Efficient generation of transgenic mice with intact yeast artificial chromosomes by intracytoplasmic sperm injection.
نویسندگان
چکیده
The production of animals with large transgenes is an increasingly valuable tool in biotechnology and for genetic studies, including the characterization and manipulation of large genes and polygenic traits. In the present study, we describe an intracytoplasmic sperm injection (ICSI) method for the stable incorporation and phenotypic expression of large yeast artificial chromosomes (YAC) constructs of submegabase and megabase magnitude. By coinjecting spermatozoa and YACs into metaphase II oocytes, we were able to produce founders exhibiting germline transmission of an intact and functional transgene of 250 kilobases, carrying the mouse tyrosinase locus, used here as a reporter gene to rescue the albinism of recipient mice. More than 35% transgenesis was obtained for this YAC transgene. When compared with the pronuclear microinjection standard method, the efficiency of the ICSI-mediated YAC transfer system was significantly greater. In summary, we describe, for the first time, stable incorporation in the host genome and correct phenotypic expression of large DNA constructs mediated by ICSI.
منابع مشابه
A method for the generation of YAC transgenic mice by pronuclear microinjection.
Yeast artificial chromosomes (YACs) represent the latest generation of vectors which have the great advantage of large insert size. The introduction of YACs into mammalian cells and organisms has become an important goal, since it offers the potential to study the control of large and complex transcription units and identify genes by complementation. Microinjection into the nucleus is the most ...
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ورودعنوان ژورنال:
- Biology of reproduction
دوره 71 6 شماره
صفحات -
تاریخ انتشار 2004