| Autor: |
Gurskaia NG, Staroverov DB, Fradkov AF, Luk'ianov KA |
| Jazyk: |
ruština |
| Zdroj: |
Bioorganicheskaia khimiia [Bioorg Khim] 2011 May-Jun; Vol. 37 (3), pp. 425-8. |
| DOI: |
10.1134/s1068162011030071 |
| Abstrakt: |
Computer analysis predicted a strong donor splice site within the 3'-part of the far-red fluorescent protein Katushka coding region. To test the functional activity of this site a model vector has been constructed. This vector encoded Katushka and green fluorescent protein TagGFP2 with a gene fragment of tafazzin in between. Normal splicing of this pre-mRNA should result in a frameshift between Katushka and TagGFP2. Alternatively, after splicing at internal katushka donor splice site appearance of Katushka-TagGFP2 fusion protein was expected. Expression of this construct in a mammalian cell culture led to bright red and green fluorescence. Therefore, katushka-specific donor splice site is functional. Disruption of this splice site by several silent substitutions resulted in red-only fluorescent cells that corresponded to normal splicing. The mutant katushka can be used for visualization of pre-mRNA splicing at single cell level by fluorescence microscopy and flow cytometry. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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