Cannon M, Platz J, O'Leary M, Sookdeo C, Cannon F
BioTechnica International, Inc., Cambridge, MA 02140.
Plant Mol Biol. 1990 Jul;15(1):39-47. doi: 10.1007/BF00017722.
We have shown leaf-specific inhibition GUS gene expression in transgenic Nicotiana plants using an antisense RNA with a 41-base homology spanning the translation start codon of the gene. GUS was expressed from the nominally constitutive 35S promoter and the antisense RNA was expressed from the light-regulated ca/b promoter of Arabidopsis thaliana. A range of GUS inhibition from 0 to 100% was obtained by screening a small population of transgenic plants and the specific levels of inhibition observed were stably inherited in two generations. An antiGUS 'gene' dosage effect was observed in plants which were homozygous for antiGUS. RNA detection results suggest that duplex formation with the 41 base pair antiGUS RNA destabilized the GUS mRNA and that an excess of antisense RNA was not required. Our results demonstrate the potential of antisense RNA as a strategy for obtaining plant mutants, especially 'down mutations' in essential genes where only a short 5' sequence of the mRNA is required. They also suggest that the 'position effect' on gene expression could be used in conjunction with an antisense RNA strategy to provide a versatile approach for crop improvement.
我们已经利用一种反义RNA在转基因烟草植株中实现了叶片特异性抑制GUS基因表达，该反义RNA具有41个碱基的同源性，跨越该基因的翻译起始密码子。GUS由名义上组成型的35S启动子表达，反义RNA由拟南芥光调控的ca/b启动子表达。通过筛选一小群转基因植株，获得了0%至100%的一系列GUS抑制效果，所观察到的特定抑制水平在两代中稳定遗传。在反义GUS纯合的植株中观察到了反义GUS“基因”剂量效应。RNA检测结果表明，与41个碱基对的反义GUS RNA形成双链会使GUS mRNA不稳定，并且不需要过量的反义RNA。我们的结果证明了反义RNA作为获得植物突变体的一种策略的潜力，特别是在仅需要mRNA短5'序列的必需基因中的“下调突变”。它们还表明，基因表达的“位置效应”可与反义RNA策略结合使用，为作物改良提供一种通用方法。
