The Na(v) channel bench series: Plasmid preparation()

Research involving recombinant voltage-gated sodium (Na(v)) channels has unique challenges. Multiple factors contribute, but undoubtedly at the top of the list is these channels’ DNA instability. Once introduced into bacterial hosts, Na(v) channel plasmid DNA will almost invariably emerge mutagenized and unusable, unless special conditions are adopted. This is particularly true for Na(v)1.1 (gene name SCN1A), Na(v)1.2 (SCN2A), and Na(v)1.6 (SCN8A), but less so for Na(v)1.4 (SCN4A) and Na(v)1.5 (SCN5A) while other Na(v) channel isoforms such as Na(v)1.7 (SCN9A) lie in between. The following recommendations for Na(v) • Bacterial propagation using Stbl2 cells at or below 30 °C. • Bias toward slow-growing, small bacterial colonies. • Comprehensive sequencing of the entire Na(v) channel coding region.