Production of fertile XY female mice from XY mouse ES cells
First Claim
1. A method for increasing the efficiency of generating embryonic stem (ES) cell-derived mice in an F0 generation, comprising:
- (a) maintaining a donor XY mouse ES cell in a medium comprising;
(i) a base medium; and
(ii) supplements suitable for growing the mouse ES cells in culture and maintaining pluripotency,wherein the base medium comprises sodium bicarbonate in a concentration of 1.5-2.2 mg/mL, comprises fetal bovine serum, and has an osmolality of 218-322 mOsm/kg;
(b) injecting a donor XY mouse ES cell from step (a) into a pre-morula stage host mouse embryo;
(c) introducing the host mouse embryo of step (b) into a recipient female mouse and gestating the host mouse embryo; and
(d) obtaining F0 XY mouse progeny, wherein the ratio of ES-cell-derived pups to total pups generated in the F0 generation is greater than the ratio of ES-cell-derived pups to total pups generated in the F0 generation produced from donor XY ES cells maintained in a medium having an osmolality of greater than 322 mOsm/kg.
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Abstract
Methods and compositions are described for making phenotypically female fertile animals from XY donor cells and suitable host embryos. Culture media and methods are provided for maintaining XY donor cells in culture that after introduction into a host embryo and gestation in a suitable host will result in fertile XY female animals. Methods and compositions are described for making fertile female animals in an F0 generation from a donor XY cell and a host embryo, as are methods for making F1 progeny that are homozygous for a modification from a heterozygous F0 fertile male and a heterozygous F0 fertile female sibling.
20 Citations
26 Claims
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1. A method for increasing the efficiency of generating embryonic stem (ES) cell-derived mice in an F0 generation, comprising:
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(a) maintaining a donor XY mouse ES cell in a medium comprising; (i) a base medium; and (ii) supplements suitable for growing the mouse ES cells in culture and maintaining pluripotency, wherein the base medium comprises sodium bicarbonate in a concentration of 1.5-2.2 mg/mL, comprises fetal bovine serum, and has an osmolality of 218-322 mOsm/kg; (b) injecting a donor XY mouse ES cell from step (a) into a pre-morula stage host mouse embryo; (c) introducing the host mouse embryo of step (b) into a recipient female mouse and gestating the host mouse embryo; and (d) obtaining F0 XY mouse progeny, wherein the ratio of ES-cell-derived pups to total pups generated in the F0 generation is greater than the ratio of ES-cell-derived pups to total pups generated in the F0 generation produced from donor XY ES cells maintained in a medium having an osmolality of greater than 322 mOsm/kg. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26)
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Specification