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The proportion of UL97 mutant and wildtype strains influenced GCV IC50 values.
Total histones were extracted from mutant and wildtype larvae at 9 dpf and H3K27me3 was analyzed by western blot using specific anti-H3K27me3 anti-H3K27me27me2 antibodies.
The small interfering RNAs targeting the KRT9 R163W mutant and wildtype KRT9 were transfected into HaCaT cells, and total RNA isolated at 72 h post-transfection.
The influence of the proportion of UL97 mutant and wildtype strains on GCV IC50 values along with the strong correlation between phenotype and genotype suggests that the cell associated plaque reduction assay is highly reliable.
Interestingly, however, our examination using previously reported data from two IDH1-mutant cell lines that possess both the IDH mutant and wildtype alleles22 revealed that these cell lines were also grouped together with our G-CIMP-demethylated (C.3) tumors by clustering analysis (Supplementary Fig. S3).
How this distant mutation influences catalysis is dependent on the comparison of the crystal structures of the mutant and wildtype Rubisco, which is now under investigation of our group.
Comparing mutant and wildtype tagged proteins in transfected cells is a well-established experimental paradigm.
The latter were then intercrossed to obtain homozygous mutant and wildtype mice for experiments.
The mutant and wildtype AIM2 proteins are of similar predicted molecular weights (39 kDa and 40 kDa respectively).
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Similarly, confocal microscopy of cells expressing the GFP-RBD reporter revealed no difference between mutant and wildtypes in overall Ras activity estimated by determining the proportion the cell periphery labelled with the RBD.
These data strongly suggest that oligomerization among DN mutants and wildtype monomers, represents the mechanism responsible of the DN phenotype.
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