DNA encoding the actin binding domain of Drosophila moesin and the SV40 terminator sequence were cloned by PCR and inserted into NotI-BglII and BglII-EcoRI sites of pCASPER2 respectively. 1160bp of Drosophila genomic DNA immediately upstream of the patched coding region was fused to DNA encoding eGFP by PCR and the resulting fragment was cloned into pCR4TOPO (Invitrogen) according to the manufacturer's instructions. Genomic DNA from 11200 to 1160bp upstream of ptc was then inserted upstream of this as a BamHI-NdeI fragment. Finally, the complete patched-GFP fragment was inserted into pCASPER2 upstream of moesin as a BamHI-NotI fragment.
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Constructing Patched-GFP Fusion Reporter
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Other organizations : University of Bristol, Gènes, synapses et cognition
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- PCR cloning of DNA encoding the actin binding domain of Drosophila moesin and the SV40 terminator sequence
- PCR fusion of 1160bp of Drosophila genomic DNA upstream of the patched coding region to DNA encoding eGFP
- Insertion of genomic DNA from 11200 to 1160bp upstream of ptc as a BamHI-NdeI fragment
- Expression of the patched-GFP fusion construct
- Not explicitly mentioned
- Not explicitly mentioned
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