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    GENETIC MODIFICATION OF RATS
    2.
    发明申请
    GENETIC MODIFICATION OF RATS 审中-公开
    RATS的遗传修饰

    公开(公告)号：WO2014130706A1

    公开(公告)日：2014-08-28

    申请号：PCT/US2014/017452

    申请日：2014-02-20

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： LEE, Jeffrey, D. ,  AUERBACH, Wojtek ,  HESLIN, David ,  FRENDEWEY, David ,  LAI, Ka-man Venus ,  VALENZUELA, David M.

    IPC: C12N5/07 ,  C12N5/10 ,  C12N5/00 ,  C12N5/02

    CPC classification number: C12N15/8509 ,  A01K67/02 ,  A01K67/0271 ,  A01K67/0275 ,  A01K2207/12 ,  A01K2217/15 ,  A01K2227/105 ,  A61D19/04 ,  C12N5/0606 ,  C12N15/8775 ,  C12N2501/235 ,  C12N2501/727 ,  C12N2501/999 ,  C12N2510/00

    Abstract: Compositions and methods are provided for making rat pluripotent and totipotent cells, including rat embryonic stem (ES) cells. Compositions and methods for improving efficiency or frequency of germline transmission of genetic modifications in rats are provided. Such methods and compositions comprise an in vitro culture comprising a feeder cell layer and a population of rat ES cells or a rat ES cell line, wherein the in vitro culture conditions maintain pluripotency of the ES cell and comprises a media having mouse leukemia inhibitor factor (LIF) or an active variant or fragment thereof. Various methods of establishing such rat ES cell lines are further provided. Methods of selecting genetically modified rat ES cells are also provided, along with various methods to generate a transgenic rat from the genetically modified rat ES cells provided herein. Various kits and articles of manufacture are further provided.

    Abstract translation: 提供组合物和方法用于制备大鼠多能全细胞，包括大鼠胚胎干（ES）细胞。 提供了用于提高大鼠遗传修饰的种系传播效率或频率的组合物和方法。 这些方法和组合物包括体外培养物，其包含饲养细胞层和大鼠ES细胞群或大鼠ES细胞系，其中体外培养条件保持ES细胞的多能性，并且包含具有小鼠白血病抑制因子（ LIF）或其活性变体或片段。 进一步提供了建立这种大鼠ES细胞系的各种方法。 还提供了选择遗传修饰的大鼠ES细胞的方法，以及从本文提供的遗传修饰的大鼠ES细胞产生转基因大鼠的各种方法。 还提供了各种套件和制品。

    SELECTION OF PLURIPOTENT CELLS FOR PRODUCTION OF FERTILE XY FEMALE MICE
    4.
    发明申请
    SELECTION OF PLURIPOTENT CELLS FOR PRODUCTION OF FERTILE XY FEMALE MICE 审中-公开
    选择用于生产可食用X射线母鼠的小鼠细胞

    公开(公告)号：WO2017049240A1

    公开(公告)日：2017-03-23

    申请号：PCT/US2016/052345

    申请日：2016-09-16

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： SCHMAHL, Jennifer ,  FRENDEWEY, David ,  KUNO, Junko ,  SIAO, Chia-Jen ,  DROGUETT, Gustavo ,  BAI, Yu ,  AUERBACH, Wojtek

    IPC: A01K67/027 ,  C12N5/0735

    CPC classification number: A01K67/0271 ,  A01K2207/12 ,  A01K2207/35 ,  A01K2227/105 ,  A01K2267/02 ,  C12N5/0606 ,  C12Q1/6876 ,  C12Q2600/136 ,  C12Q2600/156 ,  C12Q2600/158 ,  G01N33/5073

    Abstract: Methods and compositions are provided for generating F0 fertile XY female animals. The methods and compositions involve making XY pluripotent or totipotent animal cells, in vitro cell cultures, or embryos that are capable of producing a fertile female XY animal in an F0 generation. Such cells, embryos, and animals can be made by silencing a region of the Y chromosome. Optionally, the cells can also be cultured in feminizing medium such as a low-osmolality medium and/or can be modified to decrease the level and/or activity of an Sry protein. Methods and compositions are also provided for silencing a region of the Y chromosome in an XY pluripotent or totipotent animal cell, or in vitro cell cultures, embryos, or animals derived therefrom, by maintaining an XY pluripotent or totipotent animal cell in a feminizing medium. Methods and compositions are also provided for maintaining a population of XY pluripotent or totipotent animal cells in a feminizing medium and selecting cells or clones having increased capabilities for producing a fertile female XY animal in an F0 generation. Methods and compositions are also provided for screening for compounds with feminizing activity or for optimizing concentrations of components in feminizing media.

