- Function research of genes
- Deletion of specific groups of cells
Homozygous mice for the targeted mutation are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. B-ROSA26-EGFP-DTA mice have STOP cassettes preventing expression of diphtheria toxin fragment A (DTA). DTA expression will be activated, if STOP cassettes are removed – resulting in the specific ablation of the cre-expressing cells. ROSA26-EGFP-DTA mice are useful for Cre-inducible deletion of specific groups of cells.
Gene editing strategy
The ROSA26-EGFP-DTA allele has a frt-flanked STOP sequence (2xSV40 pA-lox2 stop) and a loxP-flanked STOP sequence (EGFP-Neo-3xpolyA) upstream of the diphtheria toxin A subunit (DTA) gene. After removal of the flanked STOP cassettes (2xSV40 pA-lox2 stop) by FLP-recombination, mutant mice display widespread expression of EGFP, but DTA transcription is still prevented by the strong transcriptional stop sequence(EGFP-Neo-3xpolyA). When bred to mice that express Cre recombinase under the control of a promoter of interest, the floxed-STOP cassette is removed and DTA expression is activated – resulting in the specific ablation of the cre-expressing cells. Homozygous mice are viable, normal in size, and do not display any gross physical or behavioral abnormalities. The donating investigator reports that some homozygous males are subfertile.
ROSA26-EGFP-DTA mice may be useful on their own as a fluorescent reporter, or in combination with cre-expressing mice to produce conditional deletions of specific groups of cells. ROSA26-EGFP-DTA mice may also have applications in toxicology and protein synthesis research.
B cell from spleen, Flow cytometry analysis showed that the number of B cells specifically expressing Cre recombinase was significantly reduced in single cells of spleen, which proved that B cells specifically expressed DTA, and DTA successfully cleared B cells in spleen.
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