例. 植物学报 46, 162-169.
5 杨俊波, 李洪涛, 杨世雄, 李德铢, 杨莹燕 (2006). 四个DNA片段在山茶属分子系统学研究中的应用. 云南植物研究 28, 108-114.
6 张宏达, 任善湘 (1998). 中国植物志(第49卷第3分册). 北京: 科学出版社. pp. 6-170.
7 Akaike H (1974). A new look at the statistical model identification.IEEE Trans Automat Contr 19, 716-723.
8 Álvarez I, Wendel JF (2003). Ribosomal ITS sequences and plant phylogenetic inference.Mol Phylogenet Evol 29, 417-434.
9 Bailey CD, Carr TG, Harris SA, Hughes CE (2003). Characterization of angiosperm nrDNA polymorphism, para- logy, and pseudogenes.Mol Phylogenet Evol 29, 435-455.
10 Baldwin BG, Sanderson MJ, Porter JM, Wojciechowski MF, Campbell CS, Donoghue MJ (1995). The ITS region of nuclear ribosomal DNA: a valuable source of evidence on angiosperm phylogeny.Ann Missouri Bot Gard 82, 247-277.
11 Buckler ES 4th, Holtsford TP (1996). Zea ribosomal repeat evolution and substitution patterns.Mol Biol Evol 13, 623-632.
12 Carranza S, Baguñà J, Riutort M (1999). Origin and evolution of paralogous rRNA gene clusters within the flatworm family Dugesiidae (Platyhelminthes, Tricladida).J Mol Evol 49, 250-259.
13 Copenhaver GP, Pikaard CS (1996). Two-dimensional RFLP analyses reveal megabase-sized clusters of rRNA gene variants in Arabidopsis thaliana, suggesting local spreading of variants as the mode for gene homogenization during concerted evolution.Plant J 9, 273-282.
14 Dover G (1982). Molecular drive: a cohesive mode of species evolution.Nature 299, 111-117.
15 Doyle JJ, Doyle JL (1987). A rapid DNA isolation procedure for small quantities of fresh leaf tissue.Phytochem Bull 19, 11-15.
16 Eickbush TH, Eickbush DG (2007). Finely orchestrated movements: evolution of the ribosomal RNA genes.Genetics 175, 477-485.
17 Ganley ARD, Kobayashi T (2007). Highly efficient concerted evolution in the ribosomal DNA repeats: total rDNA repeat variation revealed by whole-genome shotgun sequence data.Genome Res 17, 184-191.
18 Gu ZJ, Hua X (2003). Physical mapping of the 18S-26S rDNA by fluorescent in situ hybridization (FISH) in Camellia reticulata polyploid complex (Theaceae).Plant Sci 164, 279-285.
19 Hamada M, Sato K, Kiryu H, Mituyama T, Asai K (2009). Predictions of RNA secondary structure by combining homologous sequence information.Bioinformatics 25, i330-i338.
20 Hillis DM, Moritz C, Porter CA, Baker RJ (1991). Evidence for biased gene conversion in concerted evolution of ribosomal DNA.Science 251, 308-310.
21 Keller I, Chintauan-Marquier IC, Veltsos P, Nichols RA (2006). Ribosomal DNA in the grasshopper Podisma pedestris: escape from concerted evolution.Genetics 174, 863-874.
22 Kovarik A, Pires JC, Leitch AR, Lim KY, Sherwood AM, Matyasek R, Rocca J, Soltis DE, Soltis PS (2005). Rapid concerted evolution of nuclear ribosomal DNA in two Tragopogon allopolyploids of recent and recurrent origin.Genetics 169, 931-944.
23 Librado P, Rozas J (2009). DnaSP v5: a software for comprehensive analysis of DNA polymorphism data.Bioinformatics 25, 1451-1452.
24 Long EO, Dawid IB (1980). Repeated genes in eukaryotes.Annu Rev Biochem 49, 727-764.
25 Márquez LM, Miller DJ, MacKenzie JB, Van Oppen MJH (2003). Pseudogenes contribute to the extreme diversity of nuclear ribosomal DNA in the hard coral Acropora.Mol Biol Evol 20, 1077-1086.
26 Martin DP, Lemey P, Lott M, Moulton V, Posada D, Lefeuvre P (2010). RDP3: a flexible and fast computer program for analyzing recombination.Bioinformatics 26, 2462-2463.
27 Martin DP, Williamson C, Posada D (2005). RDP2: recombination detection and analysis from sequence align- ments.Bioinformatics 21, 260-262.
28 Mayol M, Rosselló JA (2001). Why nuclear ribosomal DNA spacers (ITS) tell different stories in Quercus.Mol Phylogenet Evol 19, 167-176.
29 Muir G, Fleming CC, Schlötterer C (2001). Three divergent rDNA clusters predate the species divergence in Quercus petraea (matt.) liebl. and Quercus robur L.Mol Biol Evol 18, 112-119.
30 Nylander JAA (2004). MrModeltest v2. Program distributed by the author. Evolutionary Biology Centre, Uppsala University.
31 Odorico DM, Miller DJ (1997). Variation in the ribosomal internal transcribed spacers and 5.8S rDNA among five species of Acropora (Cnidaria; Scleractinia): patterns of variation consistent with reticulate evolution.Mol Biol Evol 14, 465-473.
32 Padidam M, Sawyer S, Fauquet CM (1999). Possible emergence of new geminiviruses by frequent recombination.Virology 265, 218-225.
