Advances in RNA analysis of dystrophin (DMD) mutations have been significantly influenced by improvements in methods to study mRNA transcription and splicing. Early breakthroughs, such as the 1988 cDNA sequencing of the 14-kb muscle transcript isoform of the dystrophin gene, paved the way for RNA-based mutation detection via RT-PCR. These studies revealed that point mutations often cause premature polypeptide chain termination, and RNA polymerase requires approximately 16 h to transcribe the 2.1 Mb Dp427m transcription unit. Next-generation technologies for RNA sequencing (RNA-seq) have enabled broader and more quantitative transcriptome analysis of the DMD locus. This protocol outlines an RNA-seq approach using rRNA depletion to examine muscle biopsy total RNA from DMD/BMD patients, specifically focusing on resolving pseudoexon and other intronic mutations. While not achieving the ultra-deep read depths of targeted RNA-seq methods, this cost-effective strategy provides sufficient coverage to robustly evaluate DMD exon and intron mRNA levels, generating data with comparable read depths for both.