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作物学报 ›› 2016, Vol. 42 ›› Issue (04): 513-524.doi: 10.3724/SP.J.1006.2016.00513

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

人工合成甘蓝型油菜及其亲本的甲基化变异模式分析

谢涛,戎浩,蒋金金*,孔月琴,冉丽萍,吴健,王幼平   

  1. 扬州大学生物科学与技术学院, 江苏扬州 225009
  • 收稿日期:2015-08-27 修回日期:2016-01-11 出版日期:2016-04-12 网络出版日期:2016-01-25
  • 通讯作者: 蒋金金, E-mail: jjjiang@yzu.edu.cn, Tel: 0514-87997303
  • 基金资助:

    本项目由国家重点基础研究计划(计划973)项目(2015CB150201), 中国和江苏省博士后基金(2014M561719, 2015T80591, 1401078B)资助。

Analysis of DNA Methylation Patterns in Resynthesized Brassica napus and Diploid Parents

XIE Tao,RONG Hao,JIANG Jin-Jin*,KONG Yue-Qin,RAN Li-Ping,WU Jian,WANG You-Ping   

  1. College of Bioscience and Biotechnology, YangzhouUniversity, Yangzhou 225009, China
  • Received:2015-08-27 Revised:2016-01-11 Published:2016-04-12 Published online:2016-01-25
  • Contact: 蒋金金, E-mail: jjjiang@yzu.edu.cn, Tel: 0514-87997303
  • Supported by:

    This study was supported by the National Key Basic Research Program of China (2015CB150201), Chinaand Jiangsu Postdoctoral Program (2014M561719, 2015T80591, 1401078B).

摘要:

甘蓝型油菜作为多倍体起源和发生的历史较短, 遗传背景较为狭窄, 人工合成甘蓝型油菜可作为植物多倍化研究的优选模型, 本文以人工合成的甘蓝型油菜为材料, 通过HPLC分析发现白菜型油菜和甘蓝的甲基化率分别为8.33%和15.88%, 2个杂种株系的全基因组甲基化水平介于双亲之间(分别为10.29%和12.83%)。MSAP分析发现杂种F1代及其亲本的甲基化水平存在明显差异(白菜型油菜<杂种F1<甘蓝), 杂种F1代的甲基化变异(23.71%)中来自A、C基因组的变异分别占6.60%和10.16%。MSAP差异性条带的序列分析发现多倍化过程中与甲基化变化相关的基因参与了多种生物学过程, 且差异甲基化基因在人工合成甘蓝型油菜及其亲本间的表达差异与甲基化修饰模式是一致的。本研究为了解甘蓝型油菜多倍化过程中发生的表观变异奠定了基础。

关键词: 甘蓝型油菜, 人工合成种, 多倍化, 甲基化

Abstract:

The genetic background of Brassica napus, one of the most important oil crops in China, is relatively narrow due to the short history of its polyploid origin. Resynthesized B. napus provides an optimal model for researches on plant polyploidization. In the present study, we compared the DNA methylation levels in synthesized B. napus (F1 generation) and diploid parents using high-performance liquid chromatography (HPLC) and DNA methylation-sensitive amplification polymorphism (MSAP) analysis. HPLC analysis indicated methylation rates of 8.33% and 15.88% in B. rapa and B. oleracea, respectively. While the methylation rate of two hybrids was 10.29% and 12.83%, which werein-between of the parents’values. MSAP analysis proved the different methylation levels in F1 generation and diploids, with the lowest and highest methylation levels identified in B. rapa and B. oleracea, respectively.Variance of the DNA methylation in F1was 23.71%, among which 6.60% and 10.16% were inherited from A and C genome, respectively. Sequence analysis of MSAP polymorphic fragments indicated genes involved in multiple molecular functions were changed during polyploidization. Expression analysis of these genes agreed to the corresponding methylation changes. This study provides preliminary basis for understanding epigenetic variations during B. napus polyploidization.  

Key words: Bassica napus, Resynthesized species, Polyploidization, DNA methylation

[1] Solti D E, Visger C J, Soltis P S. The polyploidyrevolution then…and now: Stebbins revised. Am J Bot, 2014, 101: 1057–1078

[2] Blanc G, Wolfe K H. Widespread paleopolyploidy in model plant species inferred from age distributions of duplicate genes. Plant Cell, 2004, 16: 1667–1678

[3] Nagahararu U. Genome analysis in Brassica with special reference to the experimental formation of B. napus and peculiar mode of fertilization. Jpn J Bot, 1935, 7: 389–452

