Most of the time, loxp sites are inserted in introns around a critical exon. This should prevent interference with the orginal gene function. Once a mouse is made, you can test for the function of the gene product (Northern, PCR, or western) in the floxed/floxed cre neg animal.
Thanks Omai
Just one more for you I understand that the neo cassette can also alter gene function, causing a hypermorphic allele. is this true as well?
Look at the example papers below, in which the presence of a neo-cassette in the gene of SLC6A1, the neuronal GABA transporter 1, results in almost complete knockout of the transporter expression.
Sorry the reply took so long. The cross to a flipase mouse will remove the neocassette in the intron. Here are two examples, one from literature, one from Jackson lab.
Most of the time, loxp sites are inserted in introns around a critical exon. This should prevent interference with the orginal gene function. Once a mouse is made, you can test for the function of the gene product (Northern, PCR, or western) in the floxed/floxed cre neg animal.
Omai
Thanks Omai
Just one more for you I understand that the neo cassette can also alter gene function, causing a hypermorphic allele. is this true as well?
That is true
Look at the example papers below, in which the presence of a neo-cassette in the gene of SLC6A1, the neuronal GABA transporter 1, results in almost complete knockout of the transporter expression.
GABA transporter deficiency causes tremor, ataxia, nervousness, and increased GABA-induced tonic conductance in cerebellum.
Chiu CS, Brickley S, Jensen K, Southwell A, Mckinney S, Cull-Candy S, Mody I, Lester HA.
J Neurosci. 2005 Mar 23;25(12):3234-45.
PMID: 15788781
GABA transporter-1 (GAT1)-deficient mice: differential tonic activation of GABAA versus GABAB receptors in the hippocampus.
Jensen K, Chiu CS, Sokolova I, Lester HA, Mody I.
J Neurophysiol. 2003 Oct;90(4):2690-701. Epub 2003 Jun 18.
PMID: 12815026
Ok so if utilize a flipase cross to delete my frt-neo cassette then I can bypass these effects?
Hey Docmagus,
Sorry the reply took so long. The cross to a flipase mouse will remove the neocassette in the intron. Here are two examples, one from literature, one from Jackson lab.
www.pubmedcentral.nih.gov/articlerender.fcgi
jaxmice.jax.org/strain/009672.html
Happy mouse breeding.
Omai