| 韩德俊,陈耀锋,李春莲,郭东伟,李振岐.转甜菜碱醛脱氢酶基因油菜的获得及其耐盐性研究[J].干旱地区农业研究,2007,(4):6~11 |
| 转甜菜碱醛脱氢酶基因油菜的获得及其耐盐性研究 |
| Agrobacterium-mediated transformation with a gene encoding for betaine-aldehyde dehydrogenase(BADH) in brassica napus |
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| DOI：10.7606/j.issn.1000-7601.2007.04.02 |
| 中文关键词: 甘蓝型油菜 遗传转化 甜菜碱醛脱氢酶 耐盐 |
| 英文关键词:Brassica napus transformation BADH tolerance to salt |
| 基金项目:西北农林科技大学科研专项资助项目（04ZR007） |
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| 中文摘要: |
| 以9个甘蓝型油菜（Brassica nupas L.）材料为受体，分别以其子叶和下胚轴为外植体，利用农杆菌将来自山菠菜的BADH基因导入甘蓝型油菜，研究了转化植株的抗盐性。结果表明，依据对农杆菌侵染的反应，参试品种可划分为三种类型：抗性类型（resistant type，R）；超敏感类型（supper susceptive type，SS）和中间类型（middle type，M）。R型外植体切口易发生过敏反应导致失去再生能力；SS型外植体极易受农杆菌快速增殖影响，最终导致整体褐化死亡；只有M型外植体还保留部分再生能力，可产生少量不定芽。通过改进培养方式可提高外植体在农杆菌胁迫下的芽发生频率，在无菌滤纸上进行共培养，不仅能降低“玻璃化”现象，还能延长共培养时间，为外植体细胞的再分化提供了时间保障。分化培养基中加入3～6 mg/L AgNO3可减轻外植体褐化程度，提高不定芽分化率。PCR分析和BADH活性检测发现，有部分转化体BADH酶活性明显高于对照，表明转基因植株在盐胁迫下外源BADH基因已表达，转基因植株能够在0.5%NaCl培养基上正常生长，而对照在相同的培养基上生长缓慢且生根困难。 |
| 英文摘要: |
| Glycinebetaine(GB) is one of the organic compatible solutes that can accumulate rapidly in many plants under salinity or drought stress. Betaine aldehyde dehydrogenase(BADH) is the enzyme that catalyzes the second step in the synthesis of the GB. Cotyledon and hypocotyl explants of 9 varieties of B. napus were used to investigate the regeneration competence of them under the stress of Agrobacterium, and establish an efficient transformation experiments for the BADH gene. The result indicated that the 9 varieties could be divided into three types according to the response to the infection of Agrobacterium: resistant type(R); supper susceptive type(SS) and middle type(M). The R type could not form regeneration shoot owing to hypersusceptibility response at the cut of the explant; the SS type could not also form regeneration shoot owing to propagate so far on the surfer of the explant; and only the M type could form regeneration shoot. The frequency of regeneration shoot can be enhanced by optimizing the co-cultivate condition: using filter paper as co-cultivate support, it results in not only avoiding vitrification of explant, but also prolong the time of co-cultivate, thus the cells on the explant could have enough time to regenerate. The use of AgNO3 (3～6 mg/L) was a prerequisite for efficient shoot regeneration under selective conditions, but the mount is dependented on genotype. The hypocotyl explant was more easy death infected by Agrobacterium than cotyledon explant. two Brassica napus lines of breeding, P137 and 487, using Agrobacterium tumefaciens strain AGL1 carrying a binary vector pBin438, with cotyledon and hypocotyl regeneration system. Polymerase chain reaction analyses demonstrated that the BADH gene had integrated into the genome of B. napus. Transgenic B. napus plants showed the levels of activity of BADH were similar to or higher than wild-type plants leaves after exposure to salt stress. Observations on rooting development under the salt stress with concentrations up to 0.5% suggested that the transgenic plants exhibited tolerance to salt stress than wild-type plants. |
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