Betzner A S, Oakes M P, Huttner E
Group Limagrain Pacific Pty Limited, Research School of Biological Sciences, Australian National University, Canberra, Australia.
Plant J. 1997 Mar;11(3):587-95. doi: 10.1046/j.1365-313x.1997.11030587.x.
An artificial amber suppressor tRNA(Leu) gene (supL) was physically linked to a mutated gus reporter gene, p35S-gus(amL), which was inactivated by an amber stop codon (amL). Upon introduction into Arabidopsis thaliana, the presence of the supL gene was found to be correlated with cytotoxic effects observed during tissue culture and in mature plants. Those primary transformants that displayed cytotoxic symptoms were shown by X-Glu staining to express GUS as a result of amber stop codon suppression in vivo. Phenotypically normal lines were found by RT-PCR to express supL. GUS activity above background level was barely detectable in these plants, indicating a low level expression of supL. However, the remaining suppressor activity was still sufficient to transactivate an amber-mutated male sterility gene, pA9-barnase(amL1) when combined within the same plant by crossing. The suppressor tRNA(Leu) gene may thus be used in transgenic plants for gene transactivation.
一个人工琥珀抑制tRNA（Leu）基因（supL）与一个突变的gus报告基因p35S - gus（amL）物理连锁，该报告基因因琥珀终止密码子（amL）而失活。导入拟南芥后，发现supL基因的存在与组织培养和成熟植株中观察到的细胞毒性效应相关。那些表现出细胞毒性症状的初级转化体经X - Glu染色显示，由于体内琥珀终止密码子抑制而表达GUS。通过RT - PCR发现表型正常的株系表达supL。在这些植物中几乎检测不到高于背景水平的GUS活性，表明supL表达水平较低。然而，当通过杂交在同一植株中组合时，剩余的抑制活性仍足以反式激活一个琥珀突变的雄性不育基因pA9 - barnase（amL1）。因此，抑制tRNA（Leu）基因可用于转基因植物中的基因反式激活。
