Skeletal muscle is a complex, versatile tissue with a large variety of diverse fibre types. The heterogeneity is determined by a combination of the individual properties of the different fibre types and their proportion, making the analysis of the expression profile difficult. Understanding molecular processes regulating fiber type diversity is still complicated by the presence of additional components of other tissues. The study aimed to identify the transcriptional profiles of single fibres isolated from mouse muscles. We studied 4 distinct muscle fibre types: slow fibres (MyHC-1) from soleus and fast-glycolytic fibres (MyHC-2B) from EDL and intermediate fiber type expressing MyHC-2A or MyHC-2X isolated from same muscles. The protocol used allowed genomic analysis at single-cell level in muscles (microgenomics). The analysis was "supervised" as each isolated myofibre was classified according to the MyHC expression, prior the microarray analyses which were carried out on platforms with 13000 probes. Both qualitative and quantitative improvements were achieved. Myofiber profiles were virtually free from non-muscle transcriptional activity and a list of genes co-expressed with the MyHC isoforms, and therefore novel markers that could be useful for fibre type functional classification, was obtained. This innovative approach will allow a better understanding of the adaptive transcriptomic transitions and muscle plasticity under physiological and pathological conditions.