Aims: The present study aimed to investigate the genetic structure of A. aculeata (macaw palm) groups by analyzing individual polymorphism of different ITS regions. Study Design: Experimental design study. Place and Duration of Study: Department of Forest Engineering (Universidade Federal dos Vales do Jequitinhonha e Mucuri/MG) and Germoplasm Bank of macaw (Universidade Federal de Viçosa/MG), between February 2014 and December 2015. Methodology: DNA extracted from 42 leaf samples of Macaw palm. Five universal oligonucleotide primer pairs specific for different internal transcribed spacer (ITS) regions of nuclear ribosomal DNA of Macaw (ITS 1, ITS 2, ITS 3, ITS 4, and ITS 5) used for PCR amplification. Results: Of 42 analyzed samples, 20 resulted in reproducible bands for primers ITS 1 and ITS 2, 21 for ITS 3 and ITS 4, 24 for ITS 2 and ITS 5, and 22 for ITS 4 and ITS 5. Samples were grouped based on these results. Conclusion: Therefore, it was possible to identify genetic differences among Macaw progenies based on the quality of ITS sequences.