    Abstract translation: 提供了用于产生F0可育XY雌性动物的方法和组合物。 所述方法和组合物包括制备XY多能或全能动物细胞，体外细胞培养物或能够在F0代中产生可育雌性XY动物的胚胎。 可以通过沉默Y染色体的区域来制备这样的细胞，胚胎和动物。 任选地，细胞还可以在女性化培养基如低渗透浓度培养基中培养和/或可被修饰以降低Sry蛋白的水平和/或活性。 还提供了通过将XY多能或全能动物细胞维持在女性化培养基中，从而在XY多能或全能动物细胞或体外细胞培养物，胚胎或由其衍生的动物中沉默Y染色体区域的方法和组合物。 还提供了用于在女性化培养基中维持XY多能或全能动物细胞群的方法和组合物，并选择具有增加在F0代中产生可育雌性XY动物的能力的细胞或克隆。 还提供了用于筛选具有女性化活性的化合物或优化女性化培养基中组分浓度的方法和组合物。

    NUCLEASE-MEDIATED TARGETING WITH LARGE TARGETING VECTORS
    5.
    发明申请
    NUCLEASE-MEDIATED TARGETING WITH LARGE TARGETING VECTORS 审中-公开
    用大量目标矢量进行核心介导的瞄准

    公开(公告)号：WO2013163394A1

    公开(公告)日：2013-10-31

    申请号：PCT/US2013/038165

    申请日：2013-04-25

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： FRENDEWEY, David ,  AUERBACH, Wojtek ,  VALENZUELA, David, M. ,  YANCOPOULOS, George, D.

    IPC: C12N15/85 ,  C12N15/90 ,  C07K14/715

    CPC classification number: C12N15/85 ,  C07K14/7155 ,  C12N15/8509 ,  C12N15/907

    Abstract: Compositions and methods are provided for making one or more targeted genetic modifications at a target genomic locus by employing homologous recombination facilitated by single or double-strand break at or near the target genomic locus. Compositions and methods for promoting efficiency of homologous recombination between an LTVEC and a target genomic locus in prokaryotic or eukaryotic cells using engineered nucleases are also provided.

    Abstract translation: 提供组合物和方法，用于通过在靶基因组座位或其附近的单链或双链断裂促进的同源重组在靶基因组座位上进行一个或多个靶向遗传修饰。 还提供了使用工程化核酸酶在原核或真核细胞中促进LTVEC和靶基因组座位之间的同源重组效率的组合物和方法。

    SELECTION OF PLURIPOTENT CELLS FOR PRODUCTION OF FERTILE XY FEMALE MICE
    6.
    发明申请
    SELECTION OF PLURIPOTENT CELLS FOR PRODUCTION OF FERTILE XY FEMALE MICE 审中-公开

    公开(公告)号：WO2017049240A8

    公开(公告)日：2017-03-23

    申请号：PCT/US2016/052345

    申请日：2016-09-16

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： SCHMAHL, Jennifer ,  FRENDEWEY, David ,  KUNO, Junko ,  SIAO, Chia-Jen ,  DROGUETT, Gustavo ,  BAI, Yu ,  AUERBACH, Wojtek

    IPC: A01K67/027 ,  C12N5/0735

    Abstract: Methods and compositions are provided for generating F0 fertile XY female animals. The methods and compositions involve making XY pluripotent or totipotent animal cells, in vitro cell cultures, or embryos that are capable of producing a fertile female XY animal in an F0 generation. Such cells, embryos, and animals can be made by silencing a region of the Y chromosome. Optionally, the cells can also be cultured in feminizing medium such as a low-osmolality medium and/or can be modified to decrease the level and/or activity of an Sry protein. Methods and compositions are also provided for silencing a region of the Y chromosome in an XY pluripotent or totipotent animal cell, or in vitro cell cultures, embryos, or animals derived therefrom, by maintaining an XY pluripotent or totipotent animal cell in a feminizing medium. Methods and compositions are also provided for maintaining a population of XY pluripotent or totipotent animal cells in a feminizing medium and selecting cells or clones having increased capabilities for producing a fertile female XY animal in an F0 generation. Methods and compositions are also provided for screening for compounds with feminizing activity or for optimizing concentrations of components in feminizing media.

    METHODS AND COMPOSITIONS FOR MODIFYING A TARGETED LOCUS
    7.
    发明申请
    METHODS AND COMPOSITIONS FOR MODIFYING A TARGETED LOCUS 审中-公开
    用于修改目标位置的方法和组合

    公开(公告)号：WO2015188109A1

    公开(公告)日：2015-12-10

    申请号：PCT/US2015/034503

    申请日：2015-06-05

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： AUERBACH, Wojtek ,  FRENDEWEY, David ,  DROGUETT, Gustavo ,  GAGLIARDI, Anthony ,  KUNO, Junko ,  VALENZUELA, David M.

    IPC: C12N15/90

    CPC classification number: C12N15/907 ,  A01K67/0278 ,  A01K2217/072 ,  A01K2227/105 ,  A01K2267/0387 ,  C07K14/7051 ,  C12N15/64 ,  C12N2810/00

    Abstract: Methods and compositions are provided for modifying one or more target loci in a cell. Such methods comprise providing a cell comprising a first polynucleotide encoding a first selection marker operably linked to a first promoter active in the cell, wherein the first polynucleotide further comprises a first recognition site for a first nuclease agent. A first nuclease agent is introduced into a cell, wherein the first nuclease agent induces a nick or double-strand break at the first recognition site. Further introduced into the cell is a first targeting vector comprising a first insert polynucleotide flanked by a first and a second homology arm that correspond to a first and a second target site located in sufficient proximity to the first recognition site. At least one cell is then identified comprising in its genome the first insert polynucleotide integrated at the target locus.

