The following are steps required to develop a knockout mouse. Place them in the

Question

The following are steps required to develop a knockout mouse. Place them in the correct order.

(1) Culture the cells in growth medium with G418 and gancyclovir to select those cells that have been transformed by homologous recombination.

(2) Implant the embryos in pseudopregnant mice.

(3) Interbreed resultant chimeric progeny to produce homozygous knockout mice.

(4) Disrupt the target gene with a neo+ marker, and link the tk+ marker to the target gene.

(5) Transfect mouse embryonic stem cells to transfer the disrupted gene (along with the adjacent tk+ marker) into the cells.

(6) Inject positive cells into early-stage mouse embryos.

Explanation / Answer

1.Disrupt the target gene with a neo+ marker, and link the tk+ marker to the target gene.
2.Transfect mouse embryonic stem cells to transfer the disrupted gene (along with the adjacent tk+ marker) into the cells.
3.Culture the cells in growth medium with G418 and gancyclovir to select those cells that have been transformed by homologous recombination.
4. Inject positive cells into early-stage mouse embryos.
5. Implant the embryos in pseudopregnant mice.
6. Interbreed resultant chimeric progeny to produce homozygous knockout mice.

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