Department of Pathology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
Methods. 2010 May;51(1):92-100. doi: 10.1016/j.ymeth.2010.03.001. Epub 2010 Mar 4.
Transposon-mediated integration strategies in Xenopus offer simple and robust methods for the generation of germline transgenic animals. Co-injection of fertilized one-cell embryos with plasmid DNA harboring a transposon transgene and synthetic mRNA encoding the cognate transposase enzyme results in mosaic integration of the transposon at early cleavage stages that are frequently passed through the germline in the adult animal. Micro-injection of fertilized embryos is a routine procedure used by many laboratories that use Xenopus as a developmental model and, as such, the transposon transgenesis method can be performed without additional equipment or specialized methodologies. The methods for injecting Xenopus embryos are well documented in the literature so here we provide a step-by-step guide to other aspects of transposon transgenesis, including screening mosaic founders for germline transmission of the transgene and general husbandry considerations related to management of populations of transgenic frogs.
转座子介导的整合策略在爪蟾中为产生生殖系转基因动物提供了简单而强大的方法。将带有转座子转基因的质粒 DNA 和编码同源转座酶的合成 mRNA 共注射到受精卵中,导致转座子在早期卵裂阶段的镶嵌整合,这些整合在成年动物中经常通过生殖系传递。微注射受精卵是许多使用爪蟾作为发育模型的实验室常用的常规程序,因此,无需额外的设备或专门的方法即可进行转座子转基因方法。在文献中有详细记录的注射爪蟾胚胎的方法,因此在这里我们提供了转座子转基因的其他方面的分步指南,包括筛选具有转基因生殖系传递的嵌合体创始者以及与管理转基因青蛙种群相关的一般饲养注意事项。