研究报告

水稻基因组上一个Ds切离及其双位点插入行为的分子鉴定

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  • 1. 苏州大学基础医学与生物科学学院, 苏州 215123;
    2. 苏州市农业科学院作物研究室, 苏州215155
赵丁丁, 硕士研究生, 专业方向:植物发育生物学。Tel: 0512-65882833

收稿日期: 2014-02-10

  网络出版日期: 2014-12-20

基金资助

苏州市应用基础研究计划项目(编号:SYN201204 )资助

Molecular identification of one event of Ds excision and re-insertion at two loci in rice genome

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  • 1. School of Biology & Basic Medical Sciences, Soochow University, Suzhou 215123, China;
    2. Crop Science Research Laboratory, Suzhou Academy of Agricultural Sciences, Suzhou 215155, China

Received date: 2014-02-10

  Online published: 2014-12-20

摘要

玉米转座元件Ac/Ds是hAT转座子家族的成员, 导入水稻基因组后具有转座活性, 尽管转座机制还不完全清楚, 但它们通常经保守的非复制型“剪切-粘贴”过程转座。研究表明, 在Ac编码的转座酶作用下, Ds从原位点切离后常优先重新插入到连锁位点。文章利用TAIL-PCR技术从水稻一个Ds插入突变体及其回复突变体中分离Ds侧翼序列, 结合生物信息学分析方法, 对Ds在突变体上插入位点、回复突变体内切离足迹和重新插入位点进行了分子鉴定。结果显示, 突变体中Ds从3号染色体切离后, 在原插入位点残留了8 bp足迹序列(CATCATGA), 引起Ds标记基因外显子和内含子数目增加, 从而影响基因结构。切离后的Ds重新插入回复突变体第2和第6号染色体上, 分别编码烟草胺氨基转移酶和衰老相关蛋白的2个基因的编码区。因此, 典型的“剪切-粘贴”机制不能完全解释Ds的转座行为, Ds转座存在“剪切-复制-粘贴”的特点。

本文引用格式

赵丁丁, 乔中英, 程孝, 王建平, 焦翠翠, 孙丙耀 . 水稻基因组上一个Ds切离及其双位点插入行为的分子鉴定[J]. 遗传, 2014 , 36(12) : 1249 -1255 . DOI: 10.3724/SP.J.1005.2014.1249

Abstract

The maize Ac/Ds transposable elements are members of the hAT transposon superfamily, and have stable transpositional activity in transgenic rice plants. Ac/Ds transposable elements are considered to transpose via a conservative non-replicative “cut and paste” model, though their transposition mechanism is not completely understood. Previous studies have shown that Ds preferentially transposes to genetically linked sites after being excised from its original site in the presence of Ac-transposase. In this study, genomic sequences flanking Ds insertions from a Ds-tagged rice mutant and its rever- tant were determined by TAIL-PCR. The Ds insertion site, the excision footprint and the re-insertion sites in the mutant were identified using bioinformatics tool. The results showed that Ds element excised from its original insertion site on chromosome 3 by leaving an 8 bp footprint (CATCATGA), which resulted in exon changes in tagged gene. After the excision, Ds element was re-inserted into the coding sequences of two genes on chromosome 2 and chromosome 6, which encode a nicotianamine aminotransferase and a senescence-associated protein, respectively. The transposition behavior of Ds element in this study could not be fully explained by the "cut and paste" mechanism, while it is likely to transpose in a "cut and copy and paste" way.

