K8 or K18 wild-type gene was subcloned into downstream of the hCMV promoter in pMRB101 mammalian expression vector, and then the plasmids were used for generation of K8 or K18 mutant. Plasmids containing K8 or K18 mutations were generated by the PCR-based mutagenesis method. PCR was performed with designed primers using Pfu high-fidelity polymerase (Agilent Technologies, Santa Clara, CA, USA).
Pfu high fidelity polymerase
Manufactured by Agilent Technologies
Sourced in United States
Pfu high-fidelity polymerase is a thermostable DNA polymerase enzyme that can amplify DNA with high accuracy. It exhibits proofreading activity, which helps maintain the integrity of the amplified DNA sequence.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using pfu high fidelity polymerase
1
Engineered K8 and K18 Mutants
2
PCR Amplification of Sordellilysin
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Oligonucleotides (Integrated DNA Technologies Inc.) (Forward primer = 5′-GTACATATCCAGGAGCATTACAAC-3′; Reverse primer = 5′-CCACCATTCCCAAGCAAGACCTGT-3′) were designed to amplify sordellilysin based on the reported sequence of perfringolysin O [5 (link)]. PCR was performed using Pfu high fidelity polymerase (Agilent Genomics) and appropriate cycling conditions. Products were separated via agarose gel electrophoresis and compared with corresponding products from C. sordellii ATCC9714 (Figure 2 ).
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