Ingelbrecht I, Breyne P, Vancompernolle K, Jacobs A, Van Montagu M, Depicker A
Laboratorium voor Genetica, Universiteit Gent, Belgium.
Gene. 1991 Dec 30;109(2):239-42. doi: 10.1016/0378-1119(91)90614-h.
When a promoterless marker gene is transformed into the plant genome using the Agrobacterium vector system, on average 30% of the T-DNA inserts produce gene fusions. This suggests that the T-DNA is preferentially integrated into transcribed regions. Here, we proposed that this transcriptional activity is responsible for some of the variation in expression frequently observed among independent transformants. Using hybrid gene constructions, we show that transcriptional readthrough into a downstream gene with opposite orientation substantially reduces expression of this gene both in transient expression and in transgenic plants. Furthermore, a poly(A) signal/terminator can block readthrough and restore the expression of the gene. Finally, enzymatic analysis of calli suggests that less variation in neomycin phosphotransferase II synthesis is observed when the gene is separated from plant DNA by promoter and terminator elements.
当使用农杆菌载体系统将无启动子标记基因转化到植物基因组中时，平均30%的T-DNA插入会产生基因融合。这表明T-DNA优先整合到转录区域。在此，我们提出这种转录活性是独立转化体间经常观察到的表达变异的部分原因。使用杂交基因构建体，我们表明转录通读进入反向的下游基因会在瞬时表达和转基因植物中显著降低该基因的表达。此外，聚腺苷酸化信号/终止子可以阻断通读并恢复该基因的表达。最后，愈伤组织的酶分析表明，当基因通过启动子和终止子元件与植物DNA分离时，新霉素磷酸转移酶II合成的变异较少。
