Simple and efficient site-directed mutagenesis using two single-primer reactions in parallel to generate mutants for protein structure-function studies

Table 1 Forward (-F) and reverse (-R) primers used for point mutations in α ENaC (a-), and insertion of a 27 nt segment coding for a 9 residue epitope tag in β ENaC (b+) cDNA.

Primer	Sequence*	Length (nt)
a-R333K-F	ctgtccctgatgctgAAGgcagagcagaatgacttc	36
a-R333K-R	gaagtcattctgctctgcCTTcagcatcagggacag	36
a-D338A-F	cgcgcagagcagaatgCcttcattcccctgctg	33
a-D338A-R	cagcaggggaatgaagGcattctgctctgcgcg	33
a-R350A-F	cacagtgactggggccGCggtaatggtgcacggg	34
a-R350A-R	cccgtgcaccattaccGCggccccagtcactgtg	34
a-E358D-F	gtgcacgggcaggatgaTcctgcctttatggatg	34
a-E358D-R	catccataaaggcaggAtcatcctgcccgtgcac	34
a-K474R-F	ctgggctgtttcaccaGgtgccggaagccatgc	33
a-K474R-R	gcatggcttccggcacCtggtgaaacagcccag	33
a-R476K-F	ctgtttcaccaagtgcAAgaagccatgcagcgtg	34
a-R476K-R	cacgctgcatggcttcTTgcacttggtgaaacag	34
b+HA-F	gccattgccaccaggTACCCATACGACGTCCCAGACTACGCTaacctgaacttctcc	57
b+HA-R	ggagaagttcaggttAGCGTAGTCTGGGACGTCGTATGGGTAcctggtggcattggc	57

ISSN: 1472-6750