Mouse lambda light chain locus
AssigneeTranslocus Limited
InventorsMARIANNE BRUGGEMANN, XIANGANG ZOU
Life of this patent
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Abstract
The present invention provides in a first aspect a mouse in which the λ (lambda) light chain locus has been functionally silenced. In one embodiment, the mouse λ light chain locus was functional silenced by deletion of gene segments coding for the λ light chain locus. In a further aspect, a mouse containing functionally silenced λ and κ (kappa) L chain loci was produced. The invention is useful for the production of antibodies, for example heterologous antibodies, including heavy chain only antibodies.
Claims39
A knock-out mouse having its lambda light chain locus C2-C4-C3-C1 deleted using LoxP constructs as the targeting constructs and ubiquitous Cre-expresser mice, wherein the knock-out mouse is produced by a method comprising the steps of:
- (a) introducing at least one C3-C1 targeting construct that comprises LoxP sequences to a mouse embryonic stem (ES) cell, thereby producing a mouse ES cell having silenced C1;
- (b) introducing the mouse ES cell having silenced C1 to a mouse embryo in order to produce a mouse having silenced C1 by germline transmission;
- (c) breeding the mouse having silenced C1 with a Cre-expresser mouse, which causes targeted deletion of C3-C1;
- (d) obtaining a C3-C1 deleted mouse as a result of the breeding of (c);
- (e) obtaining a mouse ES cell from the C3-C1 deleted mouse;
- (f) introducing a C2-C4 targeting construct that comprises a LoxP sequence to the mouse ES cell from the C3-C1 deleted mouse, thereby producing a mouse ES cell having silenced C2;
- (g) introducing the mouse ES cell having silenced C2 to a mouse embryo in order to produce a mouse having silenced C2 by germline transmission;
- (h) breeding the mouse having silenced C2 with a Cre-expresser mouse, which causes targeted deletion of C2-C4;
- and
- (i) obtaining a knock-out mouse having deleted lambda locus C2-C4-C3-C1 as a result of the breeding of (h).
A method of producing a knock-out mouse, wherein lambda light chain locus is deleted through knock-out by targeted integration and gene deletion using LoxP constructs as the targeting constructs and ubiquitous Cre-expresser mice, wherein the method comprises the step of deleting at least the constant region genes C1, C2 and C3 of the lambda light chain locus.
A method of making a knock-out mouse having its lambda light chain locus deleted, wherein the method comprises the steps of:
- (a) introducing at least one C2-C4 targeting construct to a mouse embryonic stem (ES) cell, thereby producing an ES cell having silenced C2 region;
- (b) introducing the ES cell having the silenced C2 region to a mouse embryo in order to produce a mouse having silenced C2 region by germline transmission;
- (c) obtaining a C2 deleted mouse by breeding the mouse of step (b) with another mouse capable of causing deletion of the targeted C2-C4 region by germline transmission;
- (d) obtaining ES cells from a progeny mouse of step (c);
- (e) introducing a C3-C1 targeting construct to the ES cell of step (d), thereby producing an ES cell having silenced C1 region;
- (f) introducing the ES cell of step e) having the silenced C1 region to a mouse embryo in order to produce a mouse having silenced C1 region by germline transmission;
- and
- (g) obtaining a mouse having deleted lambda locus regions C2-C4-C3-C1 by breeding the mouse of step (f) with another mouse capable of causing deletion of the targeted C3-C1 region in germline transmission.
A knock-out mouse in which the lambda light chain locus is deleted through knock-out by targeted integration and gene deletion, wherein the knock-out of the lambda light chain locus is carried out by a method comprising the steps of:
- (a) introducing at least one C2-C4 targeting construct to a mouse embryonic stem (ES) cell, thereby producing an ES cell having silenced C2 region;
- (b) introducing the ES cell having the silenced C2 region to a mouse embryo in order to produce a mouse having silenced C2 region by germline transmission;
- (c) obtaining a C2 deleted mouse by breeding the mouse of step (b) with another mouse capable of causing deletion of the targeted C2-C4 region by germline transmission;
- (d) obtaining ES cells from a progeny mouse of step (c);
- (e) introducing a C3-C1 targeting construct to the ES cell of step (d), thereby producing an ES cell having silenced C1 region;
- (f) introducing the ES cell of step (e) having the silenced C1 region to a mouse embryo in order to produce a mouse having silenced C1 region by germline transmission;
- and
- (g) obtaining a mouse having deleted lambda locus regions C2-C4-C3-C1 by breeding the mouse of step (f) with another mouse capable of causing deletion of the targeted C3-C1 region in germline transmission.
