Transgenic and Knock-Out Mice

Question 1

What is a transgenic (Tg) mouse

Answer 1

mouse with an additional gene introduced in fertilized egg randomly into its genome
(gene may be wild-type or normal, mutant, ot from another species)

Question 2

What are knockout (KO) mouse?

Answer 2

mouse in which a gene is specifically inactivated (targeted deletion)

Question 3

Special cases?

Answer 3

1. Inducible transgenes
2. conditional/tissue specific knockouts
3. knock ins

Question 4

Process of gene insertion in transgenic mice?

Answer 4

1. collect fertilized eggs
2. inject cloned DNA into pronuclei
3. implant injected eggs into oviduct of pregnant female
4. 10-30% of offspring contain transgene

Question 5

Uses of transgenic mice? Their caveats?

Answer 5

1. Study gene function -> overexpression can give misleading results (caveat)
2. Visualize protein expression patterns
   Caveat –> Disruption of different genes

Question 6

Process of knockout mice gene inactivation?

Answer 6

1. Use embryonic stem cells instead of fertilized eggs
2. introduce target gene construct to cultured stem cells
3. homologous and nonhomologous recombination occurs –> only want homologous recombinants in study bc homologous incorporates the target gene
4. Inject stem cells into embryo and then transfer embryo into pregnant mouse

Question 7

How do we get rid of recombinants without target gene?

Answer 7

treat with NEOMYCIN

Question 8

Uses of KO mice?

Answer 8

study function of non-essential gene

developmental genes

Question 9

Caveats of KO mice?

Answer 9

genes required for viability cannot be knocked out

expensive and time-consuming