Download Ion Channels: Methods and Protocols by James D. Stockand, Mark S. Shapiro PDF

By James D. Stockand, Mark S. Shapiro

The varied purposes during this quantity diversity from the examine of allosteric rules of ion channel task utilizing a vintage mutagenesis strategy, to the learn of channel subunit stoichiometry utilizing a singular biophysical procedure according to fluorescence resonance strength move. Highlights comprise tools for heterologous expression of ion channels in cells, for making a choice on channel structure-function, and for learning channel rules.

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Extra resources for Ion Channels: Methods and Protocols

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Phenol/chloroform/isoamyl alcohol 25:24:1 (Sigma Fluka, St. Louis, MO; use caution as this mix is caustic). 8. Chloroform/isoamyl alcohol 49:1 (Sigma Fluka). 9. Pellet Paint coprecipitant (Novagen, Madison, WI). 10. Millipore UltraFree FilterSpin columns (Fisher). 11. Rapid DNA Ligation Kit (Roche). 12. One Shot competent Escherichia coli (Invitrogen). 13. Luria-Bertani (LB) medium. 14. Bacto Agar for plates (BD Biosciences, Clontech). 15. LB with kanamycin (KAN; 50 µg/mL). 16. KAN plates: LB agar plates with kanamycin (50 µg/mL).

Beech, D. , Shapiro, M. , and Wollmuth, 1. L. P. (1995) Multiple G-protein-coupled pathways inhibit N-type Ca2+ channels of neurons. Life Sci. 56, 989–992. 2 Gamper, N. and Shapiro, M. S. (2003) Calmodulin mediates Ca 2+-dependent 2. modulation of M-type K+ channels. J. Gen. Physiol. 122, 17–31. , Stockand, J. , and Shapiro, M. S. (2003) Subunit-specific modula3. tion of KCNQ potassium channels by Src tyrosine kinase. J. Neurosci. 23, 84–95. , and Shapiro, M. S. (2004) Phos4. phatidylinositol 4,5-bisphosphate signals underlie receptor-specific Gq/11-mediated modulation of N-type Ca2+ channels.

14. Bacto Agar for plates (BD Biosciences, Clontech). 15. LB with kanamycin (KAN; 50 µg/mL). 16. KAN plates: LB agar plates with kanamycin (50 µg/mL). 17. QIAprep Spin Miniprep Kit (Qiagen). 3. Subcloning of Complementary DNA for POI-IRES-EGFP Into pSFV 1. 2. 2. pSFV and pSFVHelper2 vectors (Invitrogen; see Note 1). 3. Guanosine 5'-triphosphate (GTP)/cytidine 5'-triphosphate (CTP) mix: Mix of 1 mM each from PCR grade 100 mM dNTP set (Invitrogen) in MBG water. 4. Large (Klenow) fragment DNA polymerase I (Klenow; New England Biolabs).

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