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However, there are a few limitations to the rational design approach: (i) requires prior knowledge of the target structure or interaction surface to identify potential ligands, (ii) identified ligands may not be amenable to structural stabilization after isolation, as proper folding may be dictated by interactions within the entire protein structure or interface, 100 and (iii) identified ligands may not represent the optimal binders as not all combinatorial space is explored compared to screening approaches

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The successful PCR product (Delhi and Che-S only) were sequenced through the Sanger method using the primer-walking strategy