公开(公告)号：US20130212725A1

公开(公告)日：2013-08-15

申请号：US13702231

申请日：2011-06-07

申请人： Ralf Kühn ,  Wolfgang Wurst ,  Melanie Meyer

发明人： Ralf Kühn ,  Wolfgang Wurst ,  Melanie Meyer

IPC分类号： C12N15/62

CPC分类号： C12N15/62 ,  A01K67/0276 ,  A01K67/0278 ,  A01K2207/05 ,  A01K2217/07 ,  A01K2217/072 ,  A01K2227/105 ,  A01K2267/03 ,  A01K2267/0393 ,  C07K2319/80 ,  C12N9/22 ,  C12N15/907

摘要： The present invention relates to a method of modifying a target sequence in the genome of a eukaryotic cell, the method comprising the step: (a) introducing into the cell a fusion protein comprising a DNA-binding domain of a Tal effector protein and a non-specific cleavage domain of a restriction nuclease or a nucleic acid molecule encoding the fusion protein in expressible form, wherein the fusion protein specifically binds within the target sequence and introduces a double strand break within the target sequence. The present invention further relates to the method of the invention, wherein the modification of the target sequence is by homologous recombination with a donor nucleic acid sequence further comprising the step: (b) introducing a nucleic acid molecule into the cell, wherein the nucleic acid molecule comprises the donor nucleic acid sequence and regions homologous to the target sequence. The present invention also relates to a method of producing a non-human mammal or vertebrate carrying a modified target sequence in its genome. Furthermore, the present invention relates to a fusion protein comprising a DNA-binding domain of a Tal effector protein and a non-specific cleavage domain of a restriction nuclease.

摘要翻译： 本发明涉及一种修饰真核细胞基因组中的靶序列的方法，所述方法包括以下步骤：（a）将包含Tal效应蛋白的DNA结合结构域的融合蛋白引入细胞， 限制性核酸酶或编码可表达形式的融合蛋白的核酸分子的特异性切割结构域，其中所述融合蛋白在靶序列内特异性结合并在靶序列内引入双链断裂。 本发明还涉及本发明的方法，其中靶序列的修饰是通过与供体核酸序列的同源重组而进一步包括步骤：（b）将核酸分子引入细胞，其中核酸 分子包含供体核酸序列和与靶序列同源的区域。 本发明还涉及在其基因组中产生携带修饰的靶序列的非人哺乳动物或脊椎动物的方法。 此外，本发明涉及包含Tal效应蛋白的DNA结合结构域和限制性核酸酶的非特异性切割结构域的融合蛋白。

公开(公告)号：US20120276537A1

公开(公告)日：2012-11-01

申请号：US13504587

申请日：2010-10-28

申请人： Ralf Kühn ,  Wolfgang Wurst ,  Melanie Meyer

发明人： Ralf Kühn ,  Wolfgang Wurst ,  Melanie Meyer

IPC分类号： C12N15/90 ,  C12Q1/68 ,  C12N15/89

CPC分类号： A01K67/0278 ,  A01K2217/072 ,  A01K2227/105 ,  A01K2267/01 ,  C12N9/22 ,  C12N15/8509 ,  C12N15/907

摘要： The present invention relates to a method of modifying a target sequence in the genome of a mammalian or avian oocyte by homologous recombination with a donor nucleic acid sequence, the method comprising the steps (a) introducing into the oocyte a zinc finger nuclease or a nucleic acid molecule encoding the zinc finger nuclease in expressible form, wherein the zinc finger nuclease specifically binds within the target sequence and introduces a double strand break within the target sequence; and (b) introducing a nucleic acid molecule into the oocyte, wherein the nucleic acid molecule comprises the donor nucleic acid sequence and regions homologous to the target sequence. The present invention further relates to a method of producing a non-human mammal or an avian carrying a modified target sequence in its genome.

摘要翻译： 本发明涉及通过与供体核酸序列同源重组来修饰哺乳动物或禽类卵母细胞基因组中的靶序列的方法，所述方法包括以下步骤：（a）向卵母细胞中引入锌指核酸酶或核酸 编码锌指核酸酶的可分解形式的酸分子，其中所述锌指核酸酶在靶序列内特异性结合并在靶序列内引入双链断裂; 和（b）将核酸分子引入所述卵母细胞中，其中所述核酸分子包含供体核酸序列和与靶序列同源的区域。 本发明还涉及在其基因组中产生携带修饰的靶序列的非人哺乳动物或禽类的方法。