Cain-Hom Carol, Splinter Erik, van Min Max, Simonis Marieke, van de Heijning Monique, Martinez Maria, Asghari Vida, Cox J Colin, Warming Søren
Department of Transgenic Technology, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA.
Cergentis BV, Yalelaan 62, 3584 CM Utrecht, the Netherlands.
Nucleic Acids Res. 2017 May 5;45(8):e62. doi: 10.1093/nar/gkw1329.
Cre/LoxP technology is widely used in the field of mouse genetics for spatial and/or temporal regulation of gene function. For Cre lines generated via pronuclear microinjection of a Cre transgene construct, the integration site is random and in most cases not known. Integration of a transgene can disrupt an endogenous gene, potentially interfering with interpretation of the phenotype. In addition, knowledge of where the transgene is integrated is important for planning of crosses between animals carrying a conditional allele and a given Cre allele in case the alleles are on the same chromosome. We have used targeted locus amplification (TLA) to efficiently map the transgene location in seven previously published Cre and CreERT2 transgenic lines. In all lines, transgene insertion was associated with structural changes of variable complexity, illustrating the importance of testing for rearrangements around the integration site. In all seven lines the exact integration site and breakpoint sequences were identified. Our methods, data and genotyping assays can be used as a resource for the mouse community and our results illustrate the power of the TLA method to not only efficiently map the integration site of any transgene, but also provide additional information regarding the transgene integration events.
Cre/LoxP技术在小鼠遗传学领域被广泛用于基因功能的空间和/或时间调控。对于通过原核显微注射Cre转基因构建体产生的Cre品系，整合位点是随机的，在大多数情况下并不清楚。转基因的整合可能会破坏内源基因，潜在地干扰对表型的解释。此外，在携带条件性等位基因和特定Cre等位基因的动物进行杂交育种时（如果等位基因位于同一条染色体上），了解转基因的整合位置对于杂交方案的设计很重要。我们利用靶向位点扩增（TLA）技术有效地定位了七个先前发表的Cre和CreERT2转基因品系中的转基因位置。在所有品系中，转基因插入都与不同复杂程度的结构变化相关，这说明了检测整合位点周围重排的重要性。在所有七个品系中，都确定了精确的整合位点和断点序列。我们的方法、数据和基因分型检测可作为小鼠研究群体的资源，我们的结果表明TLA方法不仅能有效地定位任何转基因的整合位点，还能提供有关转基因整合事件的额外信息。
