The myogenic regulatory factors (MRFs) are a group of muscle specific transcription factors important during determination and differentiation of muscle fibers. In adult muscle two of the members, MyoD and myogenin, are expressed preferentially in fast and slow muscles, respectively, and have been suggested to play a role in fiber type specific gene regulation. Overexpression of MyoD in the slow soleus muscle has been shown to cause a shift towards a faster phenotype, while overexpression of myogenin in the fast extensor digitorum longus (EDL) causes an increase in oxidative capacity. Less is known about the two last members of this family, MRF4 and Myf-5. In adult muscle MRF4 is expressed at very high levels and Myf-5 at very low levels, but their function is unknown. The expression of MRF4 has only been investigated at the transcript level, so we wanted to map the normal expression of MRF4 protein. In order to investigate whether MRF4 and Myf-5 play a role in regulation of fiber type specific genes, we wanted to knock down the expression of MRF4 and to overexpress Myf-5 in muscles of adult rats. Mapping of MRF4 protein expression showed that there was a significantly higher expression level in the slow soleus compared to the faster muscles. For the knockdown study, six siRNA was tested in cell culture, but since none of them gave a measurable knockdown, further studies in vivo were abolished. Overexpression of Myf-5 in the fast EDL resulted in an increase of 2b fibers, indicating that Myf-5 activates expression of MyHC 2b.