A kind of polypeptide and its application in the preparation of voltage-sensitive sodium channel inhibitor
A sodium channel inhibitor, voltage-sensitive technology, applied in the biological field, can solve the problems of long cycle of natural polypeptides
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[0014] 1. Yeast two-hybrid method to obtain positive clones, and self-activation verification
[0015] The yeast two-hybrid experiment was carried out using the MATCHMAKER GAL4Two-Hybrid System3 system of clontech company. The specific steps are: 1) will contain hNa v The S3-S4 extracellular linking loop of 1.5 channel Domain II (hNa v 1. The bait plasmid (BD) of 5-DII-S3-S4) was transformed into yeast strain AH109, and it was prepared to be competent by PEG / LiAc method; Competent; 3) Finally, spread the competent cells on the growth-defective medium and culture at 30°C until the positive clones grow; 4) Small-scale expansion culture of the positive clones, extraction of plasmids, transfer into AH109, and smear on Repeat the experiment and self-activation verification on a suitable defective medium; 5) Send the positive plasmid without self-activation to the sequencing company for sequencing.
[0016] Through the above steps, we found that 3 library plasmids can interac...
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