GmGBP1干涉转基因大豆种质创制及其对下游基因可变剪接的调控
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摘要
GmGBP1基因与大豆光周期开花时间相关,本试验以GmGBP1基因为对象,利用大豆子叶节转化法进行植物组织培养,得到GmGBP1干涉(GmGBP1-i) T2转基因大豆材料并进行分子检测。RNA-seq转录组测序获得可变剪接数据,进一步利用RT-PCR方法对WT与GmGBP1-i大豆叶片测序结果中的可变剪接基因进行验证,共检测到3个基因发生可变剪接,表明GmGBP1干涉表达量降低引起了下游基因可变剪接。本研究为大豆GmGBP1的基因功能分析及解析大豆成花诱导机理奠定了基础。
GmGBP1 is a gene related to photoperiod flowering time of soybean.In this experiment,GmGBP1 gene was used as the target,and plant tissue culture was carried out by soybean cotyledonary node transformation method to obtain T2 GmGBP1 interference transgenic soybean.The transcript was sequenced to obtain alternative splicing data.The alternative splicing genes in the sequencing results of WT and GmGBP1-i interference soybean leaves were further verified by RT-PCR.A total of 3 genes were alternatively spliced,indicating that the decrease of GmGBP1 interference expression led to the alternative splicing of downstream genes,which could affect the flowering time of soybean.This research provided a basis for further study of soybean GmGBP1 gene,and laid a theoretical basis for the mechanism of soybean flowering induction.
GmGBP1 is a gene related to photoperiod flowering time of soybean.In this experiment,GmGBP1 gene was used as the target,and plant tissue culture was carried out by soybean cotyledonary node transformation method to obtain T2 GmGBP1 interference transgenic soybean.The transcript was sequenced to obtain alternative splicing data.The alternative splicing genes in the sequencing results of WT and GmGBP1-i interference soybean leaves were further verified by RT-PCR.A total of 3 genes were alternatively spliced,indicating that the decrease of GmGBP1 interference expression led to the alternative splicing of downstream genes,which could affect the flowering time of soybean.This research provided a basis for further study of soybean GmGBP1 gene,and laid a theoretical basis for the mechanism of soybean flowering induction.
引文
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[23]李冬梅,陈薇,李永光,等.大豆子叶节遗传转化体系的优化研究[J].大豆科学,2018,37(4):531-538.
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[29] Liu H,Liu B,Zhao C,et al. The action mechanisms of plant cryptochromes[J]. Trends Plant Sci,2011,16(12):684-691.
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[34] Kleine T,Lockhart P,Batschauer A. An Arabidopsis protein closely related to synechocystis cryptochrome istargeted to organelles[J]. Plant J,2003,35(1):93-103.
[2]张彦威.大豆Gm GBP1基因参与光周期开花途径和逆境反应的功能分析[D].哈尔滨:东北农业大学,2013.
[3]赵琳,罗秋兰,杨春亮,等.大豆在暗诱导条件下差异表达c DNA文库的构建及分析[J].大豆科学,2007,26(2):134-139.
[4] Zhang Y W,Zhao L,Li H Y,et al. Gm GBP1,a homolog of human ski interacting protein in soybean,regulates flowering and stress tolerance in Arabidopsis[J]. BMC Plant Biol,2013,13(1):21.
[5]刘丽雪,吕庆雪,张彦威,等.大豆Gm GBP1在GA调控开花过程中的功能分析[J].作物杂志,2014(4):71-74.
[6]王萍,王罡,季静,等.大豆转基因体系的研究进展[J].遗传,2004,26(6):969-976.
[7]荣非.利用两种遗传转化方法获得抗草甘膦转基因大豆的研究[D].天津:天津大学,2015.
[8]姚丙晨,闫双勇,苏京平,等.大豆转基因研究进展[J].大豆科技,2015(5):18-26.
[9]王连铮.国内外大豆生产形势和大豆产业化问题[J].高科技与产业化,2008(7):67-69.
[10]王关林,方宏筠.植物基因工程原理与技术[M].北京:科学出版社,1998.
