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Quantitative RT-PCR RNA from the cells was extracted using the FastPure Cell/Tissue Total RNA Isolation Kit-BOX 2 (Vazyme, RC101-01)

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Using codon-randomizing mutagenesis to generate a clone library bearing nearly 80% of all possible amino acid changes, we measured the functional consequences of CBS variation by measuring the effects of selection on allele frequencies during a competitive yeast complementation assay using next-generation sequencing