What techniques can scientists use to determine if a particular transgene has been integrated into the genome of an organism?

One complication of making a transgenic animal is that the transgene may integrate at random into the coding region, or the regulatory region, of an endogenous gene. What might be the consequences of such random integrations? How might this complicate genetic analysis of the transgene?

A number of mouse models for human cystic fibrosis (CF) exist. Each of these mouse strains is transgenic and bears a different specific CFTR gene mutation. The mutations are the same as those seen in several varieties of human CF. These transgenic CF mice are being used to study the range of different phenotypes that characterize CF in humans. They are also used as models to test potential CF drugs. Unfortunately, most transgenic mouse CF strains do not show one of the most characteristic symptoms of human CF, that of lung congestion. Can you think of a reason why mouse CF strains do not display this symptom of human CF?

When disrupting a mouse gene by knockout, why is it desirable to breed mice until offspring homozygous (−/−) for the knockout target gene are obtained?

Craig Venter and others have constructed synthetic copies of viral genomes. For example, the genome for poliovirus and the 1918 influenza strain responsible for the pandemic flu have been assembled this way. The United States currently has a moratorium on federal funding for 'gain-of-function' experiments which increase the virulence or transmission potential of viruses. What concerns might ethicists have about synthetic biology studies involving potential pandemic pathogens?

Gene targeting and gene editing are both techniques for removing or modifying a particular gene, each of which can produce the same ultimate goal. What is the main technical difference in how DNA is modified that differs between these approaches?

How would you edit a specific nucleotide in a genome?

The CRISPR–Cas9 complex directs the Cas9 endonuclease to a specific genomic locus. If the endonuclease domain is inactivated and replaced with a transcriptional activator (or repressor) domain, what would be the functional consequence of directing such a complex to a specific chromosomal location?

How might you use CRISPR–Cas9 to create a large deletion?