[4] Li H T, Younas M, Wang X F, Li X M, Chen L, Zhao B, Chen X, Xu J S, Hou F, Hong B H, Liu G, Zhao H Y, Wu X L, Du H Z, Wu J S, Liu K D. Development of a core set of single-locus SSR markers for allotetraploid rapeseed (Brassica napus L.). Theor Appl Genet, 2013, 126: 937–947

[5] Rahman H. Breeding spring canola (Brassica napus L.) by the use of exotic germplasm. Can J Plant Sci, 2013, 93: 363–373

[6] Qian W, Meng J, Li M, Frauen M, Sass O, Noack J, Jung C. Introgression of genomic components from Chinese Brassica rapa contributes to widening the genetic diversity in rapeseed (B. napus L.), with emphasis on the evolution of Chinese rapeseed. Theor Appl Genet, 2006, 113: 49–54

[7] Mei J, Fu Y, Qian L, Xu X, Li J, Qian W. Effectively widening the gene pool of oilseed rape (Brassica napus L.) by using Chinese B. rapa in a ‘virtual allopolyploid’ approach. Plant Breed, 2011, 130: 333–337

[8] Gaeta R T, Pires J C, Iniguez-Luy F, Leon E, Osborn T C. Genomic changes in resynthesized Brassica napus and their effect on gene expression and phenotype. Plant Cell, 2007, 19: 3403–3417

[9] Jiang J, Shao Y, Du K, Ran L, Fang X, Wang Y. Use of digital gene expression to discriminate gene expression differences in early generations of resynthesized Brassica napus and its diploid progenitors. BMC Genom, 2013, 1: 72–82

[10] Karim M M, Siddika A, Tonu N N, Hossain D M, Meah M B, Kawanabe T, Fujimoto R, Okazaki K. Production of high yield short duration Brassica napus by interspecific hybridization between B. oleracea and B. rapa. Breed Sci, 2014, 63: 495–502

[11] Wen J, Tu J X, Li Z, Fu T D, Ma C Z, Shen J X. Improving ovary and embryo culture techniques for efficient resynthesis of Brassica napus from reciprocal crosses between yellow-seeded diploids B. rapa and B. oleracea. Euphytica, 2008, 162: 81–89

[12] Wendel JF. Genome evolution in polyploids. Plant Mol Biol, 2000, 42: 225–249

[13] Buggs R J, Chamala S, Wu W, Tate J A, Schnable P S, Soltis D E, Soltis P S, Barbazuk W B. Rapid, repeated, and clustered loss of duplicate genes in allopolyploid plant populations of independent origin.Curr Biol, 2012, 22: 248–252

[14] Feldman M, Liu B, Segal G, Abbo S, Levy A A, Vega J M. Rapid elimination of low-copy DNA sequences in polyploid wheat: a possible mechanism for differentiation of homoeologous chromosomes. Genetics, 1997, 147: 1381–1387

[15] Chen Z J. Genetic and epigenetic mechanisms for gene expression and phenotypic variation in plant polyploids. Annu Rev Plant Biol, 2007, 58: 377–406

[16] Udall J A, Quijada P A, Osborn T C. Detection of chromosomal rearrangements derived from homeologous recombination in four mapping populations of Brassica napus L. Genetics, 2005, 169: 967–979

[17] Doyle J J, Flagel L E, Paterson A H, Rapp R A, Soltis D E, Soltis P S, Wendel J F. Evolutionary genetics of genome merger and doubling in plants. Annu Rev Plant Biol, 2008, 42: 443–461

[18] Lukens L N, Pires J C, Leon E, Vogelzang R, Oslach L, Osborn T. Patterns of sequence loss and cytosine methylation within a population of newly resynthesized Brassica napus allopolyploids. Plant Physiol, 2006, 140: 336–348

[19] Comai L, Tyagi A P, Winter K, Holmes-Davis R, Reynolds S H, Stevens Y, Byers B. Phenotypic instability and rapid gene silencing in newly formed Arabidopsis allotetraploids. Plant Cell, 2000, 12: 1551–1568

[20] Lee H S, Chen Z J. Protein-coding genes are epigenetically regulated in Arabidopsis polyploids. Proc Natl Acad Sci USA, 2001, 98: 6753–6758

[21] Shaked H, Kashkush K, Ozkan H, Feldman M A, Levy A. Reproducible responses of the genome to wide hybridization and allopolyploidy in wheat. Plant Cell, 2001, 13: 1749–1759

[22] Xu Y H, Zhong L, Wu X M, Fang X P, Wang J B. Rapid alterations of gene expression and cytosine methylation in newly synthesized Brassica napus allopolyploids. Planta, 2009, 229: 471–483

