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Table 1 The primers for site-directed mutagenesis

From: A mutant screening method by critical annealing temperature-PCR for site-directed mutagenesis

Primer Sequence Length Mismatches T m
D112G(F) CTATGgCCTCCTAAAGGACCTAGAGGAAG 29 1 76.0
D112G(R) GGAGGcCATAGACGTTGCTGTCAGAG 26 1 75.3
D112K(F) CTATaAaCTCCTAAAGGACCTAGAGGAAG 29 2 68.3
D112K(R) GGAGtTtATAGACGTTGCTGTCAGAG 26 2 66.8
I4V(F) CAACCgTTCCCTTATCCAGGCTTTTTG 27 1 72.5
I4V(R) GAAcGGTTGGGAACATATGTATATCTCCTTCT 32 1 73.9
L45D(F) CATTCgatCAGAACCCCCAGACCTCCCTCTGTTTCT 36 3 76.1
L45D(R) GGTTCTGatcGAATGAATACTTCTGTTCCTTTGGGAT 37 3 71.8
E56D(F) TCAGAcTCTATTCCGACACCCTCCAAC 27 1 74.1
E56D(R) GAATAGAgTCTGAGAAACAGAGGGAGGTCTG 31 1 76.3
R64D(F) CTCCAACgacGAGGAAACACAACAG 25 3 63.8
R64D(R) CCTCgtcGTTGGAGGGTGTCGGAATAG 27 3 69.7
N109Y(F) CAGCtAtGTCTATGACCTCCTAAAGGACCTAGAGGAAG 38 2 77.9
N109Y(R) GACaTaGCTGTCAGAGGCGCCGTACACCAG 30 2 76.9
D116F(F) AAAGttCCTAGAGGAAGGCATCCAAACGCTGAT 33 2 73.6
D116F(R) CTCTAGGaaCTTTAGGAGGTCATAGACGTTGCTGTCAG 38 2 77.9
R64M(F) TCCAACAtGGAGGAAACACAACAG 24 1 68.0
R64M(R) TCCaTGTTGGAGGGTGTCGGAATAG 25 1 71.8
hGH2V3-P233S(F) TGGtCgTGGAAGgTGCTACTCCAGTGCCCACCAGCC 36 3 80.6
hGH2V3-P233S(R) GAGTAGCAcCTTCCAcGaCCAGGAGAGGCACTGGG 35 3 79.4
hGHBP-S237C(F) GATGtGCTAAGAATTCGAGCTCCGTCGACAA 31 1 76.3
hGHBP-S237C(R) TTAGCaCATCTGAGGAAGTGTTACATAGAGCACC 34 1 76.8
pGHBP-S237C(F) GATGtGCTAAGAATTCGAGCTCCGTCGACAA 31 1 76.3
pGHBP-S237C(R) TTAGCaCATCTGAGGAAGTGTTACATAGAGCACC 34 1 76.8
hGHR1-255R235C(F) AACAAtgcAACTCTGGAAATTATGGC 26 3 59.8
hGHR1-255R235C(R) TTgcaTTGTTTGGATCTCACA 21 3 58.7
hGHR1-638R235C(F) AACAAtgcAACTCTGGAAATTATGGC 26 3 59.8
hGHR1-638R235C(R) TTgcaTTGTTTGGATCTCACA 21 3 58.7
  1. Tm is the melting temperature of mutagenic oligonucleotide primers annealing with the WT template to 50%.
  2. For all primers, the mutagenesis oligonucleotides are denoted in lowercase.