The most commonly used method for site-directed mutagenesis is the QuikChange® method (Stratagene, La Jolla, CA, USA), which employs mutagenic plasmid amplification to introduce single site-directed mutations.
Many methods for site-directed mutagenesis have been developed, including QuickChange site-directed mutagenesis [ 15], the overlap extension method [ 16, 17], and the megaprimer method [ 18, 19].
This chapter presents methods for site-directed mapping of histone DNA interactions with single base pair resolution.
Please refer to online supplementary methods for site-directed mutagenesis, luciferase assay, NF-κB p65 translocation, measurement of interleukin-8 (IL-8) production, immunoblot analysis, mRNA and miR expression analysis and microRNA profiling in blood samples.
EGFP-ΔPEST vector was generated by site directed mutagenesis according to the QuickChange method (Stratagene) using the primers ΔPEST-For and ΔPEST-Rev and EGFP-HD-PTP as template.
For the site directed mutagenesis we used the Quickchange Site Directed Mutagenesis Kit (NO. 200518, Stratagene).
Molecular docking studies coupled with site directed mutagenesis were used to predict the binding sites for the compound.
In combination with site directed spin labelling, pulsed electron electron double resonance (PELDOR or DEER) could be a well-suited method for their characterization and localization.
Myc3-RHO1Q68H was created by site directed mutagenesis.
IDH1 mutants were generated using the site directed mutagenesis method.
Mutant constructs were generated using a PCR-based site directed mutagenesis method, as described previously (Nøhr and Kristiansen, 2003).
