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molecular revolution The first molecular markers: allozymes Allozymes Enzymes that diifer in amino acid sequence yet catalyze the same reaction -visible as a band on a gel -may exist at several gene loci Isozyme: allelic form of allozyme (same locus) + Allozyme assay • Gel electrophoresis • Current • Charge • Gel matrix • Starch or other medium • Protein variation • Charge • depends on pH Substrate • Size • different length of AA chain • Shape • Tertiary and quaternary structure • Visualization • Uses enzyme reactions • Specific substrate for each allozyme enzyme Visible Product Lab assay of isozymes Isozyme gels Quaternary Structure and Allozymes • Enzymes can consist of several subunits • Monomer • One subunit – Dimer • Two subunits – Tetramer • Four subunits AA Genotype AB BB Disparate viewpoints about true levels of genetic variation • classical + + + m + ... + + + ============== + + + + + ... + + + • balanced A1 B2 C3... X1 Y1 Z3 ================= A4 B2 C7... X1 Y1 Z2 Allozymes revealed unexpectedly high levels of genetic variation Allozyme questions • What type of DNA variation can you detect with isozymes? • What is the ascertainment bias? • Why were they wildly popular in the 1970’s and 1980’s, then disappeared by 2000?