[11]张艳,南相日,满为群,等.大豆遗传转化及转基因大豆安全性研究进展[J].中国农学通报,2012,28(33):7-11.
[12]胡张华,黄锐之,刘智宏,等.利用花粉管导入法获得转反义PEP基因大豆植株[J].浙江农业学报,1999,11(2):99-100.
[13]李宝健,许新萍,石和平,等.应用电注射法将外源基因导入水稻种胚及获得转基因水稻植株的研究[J].中国科学:B辑化学生命科学地学,1991,21(3):270-275,339.
[14] Hansen G,Wright M S. Recent advances in the transformation of plants[J]. Trends Plant Sci,1999,4(6):226-231.
[15]刘海坤,卫志明.大豆遗传转化研究进展[J].植物生理与分子生物学学报,2005,31(2):126-134.
[16]侯文胜,林抗雪,陈普,等.大豆规模化转基因技术体系的构建及其应用[J].中国农业科学,2014,47(21):4198-4210.
[17]刘红亮,郑丽明,刘青青,等.非模式生物转录组研究[J].遗传,2013,35(8):955-970.
[18]朱玉贤,李毅,郑晓峰,等.现代分子生物学(第4版)[M].北京:高等教育出版社,2013.
[19]孙墨可,孙晓文,田娟,等.综述高通量转录组测序技术在生物中的应用[J].南方农机,2018,49(19):16-17.
[20]邢永强,刘国庆,蔡禄. Pre-mRNA选择性剪接的调控及选择性剪接数据库[J].中国生物化学与分子生物学报,2016,32(1):17-28.
[21] Wang Z F,Burge C B. Splicing regulation:from a parts list of regulatory elements to an integrated splicing code[J]. RNA,2008,14(5):802-813.
[22] Zhao L,Li M,Xu C,et al. Natural variation in Gm GBP1 promoter affects photoperiod control of flowering time and maturity in soybean[J]. Plant J,2018,96(1):147-162.
[23]李冬梅,陈薇,李永光,等.大豆子叶节遗传转化体系的优化研究[J].大豆科学,2018,37(4):531-538.
[24]吕金海,伍贤进.二种提取植物组织DNA方法的改进及结果比较[J].农业与技术,2005,25(2):64-65.
[25]彭凌涛.控制拟南芥和水稻开花时间光周期途径的分子机制[J].植物生理学通讯,2006,42(6):1021-1031.
[26] Koo B H,Yoo S C,Park J W,et al. Natural variation in Os PRR37 regulates heading date and contributes to rice cultivation at a wide range of latitudes[J]. Mol Plant,2013,6(6):1877-1888.
[27] Campoli C,Shtaya M,Davis S J,et al. Expression conservation within the circadian clock of a monocot:natural variation at barley Ppd-H1 affects circadian expression of flowering time genes,but not clock orthologs[J].BMC Plant Biol,2012,12(1):97.
[28] Turner,A. The pseudo-response regulator Ppd-H1provides adaptation to photoperiod in barley[J]. Science,2005,310(5750):1031-1034.
[29] Liu H,Liu B,Zhao C,et al. The action mechanisms of plant cryptochromes[J]. Trends Plant Sci,2011,16(12):684-691.
[30] Liu B,Zuo Z,Liu H,et al. Arabidopsis cryptochrome 1interacts with SPA1 to suppress COP1 activity in response to blue light[J]. Genes Dev,2011,25(10):1029-1034.
[31] Wang S,Li L,Xu P B,et al. CRY1 interacts directly with HBI1 to regulate its transcriptional activity and photomorpho genesis in Arabidopsis[J]. J Exp Bot,2018,69(16):3867-3881.
[32]李红丹,闫蕾,孙蕾,等.玉米隐花色素CRY1b和CRY2基因转录丰度对不同光质处理的响应[J].作物学报,2018,44(9):1290-1300.
[33]孙磊. Gm CRY1基因转化大豆及功能鉴定分析[D].长春:吉林大学,2011.
[34] Kleine T,Lockhart P,Batschauer A. An Arabidopsis protein closely related to synechocystis cryptochrome istargeted to organelles[J]. Plant J,2003,35(1):93-103.