    Abstract translation: 提供了用于修饰细胞中一个或多个靶基因座的方法和组合物。 这样的方法包括提供包含编码第一选择标记的第一多核苷酸的细胞，所述第一选择标记可操作地连接到细胞中活性的第一启动子，其中所述第一多核苷酸还包含第一核酸酶试剂的第一识别位点。 将第一核酸酶试剂引入细胞中，其中第一核酸酶试剂在第一识别位点诱导切口或双链断裂。 进一步引入细胞中的是第一靶向载体，其包含侧翼于第一和第二同源性臂的第一插入核苷酸，所述第一和第二同源性臂对应于位于第一识别位点附近的第一和第二靶位点。 然后鉴定至少一个细胞，其在其基因组中包含整合在靶标位点的第一插入多核苷酸。

    TARGETED MODIFICATION OF RAT GENOME
    8.
    发明申请
    TARGETED MODIFICATION OF RAT GENOME 审中-公开
    大鼠基因组的靶向修饰

    公开(公告)号：WO2014172489A2

    公开(公告)日：2014-10-23

    申请号：PCT/US2014/034412

    申请日：2014-04-16

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： LEE, Jeffrey D. ,  MUJICA, Alexander, O. ,  AUERBACH, Wojtek ,  LAI, Ka-man Venus ,  VALENZUELA, David M. ,  YANCOPOULOS, George D.

    IPC: A01K67/027 ,  C12N15/85

    CPC classification number: C12N15/8509 ,  A01K67/0276 ,  A01K67/0278 ,  A01K2217/07 ,  A01K2227/105 ,  A01K2267/0362 ,  A01K2267/0381 ,  A61D19/04 ,  C07K14/7155 ,  C07K14/775 ,  C12N15/907 ,  C12N2015/8527 ,  C12N2800/30 ,  C12N2810/00

    Abstract: Compositions and methods are provided for modifying a rat genomic locus of interest using a large targeting vector (LTVEC) comprising various endogenous or exogenous nucleic acid sequences as described herein. Compositions and methods for generating a genetically modified rat comprising one or more targeted genetic modifications in their germline are also provided. Compositions and methods are provided which comprise a genetically modified rat or rat cell comprising a targeted genetic modification in the rat interleukin-2 receptor gamma locus, the rat ApoE locus, the rat Rag2 locus, the rat Rag1 locus and/or the rat Rag2/Rag1 locus. The various methods and compositions provided herein allows for these modified loci to be transmitted through the germline.

    Abstract translation: 提供了使用包含如本文所述的各种内源或外源核酸序列的大靶向载体（LTVEC）修饰感兴趣的大鼠基因组基因座的组合物和方法。 还提供了用于产生在其种系中包含一种或多种靶向遗传修饰的遗传修饰的大鼠的组合物和方法。 提供了组合物和方法，其包含在大鼠白细胞介素-2受体γ基因座，大鼠ApoE基因座，大鼠Rag2基因座，大鼠Rag1基因座和/或大鼠Rag2基因座中包含靶向遗传修饰的基因修饰的大鼠或大鼠细胞。 Rag1基因座。 本文提供的各种方法和组合物允许这些修饰的基因座通过种系传播。

    METHODS AND COMPOSITIONS FOR GENERATING OR MAINTAINING PLURIPOTENT CELLS
    10.
    发明申请
    METHODS AND COMPOSITIONS FOR GENERATING OR MAINTAINING PLURIPOTENT CELLS 审中-公开
    用于产生或维持PLUIPOTENT细胞的方法和组合物

    公开(公告)号：WO2016061374A1

    公开(公告)日：2016-04-21

    申请号：PCT/US2015/055776

    申请日：2015-10-15

    Applicant: REGENERON PHARMACEUTICALS, INC.

    Inventor： KUNO, Junko ,  AUERBACH, Wojtek ,  VALENZUELA, David M.

    IPC: C12N5/074 ,  C12N5/00

    CPC classification number: C12N5/0696 ,  C12N2500/32 ,  C12N2500/38 ,  C12N2500/44 ,  C12N2500/60 ,  C12N2501/235 ,  C12N2501/727 ,  C12N2509/00

    Abstract: Methods and compositions are provided for generating or maintaining human iPS cells in culture. Methods include the use of a low osmolality medium to make human iPS cells, or use of a low osmolality medium to maintain human iPS cells. Methods for making targeted genetic modification to human iPS cells cultured in low osmolality medium are also included. Compositions include human iPS cells cultured and maintained using the low osmolality medium defined herein.

    Abstract translation: 提供了用于在培养物中产生或维持人iPS细胞的方法和组合物。 方法包括使用低渗透浓度培养基制备人iPS细胞，或使用低渗透浓度培养基来维持人类iPS细胞。 还包括对在低渗透浓度培养基中培养的人iPS细胞进行靶向遗传修饰的方法。 组合物包括使用本文定义的低渗透浓度培养基培养和维持的人iPS细胞。

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