Oligonucleotide-directed mutagenesis of the FECH-encoding plasmid
| Mutant . | Oligonucleotide sequence . |
|---|---|
| C406Y | 5′-GC TGT CCG CTC TAT GTC AAT CCT GTC TGC AGG GAG-3′ |
| C406S | 5′-GC TGT CCG CTC TCT GTC AAT CCT GTC TGC AGG GAG-3′ |
| N408K | 5′-CTG TCC GCT CTG TGT CAAACC TGT CTG CAG-3′ |
| P409S | 5′-GCT GTC CGC TCT GTG TCA ATT CTG TCT GCA GGG-3′ |
| C411G | 5′-CTG TCG GCA GGG AGA CTA AAT CCT TCT TCA-3′ |
| N408K-P409S- C411G | 5′-CTG TGT CAAATC TGT CGG CAG GGA GAC TAA ATC CT-3′ |
| Mutant . | Oligonucleotide sequence . |
|---|---|
| C406Y | 5′-GC TGT CCG CTC TAT GTC AAT CCT GTC TGC AGG GAG-3′ |
| C406S | 5′-GC TGT CCG CTC TCT GTC AAT CCT GTC TGC AGG GAG-3′ |
| N408K | 5′-CTG TCC GCT CTG TGT CAAACC TGT CTG CAG-3′ |
| P409S | 5′-GCT GTC CGC TCT GTG TCA ATT CTG TCT GCA GGG-3′ |
| C411G | 5′-CTG TCG GCA GGG AGA CTA AAT CCT TCT TCA-3′ |
| N408K-P409S- C411G | 5′-CTG TGT CAAATC TGT CGG CAG GGA GAC TAA ATC CT-3′ |
Variants of FECH-encoding plasmid, pGEX-FECH,23 were constructed as described under “Materials and methods” using the oligonucleotides shown above. The nucleotide substitutions introduced by mutagenesis are underlined and in bold.