Adolescent idiopathic scoliosis (AIS) is a three-dimensional structural deformity of the spine that affects 2–3% of adolescents under the age of 16. AIS etiopathogenesis is not completely understood; however, the disease phenotype is correlated to multiple genetic loci and results from genetic–environmental interactions. One of the primary, still unresolved issues is the implementation of reliable diagnostic and prognostic markers. For clinical management improvement, predictors of curve progression are particularly needed. Recently, an epigenetic contribution to AIS development and progression was proposed; nevertheless, validation of data obtained in peripheral tissues and identification of the specific mechanisms and genes under epigenetic control remain limited. In this study, we propose a methodological approach for the identification of epigenetic markers of AIS progression through an original workflow based on the preliminary characterization of local expression of candidate genes in tissues directly involved in the pathology. The feasibility of the proposed methodological protocol has been originally tested here in terms of identification of the putative epigenetic markers of AIS progression, collection of the different tissues, retrieval of an appropriate amount and quality of RNA and DNA, and identification of suitable reference genes.