A YEAST ARTIFICIAL CHROMOSOME COVERING THE TYROSINASE GENE CONFERS COPY NUMBER-DEPENDENT EXPRESSION IN TRANSGENIC MICE
A YEAST ARTIFICIAL CHROMOSOME COVERING THE TYROSINASE GENE CONFERS COPY NUMBER-DEPENDENT EXPRESSION IN TRANSGENIC MICE
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DOI：
10.1038/362258a0
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发表时间：
1993-03-18
期刊：
影响因子：
64.8
通讯作者：
SCHUTZ, G
中科院分区：
文献类型：
--
作者：
SCHEDL, A;MONTOLIU, L;SCHUTZ, G
转基因在小鼠中的表达通常不能遵循正常的时间和空间模式，也不能达到与内源性拷贝相同的水平1，2。只有在特殊情况下，位置无关和拷贝数依赖的表达被再现3。标准构建体的大小限制可能会阻止重要的远程调控元件的包含。酵母人工染色体（YAC）4提供了克隆大的DNA片段的手段，并且YAC DNA到体细胞中的转移已被证实5 -9。我们先前已经产生了携带35-磷酸酶YAC构建体的转基因小鼠10。 在这里，我们报告了转移的250个酪氨酸酶YAC覆盖小鼠酪氨酸酶基因到小鼠的原核注射凝胶纯化的YAC DNA。将YAC插入到小鼠基因组中而没有主要重排，并且YAC携带的酪氨酸酶基因的表达导致受体小鼠的白化病表型的完全拯救。从转基因的表达达到的水平与内源基因的水平相当，并显示拷贝数依赖性和位置的独立性。
EXPRESSION of transgenes in mice often fails to follow the normal temporal and spatial pattern and to reach the same level as the endogenous copies1,2. Only in exceptional cases has position-independent and copy number-dependent expression been reproduced3. The size constraint of standard constructs may prevent the inclusion of important remote regulatory elements. Yeast artificial chromosomes (YACs)4 provide a means of cloning large DNA fragments and the transfer of YAC DNA into somatic cells has been reported5-9. We have previously produced transgenic mice carrying a 35 kilobase YAC construct10. Here we report the transfer of a 250 kilobase YAC covering the mouse tyrosinase gene into mice by pronuclear injection of gel-purified YAC DNA. The YAC was inserted into the mouse genome without major rearrangements and expression of the YAC-borne tyrosinase gene resulted in complete rescue of the albino phenotype of the recipient mice. Expression from the transgene reached levels comparable to that of the endogenous gene and showed copy number dependence and position independence.