A double knock-out mouse that has both the lambda light chain and the κ light chain loci functionally silenced by gene targeting of the κ light chain locus, targeted integration, gene deletion, homologous integration of a selectable marker gene in Cκ or targeted removal of Cκ or Jκ, wherein the double knock-out mouse is produced by breeding (1) a knock-out mouse having its κ light chain locus functionally silenced with (2) a knock-out mouse having its lambda light chain locus C2-C4-C3-C1 deleted by:
- (a) introducing at least one C3-C1 targeting construct that comprises LoxP sequences to a mouse embryonic stem (ES) cell, thereby producing a mouse ES cell having silenced C1;
- (b) introducing the mouse ES cell having silenced C1 to a mouse embryo in order to produce a mouse having silenced C1 by germline transmission;
- (c) breeding the mouse having silenced C1 with a Cre-expresser mouse, which causes targeted deletion of C3-C1;
- (d) obtaining a C3-C1 deleted mouse as a result of the breeding of (c);
- (e) obtaining a mouse ES cell from the C3-C1 deleted mouse;
- (f) introducing a C2-C4 targeting construct that comprises a LoxP sequence to the mouse ES cell from the C3-C1 deleted mouse, thereby producing a mouse ES cell having silenced C2;
- (g) introducing the mouse ES cell having silenced C2 to a mouse embryo in order to produce a mouse having silenced C2 by germline transmission;
- (h) breeding the mouse having silenced C2 with a Cre-expresser mouse, which causes targeted deletion of C2-C4;
- and
- (i) obtaining a knock-out mouse having deleted lambda locus C2-C4-C3-C1 as a result of the breeding of (h);
- wherein the breeding of (1) the knock-out mouse having its κ light chain locus functionally silenced with (2) the knock-out mouse having its lambda light chain locus C2-C4-C3-C1 deleted provides a progeny double knock-out mouse that has both the lambda light chain and the κ light chain loci deleted.
A double knock-out mouse in which both the lambda light chain and the κ light chain loci are functionally silenced by gene targeting of the κ light chain locus, targeted integration, gene deletion, homologous integration of a selectable marker gene in Cκ or targeted removal of Cκ or Jκ, wherein the mouse is produced by
- (I) breeding (1) a knock-out mouse having its κ light chain locus functionally silenced with (2) a knock-out mouse having its lambda light chain locus C2-C4-C3-C1 deleted by:
- (a) introducing at least one C3-C1 targeting construct that comprises LoxP sequences to a mouse embryonic stem (ES) cell, thereby producing a mouse ES cell having silenced C1;
- (b) introducing the mouse ES cell having silenced C1 to a mouse embryo in order to produce a mouse having silenced C1 by germline transmission;
- (c) breeding the mouse having silenced C1 with a Cre-expresser mouse, which causes targeted deletion of C3-C1;
- (d) obtaining a C3-C1 deleted mouse as a result of the breeding of (c);
- (e) obtaining a mouse ES cell from the C3-C1 deleted mouse;
- (f) introducing a C2-C4 targeting construct that comprises a LoxP sequence to the mouse ES cell from the C3-C1 deleted mouse, thereby producing a mouse ES cell having silenced C2;
- (g) introducing the mouse ES cell having silenced C2 to a mouse embryo in order to produce a mouse having silenced C2 by germline transmission;
- (h) breeding the mouse having silenced C2 with a Cre-expresser mouse, which causes targeted deletion of C2-C4;
- and
- (i) obtaining a knock-out mouse having deleted lambda locus C2-C4-C3-C1 as a result of the breeding of (h);
- and
- (II) obtaining a progeny double knock-out mouse that has both the lambda light chain and the κ light chain loci deleted by the breeding of (I).
Dependencies are inferred from the claim text.