EMBL-CRG Systems Biology Unit, Centre for Genomic Regulation (CRG), Dr. Aiguader 88, 08003 Barcelona, Spain; Universitat Pompeu Fabra (UPF), Dr. Aiguader 88, 08003 Barcelona, Spain.
EMBL-CRG Systems Biology Unit, Centre for Genomic Regulation (CRG), Dr. Aiguader 88, 08003 Barcelona, Spain; Universitat Pompeu Fabra (UPF), Dr. Aiguader 88, 08003 Barcelona, Spain; Institucio ́Catalana de Recerca i Estudis Avançats (ICREA), Passeig Lluis Companys 23, 08010 Barcelona, Spain.
Methods. 2014 Aug 1;68(3):409-16. doi: 10.1016/j.ymeth.2014.05.004. Epub 2014 May 20.
The use of drugs and drug resistance genes is a powerful method to select for the presence of a transgene. Unlike methods that require the complementation of a genetic mutation, this system can be used on any genetic background. Drug selection does not require extensive manipulation or costly equipment, yet it is very rapid and can achieve extremely high efficiency, selecting a small number of transgenic worms from among millions of non-transgenic worms. Introducing integrated transgenes into Caenorhabditis elegans by microparticle bombardment represents just such a challenge. Here we describe in detail the protocol we have developed for dual-drug selection in liquid with puromycin and G418 which works well in a variety of Caenorhabditis species. We also show that single drug selection with only puromycin or only G418 is effective in C. elegans. The growing number of drug selection markers that have been adapted to C. elegans are an important addition to the genetic toolkit at our disposal.
药物和耐药基因的使用是选择转基因存在的有力方法。与需要遗传突变互补的方法不同,该系统可用于任何遗传背景。药物选择不需要广泛的操作或昂贵的设备,但非常快速且效率极高,可从数百万个非转基因蠕虫中选择少数转基因蠕虫。通过微粒轰击将整合的转基因引入秀丽隐杆线虫代表了这样的一个挑战。在这里,我们详细描述了我们开发的用于液体中嘌呤霉素和 G418 双重药物选择的方案,该方案在多种秀丽隐杆线虫物种中均有效。我们还表明,仅用嘌呤霉素或仅用 G418 进行单一药物选择在秀丽隐杆线虫中也是有效的。越来越多的已适应秀丽隐杆线虫的药物选择标记物是我们可用遗传工具包的重要补充。