33 Parkin EJ, Butlin RK (2004). Within- and between- individual sequence variation among ITS1 copies in the meadow grasshopper Chorthippus parallelus indicates frequent intrachromosomal gene conversion.Mol Biol Evol 21, 1595-1601.
34 Parks CR (1990). Cross-compatibility studies in the genus Camellia. A presentation for the ICS congress at Maizura.Int Camellia J 22, 37-54.
35 Pedersen C, Linde-Laursen I (1994). Chromosomal locations of four minor rDNA loci and a marker microsatellite sequence in barley.Chromosome Res 2, 65-71.
36 Posada D, Crandall KA (2001). Evaluation of methods for detecting recombination from DNA sequences: computer simulations.Proc Natl Acad Sci USA 98, 13757-13762.
37 Prokopowich CD, Gregory TR, Crease TJ (2003). The correlation between rDNA copy number and genome size in eukaryotes.Genome 46, 48-50.
38 Rannala B, Yang ZH (1996). Probability distribution of molecular evolutionary trees: a new method of phylogenetic inference.J Mol Evol 43, 304-311.
39 Ronquist F, Huelsenbeck JP (2003). MrBayes 3: Bayesian phylogenetic inference under mixed models.Bioinformatics 19, 1572-1574.
40 Saitou N, Nei M (1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees.Mol Biol Evol 4, 406-425.
41 Schlötterer C, Tautz D (1994). Chromosomal homogeneity of Drosophila ribosomal DNA arrays suggests intrachromosomal exchanges drive concerted evolution.Curr Biol 4, 777-783.
42 Smith JM (1992). Analyzing the mosaic structure of genes.J Mol Evol 34, 126-129.
43 Swofford DL (2002). PAUP*: Phylogenetic Analysis Using Parsimony (*and Other Methods). Version 4. Sinauer Associates, Sunderland, MA.
44 Swofford DL, Berlocher SH (1987). Inferring evolutionary trees from gene frequency data under the principle of maximum parsimony.Syst Zool 36, 293-325.
45 Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011). MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods.Mol Biol Evol 28, 2731-2739.
46 Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997). The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.Nucleic Acids Res 25, 4876-4882.
47 Van Oppen MJH, Willis BL, Van Rheede T, Miller DJ (2002). Spawning times, reproductive compatibilities and genetic structuring in the Acropora aspera group: evidence for natural hybridization and semi-permeable species boundaries in corals.Mol Ecol 11, 1363-1376.
48 Vijayan K, Zhang WJ, Tsou CH (2009). Molecular taxo- nomy of Camellia (Theaceae) inferred from nrITS sequences.Am J Bot 96, 1348-1360.
49 Xiao LQ, Möller M, Zhu H (2010). High nrDNA ITS polymorphism in the ancient extant seed plant Cycas: incomplete concerted evolution and the origin of pseudogenes.Mol Phylogenet Evol 55, 168-177.
50 Xu JP, Zhang QQ, Xu XF, Wang ZG, Qi J (2009). Intragenomic variability and pseudogenes of ribosomal DNA in Stone flounder Kareius bicoloratus.Mol Phylogenet Evol 52, 157-166.
51 Young ND, Healy J (2003). GapCoder automates the use of indel characters in phylogenetic analysis.BMC Bioinformatics 4, 6.
52 Zheng XY, Cai DY, Yao LH, Teng YW (2008). Non-concerted ITS evolution, early origin and phylogenetic utility of ITS pseudogenes in Pyrus.Mol Phylogenet Evol 48, 892-903.
53 Zimmer EA, Martin SL, Beverley SM, Kan YW, Wilson AC (1980). Rapid duplication and loss of genes coding for the alpha chains of hemoglobin.Proc Natl Acad Sci USA 77, 2158-2162.
No related articles found!
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed   
[1] 杨礼锐 陈木楚. 植物抗逆性与光呼吸作用之间的关系[J]. 植物学报, 1991, 8(01): 43 -47 .
[2] 贺萍. 北京植物园展览温室植物的害虫调查及主要害虫的防治[J]. 植物学报, 1996, 13(02): 44 -47 .
[3] 崔凯荣 陈克明 王晓哲 王亚馥. 植物体细胞胚胎发生研究的某些现状[J]. 植物学报, 1993, 10(03): 14 -20 .
[4] 黄瑶 李朝銮 马诚 吴乃虎. 叶绿体DNA及其在植物系统学研究中的应用[J]. 植物学报, 1994, 11(02): 11 -25 .
[5] 王璞 赵秀琴. 浸提条件对小麦秸秆中化感物质检测结果的影响[J]. 植物学报, 2001, 18(06): 735 -738 .
[6] 云自厚 梁明霞 张存洁 谭志一. 用气相色谱测定植物样品中的微量细胞分裂素[J]. 植物学报, 1988, 5(01): 60 -63 .
[7] 何艳霞;王子成*. 拟南芥幼苗超低温保存后DNA甲基化的遗传变异[J]. 植物学报, 2009, 44(03): 317 -322 .
[8] 施怡婷, 杨淑华. 中国科学家在乙烯信号转导领域取得突破性进展[J]. 植物学报, 2016, 51(3): 287 -289 .
[9] 吕超群, 孙书存. 陆地生态系统碳密度格局研究概述[J]. 植物生态学报, 2004, 28(5): 692 -703 .
[10] 龙文兴, 丁易, 臧润国, 杨民, 陈少伟. 海南岛霸王岭热带云雾林雨季的环境特征[J]. 植物生态学报, 2011, 35(2): 137 -146 .