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Quikchange site directed mutagenesis

Manufactured by Thermo Fisher Scientific
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QuikChange site-directed mutagenesis is a PCR-based method for introducing site-specific mutations into double-stranded plasmid DNA. The kit provides a rapid, efficient, and reliable method for creating point mutations, small insertions, or deletions in any plasmid.

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Lab products found in correlation

Pet sumo, by Thermo Fisher Scientific (1 mentions) Pmsp1d1 plasmid, by Addgene (1 mentions) Mycoalert mycoplasma detection kit, by Lonza (1 mentions)

Close spelling variants of this product

QuikChange Site-Directed Mutagenesis Kit QuikChange II Site-Directed Mutagenesis Kit Quikchange Lightning site-directed mutagenesis kit Site-directed mutagenesis QuikChange

2 protocols using quikchange site directed mutagenesis

1

Recombinant Anthrax Protein Production

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Recombinant wild-type (WT) PA was expressed in the periplasm of Escherichia coli BL21 (DE3) and purified by anion exchange chromatography [40 (link)] after activation of PA with trypsin [41 (link)]. QuikChange site-directed mutagenesis (Stratagene) was used to introduce mutations into the plasmid (pET SUMO (Invitrogen)) encoding a truncated recombinant portion of lethal factor. LFN E126C and was expressed as His6-SUMO-LFN, which was later cleaved by SUMO (small ubiquitin-related modifier) protease, revealing the native LFN E126C N-terminus [41 (link)]. Membrane scaffold protein 1D1 (MSP1D1) was expressed from the pMSP1D1 plasmid (AddGene) with an N-terminal His-tag and was purified by immobilized Ni-NTA affinity chromatography as previously described [42 (link)].
Machen A.J., Akkaladevi N., Trecazzi C., O’Neil P.T., Mukherjee S., Qi Y., Dillard R., Im W., Gogol E.P., White T.A, & Fisher M.T. (2017). Asymmetric Cryo-EM Structure of Anthrax Toxin Protective Antigen Pore with Lethal Factor N-Terminal Domain. Toxins, 9(10), 298.
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2

Mutant Receptor Generation and Characterization

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Mutant receptors were generated as previously described (Valant et al., 2012) using QuikChange site-directed mutagenesis (Invitrogen) and all sequences were confirmed by DNA sequencing (Australian Genome Research Facility). All receptor constructs, wild type (WT) and mutants, were stably expressed in CHO-FlpIn cells using FlpIn Gateway technology system and selected using 600 μg/mL of hygromyocin B. Cells were maintained in DMEM supplemented with 5% FBS and 600 μg/mL hygromyocin B at 37 o C in a humidified incubator (5% CO 2 , 95% O 2 ). Cells were regularly monitored for mycoplasma contamination using the Lonza MycoAlert Mycoplasma Detection Kit (Lonza, Besel, Switzerland) .
Pham V., Habben Jansen M.C.C., Thompson G., Heitman L.H., Christopoulos A., Thal D.M, & Valant C. (2023). Role of Conserved Tyrosine Lid Residues in the Activation of the M2 Muscarinic Acetylcholine Receptor. Molecular pharmacology, 104(3).
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