以皖豆28为受体的抗草甘膦转基因大豆的获得及后代抗性鉴定与分析
|
摘要
研究以皖豆28(WD28)为受体,采用农杆菌介导法,将抗草甘膦g10-epsps基因转入到大豆中,先后获得6株T_0代植株,转化效率为1.23%。对获得的转化体的下代进行PCR检测,均获得目的片段。同时取其根、茎、叶、花、荚、种子进行Western blot检测,均有外源蛋白表达。T_1代通过喷施田间推荐浓度的草甘膦进行抗性鉴定,统计抗性单株数,进行X~2分析,除转化体WD28-3外,其余5个后代的抗感分离比均符合3∶1的比例。
In this study,six glyphosate resistant soybean plants(T_0 generation) were obtained through Agrobacterium mediated method.The efficiency of transformation was 1.23%.PCR and Western blot confirmed that the g10-epsps gene was intergrated into the soybean(Wandou 28) genome and expressed quite well in root,stem,leaf,flower,pod and seed.Resistance identification of T_1 plants was carried out by spraying glyphosate under the recommended concentration.The resistant to sensitive ratios of the plants derived from five T_0 plants,except WD28-3,all fit to 3∶1.
In this study,six glyphosate resistant soybean plants(T_0 generation) were obtained through Agrobacterium mediated method.The efficiency of transformation was 1.23%.PCR and Western blot confirmed that the g10-epsps gene was intergrated into the soybean(Wandou 28) genome and expressed quite well in root,stem,leaf,flower,pod and seed.Resistance identification of T_1 plants was carried out by spraying glyphosate under the recommended concentration.The resistant to sensitive ratios of the plants derived from five T_0 plants,except WD28-3,all fit to 3∶1.
引文
[1] 赵团结,盖钧镒,李海旺,等.超高产大豆育种研究的进展与讨论[J].中国农业科学,2006,39(1):29-37.
[2] 王宜林,李宏.抗除草剂转基因植物的研究现状[J].生物学通报,2005,40(11):15-17.
[3] 邱丽娟,王昌陵,周国安,等.大豆分子育种研究进展[J].中国农业科学,2007,40:2418-2436.
[4] Owens L D,Cress D E.Genotypic variability of soybean response to Agrobacterium strains harboring the Ti or Ri plasmids[J].Plant Physiology,1985,77(1):87-94.
[5] McKenzie M,Cress W.The evaluation of South African cultivars of soybean for their susceptibility to Agrobacterium tumefacious and the production of transgenic soybean[J].South African Journal of Science,1992,88(4):193-196.
[6] Bailey M,Boerma H,Parrot W.Genotype effects on proliferative embryogenesis and plant regeneration of soybean[J].In Vitro Cellular & Developmental Biology-Plant,1993,29(3):102-108.
[7] Droste A,Bodanese-Zanettini M,Mundstock E,et al.Susceptibility of Brazilian soybean cultivars to Agrobacterium tumefacious[J].Revista Brasileira de Genetica,1994,17(1):83-88
[8] Mauro A,Pfeiffer T,Collins G.Inheritance of soybean susceptibility to Agrobacterium tumefacious and its relationship to transformation[J].Crop Science,1995,35(4):1152-1156.
[9] Dale P J,Clarke B,Fontes E M G.Potential for the environmental impact of transgenic crops[J].Nature Biotechnology,2002,20(6):567-574.
[10] 钟金传,吴文良,夏友富,等.转基因大豆发展及中国大豆产业对策[J].中国农业大学学报,2005,10(4):43-50.
[11] Beckie H J,Hall L M,Warwick S I.Impact of herbicide-resistant crops as weeds in Canada[C]//The BCPC Conference.Weeds.Brighton,UK:BCPC,2001:135-142.
[12] DeCleene M,De Lay J.The host range of crown gall[J].The Botanical Review,1976,42(4):389-466.
[13] Trick H N,Dinkins R D,Santarem E R,et al.Recent advances in soybean transformation[J].Plant Tissue Culture and Biotechnology,1997,24:39-26.
[14] Barwale U,Kerns H,Widholm J.Plant regeneration from callus cultures of several soybean genotypes via embryogenesis and organogenesis[J].Planta,1986,167(4):473-481.
[15] Hinchee M A W,Connor Ward D V,Newell C A.Production of transgenic soybean plants using Agrobacterium mediated DNA transfer[J].Nature Biotechnology,1988,6:915-922.
[16] Danaldson P A,Simmonds D H.Susceptibility to Agrobacterium tumefacious and cotyledonary node transformation in short season soybean[J].Plant Cell Reports,2000,19:478-484.
[2] 王宜林,李宏.抗除草剂转基因植物的研究现状[J].生物学通报,2005,40(11):15-17.
[3] 邱丽娟,王昌陵,周国安,等.大豆分子育种研究进展[J].中国农业科学,2007,40:2418-2436.
[4] Owens L D,Cress D E.Genotypic variability of soybean response to Agrobacterium strains harboring the Ti or Ri plasmids[J].Plant Physiology,1985,77(1):87-94.
[5] McKenzie M,Cress W.The evaluation of South African cultivars of soybean for their susceptibility to Agrobacterium tumefacious and the production of transgenic soybean[J].South African Journal of Science,1992,88(4):193-196.
[6] Bailey M,Boerma H,Parrot W.Genotype effects on proliferative embryogenesis and plant regeneration of soybean[J].In Vitro Cellular & Developmental Biology-Plant,1993,29(3):102-108.
[7] Droste A,Bodanese-Zanettini M,Mundstock E,et al.Susceptibility of Brazilian soybean cultivars to Agrobacterium tumefacious[J].Revista Brasileira de Genetica,1994,17(1):83-88
[8] Mauro A,Pfeiffer T,Collins G.Inheritance of soybean susceptibility to Agrobacterium tumefacious and its relationship to transformation[J].Crop Science,1995,35(4):1152-1156.
[9] Dale P J,Clarke B,Fontes E M G.Potential for the environmental impact of transgenic crops[J].Nature Biotechnology,2002,20(6):567-574.
[10] 钟金传,吴文良,夏友富,等.转基因大豆发展及中国大豆产业对策[J].中国农业大学学报,2005,10(4):43-50.
[11] Beckie H J,Hall L M,Warwick S I.Impact of herbicide-resistant crops as weeds in Canada[C]//The BCPC Conference.Weeds.Brighton,UK:BCPC,2001:135-142.
[12] DeCleene M,De Lay J.The host range of crown gall[J].The Botanical Review,1976,42(4):389-466.
[13] Trick H N,Dinkins R D,Santarem E R,et al.Recent advances in soybean transformation[J].Plant Tissue Culture and Biotechnology,1997,24:39-26.
[14] Barwale U,Kerns H,Widholm J.Plant regeneration from callus cultures of several soybean genotypes via embryogenesis and organogenesis[J].Planta,1986,167(4):473-481.
[15] Hinchee M A W,Connor Ward D V,Newell C A.Production of transgenic soybean plants using Agrobacterium mediated DNA transfer[J].Nature Biotechnology,1988,6:915-922.
[16] Danaldson P A,Simmonds D H.Susceptibility to Agrobacterium tumefacious and cotyledonary node transformation in short season soybean[J].Plant Cell Reports,2000,19:478-484.