To produce mutated 3′UTR, the mutations were generated using QuikChange II XL Site-Directed Mutagenesis Kit (Stratagene).
The mutations were generated with the "Mutate Residue" Plugin of the VMD software [42].
Mutations were generated such that each VDRE (containing two half-sites, each) was mutated in all seven possible combinations of single mutations, double mutations, and one triple mutation.
All constructs and point mutations were generated using a standard PCR-based cloning strategy and verified through DNA sequencing.
He successfully corrected the gene responsible for the blood disorder β-thalassaemia (a potentially fatal blood disorder), though some unexpected mutations were generated.
There have been several studies in which intestinal organoids carrying specific disease-relevant mutations were generated for pathophysiological study and for screening of new pharmaceutical interventions.
In the present study, two sets of homodimeric site-directed IgG1 mutations were generated to understand how individual Fc Met oxidation affects FcRn binding.
Affinity mutations were generated in CDR2 and FR3 residues, yielding improvements in affinity of greater than 10,000-fold (a KD value of 180 pmol).
To confirm that these amino-acid residues are important for ubiquinol-binding and hence activity substitution mutations were generated and characterised.
Point mutations were generated by site-directed mutagenesis.
All constructs were entirely sequenced to confirm that no unintended mutations were generated during PCR reaction.
