Enhanced transgene expression in rice following selection controlled by weak promoters.
Enhanced transgene expression in rice following selection controlled by weak promoters.
复制标题
弱启动子控制选择后转基因表达增强
DOI：
10.1186/1472-6750-13-29
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发表时间：
2013-03-27
影响因子：
3.5
通讯作者：
Chen J
中科院分区：
文献类型：
--
作者：
Zhou J;Yang Y;Wang X;Yu F;Yu C;Chen J;Cheng Y;Yan C;Chen J
背景能够在植物中实现高水平转基因表达的技术对于植物制造的重组药物蛋白的商业生产或其它基因转移相关策略是有吸引力的。提高所需转基因产物产量的常规方法是使用强启动子来控制转基因的表达。虽然许多这样的启动子已被确定和表征，从一个单一的启动子获得的增加最终是有限的，在一定程度上results在这项研究中，我们报告了一种方法，通过使用一个弱启动子为基础的选择系统在转基因水稻放大一个单一的启动子的效果。tCUP 1是一个来源于烟草隐性启动子（tCUP）的片段，通过融合到一个β-葡萄糖醛酸酶（GUS）报告基因和一个潮霉素磷酸转移酶（HPT）选择标记上来检测其在水稻中的活性。tCUP 1启动子允许转化的水稻植株的恢复，并赋予GUS报告基因的组织特异性表达，但在驱动抗性愈伤组织生长的选择标记方面比CaMV 35 S启动子弱得多。然而，在通过使用tCUP 1选择的抗性愈伤组织和再生的转基因植株中，由于多个T-DNA插入的累加效应，绿色荧光蛋白（GFP）的组成型表达显著增加。通过使用另一个相对较弱的启动子从胭脂碱合成酶（Nos）.ConclusionsThe使用弱启动子衍生的选择标记导致高的T-DNA拷贝数，然后大大增加外源基因的表达的衰减选择弱启动子和海拔之间的相关性进行了确认。本文所述的方法提供了一种有效的途径，通过拷贝数操纵来稳健地增强外源转基因在水稻中的表达。
BackgroundTechniques that enable high levels of transgene expression in plants are attractive for the commercial production of plant-made recombinant pharmaceutical proteins or other gene transfer related strategies. The conventional way to increase the yield of desired transgenic products is to use strong promoters to control the expression of the transgene. Although many such promoters have been identified and characterized, the increase obtainable from a single promoter is ultimately limited to a certain extent.ResultsIn this study, we report a method to magnify the effect of a single promoter by using a weak promoter-based selection system in transgenic rice. tCUP1, a fragment derived from the tobacco cryptic promoter (tCUP), was tested for its activity in rice by fusion to both a β-glucuronidase (GUS) reporter and a hygromycin phosphotransferase (HPT) selectable marker. The tCUP1 promoter allowed the recovery of transformed rice plants and conferred tissue specific expression of the GUS reporter, but was much weaker than the CaMV 35S promoter in driving a selectable marker for growth of resistant calli. However, in the resistant calli and regenerated transgenic plants selected by the use of tCUP1, the constitutive expression of green fluorescent protein (GFP) was dramatically increased as a result of the additive effect of multiple T-DNA insertions. The correlation between attenuated selection by a weak promoter and elevation of copy number and foreign gene expression was confirmed by using another relatively weak promoter from nopaline synthase (Nos).ConclusionsThe use of weak promoter derived selectable markers leads to a high T-DNA copy number and then greatly increases the expression of the foreign gene. The method described here provides an effective approach to robustly enhance the expression of heterogenous transgenes through copy number manipulation in rice.