Gfp fusion proteins and their use
a technology of fusion proteins and fusion proteins, applied in the field of gfp fusion proteins and their, can solve the problems of difficult to prove and not give results reflective of the natural sta
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[0029]Construction of Gαs-GFP
[0030]Full length cDNAs encoding Gαs were excised from the PcDNA-1 vector by digesting with Sam I and Xba I restriction enzymes. The full length EGFP cDNA was obtained by PCR from the PEGFP-N3 using appropriate primers (sense 5′ GGAATTCATGAGCAAGGGCGAGGAACTG-3′ (SEQ ID NO: 8); antisense 5′-GCTCTAGACGACTTGTACAGCTCGT-3′) (SEQ ID NO: 9) and adding restriction sites to its cDNA (EcoR I at the initiation codon and Xba I at end of cDNA). To insert the EGFP within the sequence of Gαs, the first fragment of Gαs (from 1 to 71 amino acids) was amplified by PCR with restriction sites for Kap 1 at initiation codon and EcoR I at end of the fragment. The cDNA of the fragment was cloned into PcDNA3 vector by the Kap 1 and EcoR 1 restriction sites using primers (sense 5′GGGTACCATGGGCTGCCTCGGCAACA-3′ (SEQ ID NO: 10); antisense 5′-GGAATTCGTCCTCTTCGCCGCCCTTCT-3′) (SEQ ID NO: 11). Modified 7 EGFP cDNA was spliced into the first fragment of Gαs by EcoR 1 and Xba 1 restriction...
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