CO-SUPPRESSION OF NITRATE REDUCTASE HOST GENES AND TRANSGENES IN TRANSGENIC TOBACCO PLANTS
CO-SUPPRESSION OF NITRATE REDUCTASE HOST GENES AND TRANSGENES IN TRANSGENIC TOBACCO PLANTS
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DOI：
10.1007/bf00279570
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发表时间：
1994-06-15
期刊：
影响因子：
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通讯作者：
VAUCHERET, H
中科院分区：
文献类型：
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作者：
DEBORNE, FD;VINCENTZ, M;VAUCHERET, H
将带有烟草硝酸还原酶cDNA（NIA）的全部或部分克隆在花椰菜花叶病毒的35 S RNA的启动子和终止子序列之间的构建体引入烟草中，试图改善硝酸盐同化。获得了几株NIA mRNA和硝酸还原酶（NR）活性均升高的转基因植株。此外，还获得了少数表现出NR缺陷突变体的褪绿表型特征的植物。其中一株植株不含NIA mRNA，无NR活性，硝酸盐积累。因此，假设这种表型是由35 S-NIA转基因和宿主NIA基因的共同抑制引起的。NR-缺陷植物中也发现过表达NIA mRNA的转化体的后代。遗传分析表明，这些NR-缺陷的植物是纯合的35 S-NIA转基因，虽然不是所有的纯合植物的NR缺陷。在纯合植物的后代中，正常植物与NR缺陷植物的比率在每一代保持恒定，而不管上一代中NIA基因的表达状态（活性或非活性）如何。当在法国的两个地区进行田间试验时，这一比例也保持不变：凡尔赛和贝尔热拉克。对纯合植株的分析表明，在有性生殖的某些阶段，共抑制是可逆的。事实上，宿主基因和转基因在每一代都会重新激活，并且共抑制总是在正常生长的滞后期之后出现，这表明这种现象是发育调节的。我们观察到，当植物最初生长在有限的光照和/或有限的硝酸盐供应（光和硝酸盐都需要宿主NIA基因的表达）浇水时，共抑制的触发被延迟。然而，这种延迟并没有影响正常和NR-缺陷植物转移到硝酸盐施肥领域后的最终比例。独立的转化体表现出不同的共抑制率或根本没有共抑制，无论转基因拷贝数，这表明转基因周围的基因组序列可能在确定共抑制中发挥作用。
Constructs carrying the entire or part of the tobacco nitrate reductase cDNA (NIA) cloned between the promoter and terminator sequences of the 35S RNA of the cauliflower mosaic virus were introduced into tobacco, in an attempt to improve nitrate assimilation. Several transgenic plants that had elevated NIA mRNA and nitrate reductase (NR) activity were obtained. In addition, a few plants that exhibited a chlorotic phenotype characteristic of NR-deficient mutants were also obtained. One of these plants contained no NIA mRNA, no NR activity and accumulated nitrate. This phenotype was therefore assumed to result from co-suppression of 35S-NIA transgenes and host NIA genes. NR-deficient plants were also found among the progeny of transformants overexpressing NIA mRNA. Genetic analyses indicated that these NR-deficient plants were homozygous for the 35S-NIA transgene, although not all homozygous plants were deficient for NR. The ratio of normal to NR-deficient plants in the progeny of homozygous plants remained constant at each generation, irrespective of the state of expression of the NIA genes (active or inactive) in the previous generation. This ratio also remained unchanged when field trials were performed in two areas of France: Versailles and Bergerac. The analysis of homozygous plants revealed that co-suppression was reversible at some stage of sexual reproduction. Indeed, host genes and transgenes reactivated at each generation, and co-suppression always appeared after a lag period of normal growth, suggesting that the phenomenon is developmentally regulated. We observed that the triggering of co-suppression was delayed when plants were initially grown under limited light and/or watered with limited nitrate supply (light and nitrate both being required for the expression of the host NIA genes). However, this delay did not affect the final ratio between normal and NR-deficient plants after transfer to nitrate-fertilized fields. Independent transformants exhibited either different co-suppression ratios or no co-suppression at all, irrespective of the transgene copy number, suggesting that genomic sequences surrounding the transgene might play a role in determining co-suppression.