[23] Doyle J J, Doyle J L. Isolation of plant DNA from fresh tissue. Focus, 1990, 12: 13–15

[24] Demeulemeester M A C, Stallen N Van, De Proft M P. Degree of DNA methylation in chicory (Cichorium intybus L.): in?uence of plant age and vernalization. Plant Sci, 1999, 142: 101–108

[25] Xu M L, Li X Q, Korban S S. DNA-methylation alterations and exchanges during in vitro cellular differentiation in rose (Rosa hybrida L.). Theor Appl Genet, 2004, 109: 899–910

[26] Kim J K, Samaranayake M, Pradhan S. Epigenetic mechanisms in mammals. Cell Mol Life Sci, 2009, 66: 596–612

[27] Meilinger D, Fellinger K, Bultlnann S, Rothbauer U, Bonapace I M, Klinkert W E F. Np95 interacts with de novo DNA methyltransferases, Dnmt3a and Dnmt3b, and mediates epigenetic silencing of the viral CMV promoter in embryonic stem cells. EMBO Rep, 2009, 10: 1259–1264

[28] Hwang I S, Choi D S, Kim N H, Kim D S, Hwang B K. The pepper cysteine/histidine-rich DC1 domain protein CaDC1 binds both RNA and DNA and is required for plant cell death and defense response. New Phytol, 2014, 201: 518–530

[29] Tan X, Yan S Z, Tan R, Zhang Z Y, Wang Z, Chen J. Characterization and expression of a GDSL-Like lipase gene from Brassica napus in Nicotiana benthamiana. Protein J, 2014, 33: 18–23

[30] Gao Y, Zhao Y, Li T T, Liu Y, Ren C X, Wang M L. Molecular cloning and expression analysis of an F-box protein gene responsive to plant hormones in Brassica napus. Mol Biol Rep, 2010, 37: 1037–1044

[31] Lou P, Wu J, Cheng F, Cressman L G, Wang X W, McClung C R. Preferential retention of circadian clock genes during diploidization following whole genome triplication in Brassica rapa. Plant Cell, 2012, 24: 2415–2426

[32] Ferry N, Jouanin L, Ceci L R, Mulligan E A, Emami K, Gatehouses J A, Gatehouse A M R. Impact of oilseed rape expressing the insecticidal serine protease inhibitor, mustard trypsin inhibitor-2 on the beneficial predator Pterostichus madidus. Mol Ecol, 2005, 14: 337–349

[33] Chao Y, Yang Q, Kang J, Zhang T, Sun Y. Expression of the alfalfa FRIGIDA-like gene, MsFRI-L delays flowering time in transgenic Arabidopsis thaliana. Mol Biol Rep, 2013, 40: 2083–2090

[34] Baumert A, Milkowski C, Schmidt J, Nimtz M, Wray V, Strack D. Formation of a complex pattern of sinapate esters in Brassica napus seeds, catalyzed by enzymes of a serine carboxypeptidase-like acyltransferase family? Phytochemistry, 2005, 66: 1334–1345

[35] Cervera M T, Ruiz-Garcia L, Martinez-Zapater J. Analysis of DNA methylation in Arabidopsis thaliana based on methylation-sensitive AFLP markers. Mol Genet Genom, 2002, 268: 543–552

[36] Madlung A, Masuelli R W, Watson B, Reynolds S H, Davison J, Comai L. Remodeling of DNA methylation and phenotypic and transcriptional changes in synthetic Arabidopsis allotetraploids. Plant Physiol, 2002, 129: 733–746

[37] Xiong L Z, Xu C G, Maroof M A S, Zhang Q F. Patterns of cytosine methylation in an elite rice hybrid and its parental lines, detected by a methylation-sensitive amplification polymorphism technique. Mol Gen Genet, 1999, 261: 439–446

[38] Cai Y F, Xiang F N, Zhi D Y, Liu H, Xia G M. Genotyping of somatic hybrids between Festuca arundinacea Schred and Triticum aestivum L. Plant Cell Rep, 2007, 26: 1809–1819

[39] Salmon A, Ainouche M L, Wendel J F. Genetic and epigenetic consequences of recent hybridization and polyploidy in Spartina (Poaceae). Mol Ecol, 2005, 14: 1163–1175

[40] Gaeta R T, Pires J C, Iniguez-Luy F, Leon E, Osborn T C. Genomic changes in resynthesized Brassica napus and their effect on gene expression and phenotype. Plant Cell, 2007, 19: 3403–3417

[41] Birchler J A. Heterosis: The genetic basis of hybrid vigour. Nature Plants, 2015,15020, DOI: 10.1038/NPLANTS.2015.20

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