Genome editing and animal models

Transgenic technology allows a gene of interest to be introduced into the genome of a laboratory animal, and provides an extremely powerful tool to dissect the molecular mechanisms of disease. Transgenic mouse models made by microinjection of DNA into zygotic pro- nuclei in particular have been widely used by the genetics community for 30 years. However, it remains a rather crude approach： injected sequences randomly insert in multiple copies as concatamers, they can be mutagenic, and they have variable or silenced expression depending on the site of integration, a phenomenon called position effects. As a result, multiple lines are required in order to confirm appropriate transgene expression. This can be partially overcome by flanking transgenes with insulator sequences to protect the transgene from the influence of the sur- rounding regulatory elements. Large （〈300 kb） BAC- based transgenic vectors have also been shown to be more resistant to position effects. However, animals carrying extra copies of fairly large regions of the genome could have unpredictable phenotypes. The most effective method used to control for position effects is to target transgene insertion to specific genomic loci, the so-called targeted transgenesis; for instance, the fast, site-specific transgenic technology TargattTM. The purpose of this review is to provide an overview on the current existing methods for making targeted transgenic mouse models.     

利用TALEN技术制备HE4（WFDC2）敲除小鼠模型

目的 利用显微注射技术和 TALEN技术构建HE4（WFDC2）敲除小鼠并对其进行表型分析。方法 设计Wfdc2敲除位点,构建TALEN载体,体外转录TALENs获得mRNA并通过显微注射技术注射到C57BL/6 J小鼠受精卵,对F0代小鼠进行DNA鉴定获得HE4（WFDC2）敲除小鼠,观察并统计 HE4（WFDC2）敲除小鼠出生及存活情况。结果 在Wfdc2第一个外显子上设计了TALEN识别剪切位点,向受精卵注射TALENs mRNA获得了24只F0代小鼠,其中1只发生移码突变,成功制备了HE4（WFDC2）敲除小鼠,并发现纯合 HE4敲除小鼠出生后在短时间内致死。结论 通过TALEN技术成功制备HE4（WFDC2）敲除小鼠,纯合HE4（WFDC2）敲除小鼠出生致死。