Pi-ta+基因和几丁质酶基因双价基因导入水稻的研究
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云南省基金项目(2011FZ217);云南省科技项目(2012CH010);国家转基因专项(2009ZX08001-007B,2011ZX08001-002)


Pi-ta+ Gene and Bean Chitinase Gene Transformation into Rice
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    摘要:

    为了提高‘云资粳41’和‘云资粳43’的抗稻瘟病能力,利用农杆菌介导法将二价表达载体pCAMBIA1300-Pi-ta+-Bchi转化到水稻愈伤组织中。经组织培养获得再生苗,再通过氯酚红显色法、PCR检测和抗稻瘟病鉴定法获得抗稻瘟病的转基因植株,为进一步创建持久、广谱抗稻瘟病水稻新材料奠定基础。结果显示:(1)抗性愈伤组织经分化和生根培养后,共获得T0代水稻再生苗137株,其中‘云资粳41’14株,‘中花11’82株,‘云资粳43’41株。(2)经氯酚红显色法和PCR对再生苗检测,‘中花11’、‘云资粳41’、‘云资粳43’的转化苗阳性率分别为66%、43%和55%。证明2个外源基因已经整合到水稻基因组中。(3)对转化阳性植株温室接种稻瘟病病菌66b鉴定结果显示,转基因植株较非转基因植株对稻瘟病的抗性明显增强,而且转Pi-ta+基因和几丁质酶基因双价的水稻植株比转单价Pi-ta+基因或几丁质酶基因的水稻植株抗稻瘟病能力强。(4)氯酚红检测结果存在一定的假阳性,PCR检测结果更真实可靠,但氯酚红显色法方便、快速,结果观察直观,可对大量的转基因植株进行初步筛选。研究表明,转Pi-ta+基因和几丁质酶基因双价基因的水稻植株具有更高的抗稻瘟病能力。

    Abstract:

    In order to improve the rice blast resistance of ‘Yunzijing 41’and‘Yunzijing 43’,we transformed bivalent expression vector pCAMBIA1300-Pi-ta+-Bchi into rice calli using the agrobacterium- mediated method.Regeneration seedlings through tissue culture are tested through CPR assay,PCR and rice blast resistant assay method and a part of these are determined to be transgenic plants.This experiment laid the foundation for breeding the longer and higher blast resistance rice variety in future.Results show:(1)Resistant callus passed through differentiation and rooting cultivation,we received 137 T0 generation of rice seedlings,including 14 ‘Yunzijing 41’,82 ‘Zhonghua 11’,41 ‘Yunzijing 43’.(2)After CPR assay and PCR,the conversions of ‘Yunzijing 41’,‘Yunzijing 43’ and ‘Zhonghua 11’transgenic positive rice plants were 66%,43% and 55%,respectively.Transgenic plants were confirmed that two exogenous genes had been integrated into the rice genome.(3)After be identified by inoculation with rice blast strain 66 b,transgenic plants enhanced clearer resistance to rice blast than non-transgenic plants;at the same time,transgenic plants of Pi-ta+ gene and bean chitinase gene had more obvious ability of resistance to rice blast than transgenic plants of Pi-ta+ gene or bean chitinase gene.(4)CPR assay results existed in some false positives,PCR detection results are more reliable.But CPR assay is more convenient,faster and the results is more intuitive.This can be used for preliminary screening.The research indicates that transgenic plants of Pi-ta+ gene and bean chitinase gene had a higher ability of resistance to rice blast.

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张 秀,钟巧芳,付 坚,等.Pi-ta+基因和几丁质酶基因双价基因导入水稻的研究[J].西北植物学报,2013,33(10):1953-1957

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  • 在线发布日期: 2013-11-13
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