In genetic dissection, the supplement that restores growth indicates
Supplementation test interprets growth restoration as evidence that supplied compound lies downstream of metabolic block in pathway. Auxotroph unable to synthesise its own product depends on exogenous molecule, and only molecules occurring after defective step can be utilised because earlier molecules still need defective enzyme for conversion. Pattern of rescue across many mutants therefore distinguishes upstream versus downstream intermediates enabling determination of gene order. Column with many positives indicates early metabolite rescuing many blocks while narrow rescue indicates late metabolite position in sequence.
Ref: Hartl & Ruvolo, Genetics, 6th ed., Chapter 14: Pathway Analysis Using Auxotrophs Rescue Principle