Table 1 |
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|
Primers used for mutagenesis |
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|
Primers |
Sequences a |
Tm pp (°C)b |
Tm no (°C)c |
Usage d |
|
|
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|
3026L/MF |
5'CTTCTAGAATTATGAAGATAAAAGGAATAAAAAGAATAGTAGTAC3' |
41.1 |
47.5 |
26L/M |
|
3026L/MR |
5'TATCTTCATAATTCTAGAAGCTATTTTGTTTTGTTCTTTAGCTTG3' |
41.1 |
50.5 |
26L/M |
|
3051L/MF |
5'ATAAGGTATTCGATGACAATACACAGTCAAAACAACTTCAG3' |
45.6 |
51.7 |
51L/M |
|
3051L/MR |
5'TTGTCATCGAATACCTTATCTTTCCATCCTGTGGTACTG3' |
45.6 |
52.5 |
51L/M |
|
3327L/MF |
5'AATATTATTTTTATGGTTGAACCTCAAATCTTGTTTTATGCAAAA3' |
43.4 |
49.3 |
27L/M |
|
3327L/MR |
5'TTCAACCATAAAAATAATATTCTTGGTTATTATATCTTTTATATCTTTTTT3' |
43.4 |
48.5 |
27L/M |
|
3356L/MF |
5'GATAGTAGGATATATGGAAATTGCAGAAAAACATTATCGTTTG3' |
43.5 |
51.5 |
56L/M |
|
3356L/MR |
5'ATTTCCATATATCCTACTATCCTATTTCTCTTTTTTGCATAAAACAAG3' |
43.5 |
51.2 |
56L/M |
|
FabH2111C/AF |
5'GCCGCCGCCACCGGCTTCGTCTACGGCCTGGCCAG3' |
50.3 |
60.2 |
111C/A |
|
FabH2111C/AR |
5'GCCGGTGGCGGCGGCGGACAGGTCGAACGCGAGC3' |
50.3 |
60.0 |
111C/A |
|
FabH2111C/SF |
5'GCCGCCTCCACCGGCTTCGTCTACGGCCTGGCCAG3' |
50.0 |
60.2 |
111C/S |
|
FabH2111C/SR |
5'GCCGGTGCAGGCGGCGGACAGGTCGAACGCGAGC3' |
50.0 |
60.0 |
111C/S |
|
VRARN3F |
5'TTTTCAGGGCAAAGTATTGTTTGTGGATGATCATGAAATGG3' |
45.0 |
54.5 |
N3/D |
|
VRARN3R |
5'ATACTTTGCCCTGAAAATACAGGTTTTCGGTCGTTGGGATG3' |
45.0 |
59.6 |
N3/D |
|
VRARC5F |
5'TATGCATTCCAATAGGGATCCGAATTCGAGCTCCGTCGAC3' |
49.0 |
58.1 |
C5/D |
|
VRARC5R |
5'GATCCCTATTGGAATGCATAGATAACAGCTTGTGTTCTATCTTGCAC3' |
49.0 |
56.7 |
C5/D |
|
VRARIHISF |
5'CATAATTTAATTCAAAAGCACCATCACCATCATCACTGAGAATTCGAGCTCCGTCG3' |
44.6 |
56.8 |
CT/I |
|
VRARIHISR |
5'GTGCTTTTGAATTAAATTATGTTGGAATGCATAGATAACAGCTTGTG3' |
44.6 |
55.9 |
CT/I |
|
VRARDHISF |
5'GTTTAAGAAGGAGATATACATATGACGATTAAAGTATTGTTTGTG3' |
43.8 |
50.7 |
NT/D |
|
VRARDHISR |
5'TGTATATCTCCTTCTTAAACTTAAACAAAATTATTTCTAGAGGGGT3' |
43.8 |
51.3 |
NT/D |
|
|
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|
a: Primer-primer overlapping sequences are in italic. The sequence insertion is in bold face. b: Tm pp (°C) was calculated from the primer-primer overlap sequence. c: Tm no (°C) was calculated from the primer sequence matched to the template. d: N3/D: three amino acids deleted from the N-terminus; C5/D: five amino acids deleted from the C-terminus: NT/D: N-terminal His tag and TEV protease recognition site deleted and CT/I: sequence encoding 6 × His tag inserted onto the C-terminus. |
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|
Liu and Naismith BMC Biotechnology 2008 8:91 doi:10.1186/1472-6750-8-91 |
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