参考文献

[1] Fedoroff N, Wessler S, Shure M. Isolation of the trans-posable maize controlling elements Ac and Ds. Cell, 1983, 35(1): 235–242.
[2] Chin HG, Choe MS, Lee SH, Park SH, Koo JC, Kim NY, Lee JJ, Oh BG, Yi GH, Kim SC, Choi HC, Cho MJ, Han CD. Molecular analysis of rice plants harboring an Ac/Ds transposable element-mediated gene trapping system. Plant J, 1999, 19(5): 615–623.
[3] Murai N, Li ZJ, Kawagoe Y, Hayashimoto A. Transposition of the maize activator element in transgenic rice plants. Nucleic Acids Res, 1991, 19(3): 617–622.
[4] Greco R, Ouwerkerk PB, Taal AJ, Favalli C, Beguiristain T, Puigdomènech P, Colombo L, Hoge JHC, Pereira A. Early and multiple Ac transpositions in rice suitable for efficient insertional mutagenesis. Plant Mol Biol, 2001, 46(2): 215–227.
[5] Zhu ZG, Fu YP, Xiao H, Hu GC, Si HM, Yu YH, Sun ZX. Ac/Ds transposition activity in transgenic rice population and DNA flanking sequence of Ds insertion sites. Acta Botanica Sinica, 2003, 45(1): 102–107.
[6] Jin WZ, Wang SM, Xu M, Duan RJ, Wu P. Characterization of enhancer trap and gene trap harboring Ac/Ds transposon in transgenic rice. J Zhejiang Univ Sci, 2004, 5(4): 390–399.
[7] Kolesnik T, Szeverenyi I, Bachmann D, Kumar CS, Jiang S, Ramamoorthy R, Cai M, Ma ZG, Sundaresan V, Ra-machandran S. Establishing an efficient Ac/Ds tagging system in rice: large-scale analysis of Ds flanking se-quences. Plant J, 2004, 37(2): 301–314.
[8] 刘芳, 张向前, 张泽民, 陈兆贵, 朱海涛, 王江, 张景六, 张桂权. 水稻Ac/Ds系统的Ds转座行为. 科学通报, 2007, 52(14): 1649–1655.
[9] Bancroft I, Dean C. Transposition pattern of the maize element Ds in Arabidopsis thaliana. Genetics, 1993, 134(4): 1221–1229.
[10] Greco R, Ouwerkerk PBF, De Kam RJ, Sallaud C, Favalli C, Colombo L, Guiderdoni E, Meijer AH, Hoge JHC, Pereira A. Transpositional behaviour of an Ac/Ds system for reverse genetics in rice. Theor Appl Genet, 2003, 108(1): 10–24.
[11] Wang N, Long T, Yao W, Xiong L, Zhang Q, Wu C. Mutant resources for the functional analysis of the rice genome. Mol Plant, 2013, 6(3): 596–604.
[12] Fujino K, Sekiguchi H. Transposition behavior of nonau-tonomous a hAT superfamily transposon nDart in rice (Oryza sativa L.). Mol Genet Genomics, 2011, 286(2): 135–142.
[13] Park SJ, Piao HL, Xuan YH, Park SH, Je BI, Kim CM, Lee EJ, Park SH, Ryu B, Lee KH, Lee GH, Nam MH, Yeo US, Lee MC, Yun DW, Eun MY, Han CD. Analysis of intragenic Ds transpositions and excision events generating novel allelic variation in rice. Mol Cells, 2006, 21(2): 284–293.
[14] Ros F, Kunze R. Regulation of activator/dissociation transposition by replication and DNA methylation. Ge-netics, 2001, 157(4): 1723–1733.
[15] 孙丙耀, 谈建中, 陆小平, 曲春香, 万志刚, 顾福根. 水稻Ac×Ds后代基因组DNA中Ds侧翼序列的扩增及其Ds插入分析. 遗传, 2006, 28(12): 1555–1561.
[16] Kang TJ, Yang MS. Rapid and reliable extraction of ge-nomic DNA from various wild-type and transgenic plants. BMC Biotechnol, 2004, 4: 20.
[17] Sun BY, Piao HL, Park SH, Han CD. Selection of optimal primers for TAIL-PCR in identifying Ds flanking sequences from Ac/Ds insertion rice lines. Chin J Biotechnol, 2004, 20(6): 821–826.
[18] Engels WR, Johnson-Schlitz DM, Eggleston WB, Sved J. High-frequency P element loss in Drosophila is homolog dependent. Cell, 1990, 62(3): 515–525.
[19] Hirochika H. Contribution of the Tos17 retrotransposon to rice functional genomics. Curr Opin Plant Biol, 2001, 4(2): 118–122.
[20] Bai L, Singh M, Pitt L, Sweeney M, Brutnell TP. Generating novel allelic variation through Activator insertional mutagenesis in maize. Genetics, 2007, 175(3): 981–992.
[21] Baran G, Echt C, Bureau T, Wessler S. Molecular analysis of the maize wx-B3 allele indicates that precise excision of the transposable Ac element is rare. Genetics, 1992, 130(2): 377–384.
[22] Scott L, LaFoe D, Weil CF. Adjacent sequences influence DNA repair accompanying transposon excision in maize. Genetics, 1996, 142(1): 237–246.
[23] Huang
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