Bai Xianquan, Wang Qiuyun, Chu Chengcai
State Key Laboratory of Plant Genomics and National Centre for Plant Gene Research (Beijing), Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China.
Transgenic Res. 2008 Dec;17(6):1035-43. doi: 10.1007/s11248-008-9182-7. Epub 2008 Apr 24.
Based on the Cre/loxP system, we have developed a novel marker-free system mediating a direct auto-excision of loxP-flanked marker genes from T(1 )transgenic rice without any treatment or further offspring crossing. To achieve this, the floral-specific promoter OsMADS45 was isolated from rice and the expression patterns of OsMADS45 promoter was characterised by using the pOs45:GUS transgenic plants. Furthermore, the binary vector with Cre recombinase under the control of OsMADS45 promoter was constructed and introduced into rice by Agrobacterium-mediated transformation and transgenic rice plants were generated. Southern blot analysis showed that auto-excision of the selective markers occurred in some T(1) progeny of the transgenic plants, suggesting that a high auto-excision frequency can be achieved with our Cre/loxP system. This auto-excision strategy provides an efficient way of removing the selectable marker gene from transgenic rice.
基于Cre/loxP系统，我们开发了一种新型无标记系统，可在无需任何处理或进一步子代杂交的情况下，直接从T(1)转基因水稻中自动切除loxP侧翼的标记基因。为实现这一目标，从水稻中分离出花特异性启动子OsMADS45，并利用pOs45:GUS转基因植物对OsMADS45启动子的表达模式进行了表征。此外，构建了在OsMADS45启动子控制下带有Cre重组酶的二元载体，并通过农杆菌介导的转化将其导入水稻，从而获得了转基因水稻植株。Southern杂交分析表明，转基因植株的一些T(1)子代中发生了选择标记的自动切除，这表明我们的Cre/loxP系统能够实现较高的自动切除频率。这种自动切除策略为从转基因水稻中去除选择标记基因提供了一种有效的方法。
