Final answer:
When creating transgenic mouse lines via homologous recombination, the selection for positive or negative Gene of Interest (GoI) integration is typically achieved through the use of antibiotic resistance genes. Positive selection involves introducing a selectable marker, such as an antibiotic resistance gene, within the construct containing the GoI. Negative selection, on the other hand, employs counter-selectable markers like thymidine kinase to eliminate unwanted integration events.
Step-by-step explanation:
Positive selection is a crucial step in the creation of transgenic mouse lines via homologous recombination. It involves introducing a selectable marker, often an antibiotic resistance gene, along with the Gene of Interest (GoI) within the gene construct.
For example, a neomycin resistance gene (neo) can be incorporated into the construct. Cells that successfully integrate the construct into their genome will express the antibiotic resistance gene, allowing them to survive in the presence of the corresponding antibiotic. This positive selection ensures that only cells with the desired GoI integration are able to proliferate and contribute to the development of the transgenic mouse.
Negative selection is employed to eliminate cells with random or non-homologous integrations, enhancing the specificity of the process. A counter-selectable marker, such as the herpes simplex virus thymidine kinase gene (tk), can be included in the construct.
Cells expressing this marker are sensitive to certain drugs like ganciclovir. During negative selection, cells with unwanted integrations are eliminated when exposed to the counter-selective drug, leaving behind only those with the desired homologous recombination of the GoI.
In conclusion, the combination of positive and negative selection strategies, utilizing antibiotic resistance and counter-selectable markers, respectively, facilitates the precise identification and isolation of cells with the intended Gene of Interest integration during the creation of transgenic mouse lines via homologous recombination. These selection methods are essential for obtaining transgenic mice with accurate and controlled genomic modifications.