Peterhans A, Datta S K, Datta K, Goodall G J, Potrykus I, Paszkowski J
Institute of Plant Sciences, Swiss Federal Institute of Technology, ETH-Zürich, Switzerland.
Mol Gen Genet. 1990 Jul;222(2-3):361-8. doi: 10.1007/BF00633841.
Heterologous gene expression experiments have shown that genes of Monocotyledoneae are often not transcribed in Dicotyledoneae, or produce pre-mRNA that is inefficiently or aberrantly processed. It is however not known how correctly and efficiently dicotyledon-specific gene expression signals are recognized in cells of Monocotyledoneae. Here we address this question using tobacco (Nicotiana tabacum) and rice (Oryza sativa) protoplasts transformed with the same hybrid gene constructs. Constructs including the nptII protein coding sequence fused to Cauliflower Mosaic Virus (CaMV) promoter and polyadenylation signals were used to obtain stably transformed cell lines of tobacco and rice. In one of the constructs the nptII coding region is interrupted by a modified intron-3 sequence from the soybean phaseolin gene. Although the mean number of hybrid gene copies integrated into the rice genome was on average 5- to 10-fold higher than in tobacco, the steady-state transcript level was 3 times lower. A lower level of transcript was also observed in transient expression experiments. The amount of the mature mRNA was not influenced by the presence of the intron. The phaseolin intron was processed in rice with high efficiency and an accuracy indistinguishable from that seen in tobacco.
异源基因表达实验表明，单子叶植物的基因通常不在双子叶植物中转录，或者产生加工效率低下或异常的前体mRNA。然而，目前尚不清楚在单子叶植物细胞中，双子叶植物特异性基因表达信号是如何被正确且高效识别的。在这里，我们使用用相同的杂种基因构建体转化的烟草（Nicotiana tabacum）和水稻（Oryza sativa）原生质体来解决这个问题。使用包含与花椰菜花叶病毒（CaMV）启动子和聚腺苷酸化信号融合的nptII蛋白编码序列的构建体来获得烟草和水稻的稳定转化细胞系。在其中一个构建体中，nptII编码区被来自大豆菜豆蛋白基因的修饰内含子-3序列打断。尽管整合到水稻基因组中的杂种基因拷贝平均数比烟草中平均高5至10倍，但稳态转录水平却低3倍。在瞬时表达实验中也观察到较低水平的转录本。成熟mRNA的量不受内含子存在的影响。菜豆内含子在水稻中高效加工，准确性与在烟草中所见无异。
