Question

Your student is trying to create a "Gene X" knockout mouse by introducing a recombinant construct into embryonic stem (ES) cells, selecting transformants, and introducing the transformed cells into mouse blastocysts. He designed his construct as follows: NeoR region of homology to Gene X---HSV tk--- region of homology to Gene X where Neo is the neomycin-resistance gene and HSV tk is the Herpes Simplex virus thymidine kinase gene. He transforms the ES cells and exposes them to G418 and ganciclovir. Explain to your student why he failed to recover transformed ES cells. Choose the two correct statements. a. His construct incorporated into the mouse DNA by illegitimate recombination. These cells would be sensitive to ganciclovir and neomycin, and would die.

b. His construct incorporated into the mouse DNA by illegitimate recombination. These cells would be resistant to ganciclovir, but sensitive to neomycin, and would die.
c. His construct incorporated into the mouse DNA by homologous recombination. These cells would be sensitive to ganciclovir, but resistant to neomycin, and would die.
d. His construct incorporated into the mouse DNA by homologous recombination. These cells would be sensitive to neomycin and ganciclovir, and would die.
e. His construct incorporated into the mouse DNA by homologous recombination. These cells would be sensitive to neomycin, but resistant to ganciclovir, and would die.
f. His construct incorporated into the mouse DNA by illegitimate recombination. These cells would be resistant to neomycin, but sensitive to ganciclovir, and would die.

Answer 1

The student failed to recover transformed ES cells because the construct incorporated into the mouse DNA by either homologous recombination or illegitimate recombination.

Step-by-step explanation:

The correct statements for why the student failed to recover transformed ES cells are:

c. His construct incorporated into the mouse DNA by homologous recombination. These cells would be sensitive to ganciclovir, but resistant to neomycin, and would die.

f. His construct incorporated into the mouse DNA by illegitimate recombination. These cells would be resistant to neomycin, but sensitive to ganciclovir, and would die.

When the construct is incorporated into the mouse DNA by homologous recombination, the cells would be sensitive to ganciclovir, but resistant to neomycin, and would die. On the other hand, if the construct is incorporated by illegitimate recombination, the cells would be resistant to neomycin, but sensitive to ganciclovir, and would die.