Cell Lineage Trees in Mice Figure 4. Lineage analysis of satellite cells. A muscle fiber from the Gastrocnemius muscle of ML4 was stained for DAPI (blue, all nuclei within the fiber) and Pax7 (pink, satellite cells). A. DAPI staining. B. Pax7 staining showing two satellite cells. C. Overlay of DAPI and Pax7. D. Single satellite cells were isolated from muscle fibers and cultured ex vivo, creating clones whose DNA approximates the DNA of the founder cell. Part of a Petri dish containing a single cell culture is shown. Myofiber structures are evident also in culture. E. Satellite cells can be categorized based on a hierarchy of body-side, muscle and myofiber. Myofibers are numbered from most superficial (#1) until deepest in the muscle. F. The average MRCA depth between pairs of satellite cells from the same myofiber/muscle/body-side was compared to the average MRCA depth between all pairs of cells. Pairs of satellite cells from the same myofiber had on average a significantly larger MRCA depth than a random pair of cells (p,0.0001) demonstrating that myofiber is a significant lineage unit. G. Correlation between lineage and physical proximity in the two main muscles of ML7. The physical distance between myofibers was defined as the difference between myofiber indices (e.g. the distance between a satellite cell on myofiber #3 and a satellite cell on myofiber #1 is 2). Physical distance and MRCA depth are significantly anti-correlated (p = 0.02 and p = 0.0001 for Gastrocnemius left and right, respectively; values stand for anti-correlation coefficients), demonstrating that satellite cells which are physically close to each other tend to share a common developmental path. doi:10.1371/journal.pone.0001939.g004 PLoS ONE | www.plosone.org 7 April 2008 | Volume 3 | Issue 4 | e1939