A simple and highly efficient transgenesis method in mice with the Tol2 transposon system and cytoplasmic microinjection.
Creating transgenic mice is an important technology for genetic studies and is currently performed by pronuclear microinjection of plasmid DNA into fertilized eggs. Since survival of injected embryos and integration of plasmid DNA are not efficient, total efficiency is only around 3% with a standard protocol. To circumvent this problem, here we describe a novel transgenesis method, the Tol2-mediated cytoplasmic injection method (Tol2:CI). We injected a foreign DNA cloned in a Tol2-transposon vector together with the transposase mRNA into the cytoplasm of fertilized eggs. As expected, the survival rate of the injected embryos was increased drastically. Also, the foreign DNA was transposed from the plasmid to the genome and transmitted to the next generation very efficiently. Together, the overall transgenic efficiency became more than 20%. Considering its simplicity and perfect compatibility with existing pronuclear microinjection facilities, we propose that the Tol2:CI method is applicable to high throughput functional genomics studies.
Sumiyama
Kenta
K
Division of Population Genetics, National Institute of Genetics, Mishima, Shizuoka, Japan. ksumiyam@lab.nig.ac.jp
Kawakami
Koichi
K
Yagita
Kazuhiro
K
D000818:Animals; D003593:Cytoplasm; D004251:DNA Transposable Elements; D051379:Mice; D008822:Mice, Transgenic / Q000235:genetics*; D008845:Microinjections; D012333:RNA, Messenger / Q000235:genetics / Q000494:pharmacology*; D015053:Zygote
D004251:DNA Transposable Elements; D012333:RNA, Messenger