Re-verification of identified pathways. Since not all the identified pathways from Step (1) are directly related to the overproduction, a re-verification is necessary to filter the low-relevant targets. For these identified pathways, their fluxes under overproducing networks were considered as the optimum fluxes (fluxopt) to achieve theoretical maximum yield of the product. The simulated product yields (Yield) constrained by each fluxopt for the wild-type network were used to evaluate the best contribution of each up-regulated pathway toward overproduction. Pathways with Yield > 0 are considered as the key targets that directly lead to the overproduction.