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Q4_final - For instance there are large differences in...

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3269557 QUESTION 4 This question presupposes that we already know the mRNA/amino acid sequence of the receptor. Create a complimentary mRNA sequence and conjugate it to the beads of a cellulose chromatography column. Running a homogenized, tissue specific, region of the brain through this column would bind the receptors mRNA due to the complimentary sequences. The first eluent would not be our mRNA of interest and could be discarded. The second eluent contains the mRNA of interest. Using reverse transcriptase we can create a cDNA sequence and inject it into a frog oocyte. I am assuming that this process only encodes the protein of interest, although, expression of other proteins is not completely out of the question. Experimental Characterization of the Receptor – ADD MORE Depending on the relative subunit contributions, electrophysiological and pharmacological studies will reveal different things.
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Unformatted text preview: For instance, there are large differences in calcium permeabilities depending on the neuronal nAChRs subunit constitution. These channels are also selective for cations and must be expressed with both α and β subunits to be functional. By applying different chemicals you can elucidate which type of agonists/antagonists bind the receptor. Using inside-out/outside-out patches, you could determine stoichiometric relationships of ion flux, and permeability. Using a voltage-clamp you could determine any sort of voltage-dependence. Member of the cys-loop family? Comparison of our derived cDNA sequence to that of genetic databases will reveal similarities between out receptor of interest and know cys-loop family receptors. The M2 domain is evolutionarily conserved in glycine, GABA A, and nAChRs, and would presumably show up in a novel receptor as well....
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