Abstract
β-galactosidase (β-gal) is among the most frequently used markers for studying a wide variety of biological mechanisms, e.g., gene expression, cell migration, stem cell conversion to different cell types, and gene silencing. Many of these studies require the histochemical detection of relative β-gal levels in tissue cross-sections mounted onto glass slides and visualized by microscopy. This is particularly useful for the analysis of promoter activity in skeletal muscle tissue since the β-gal levels can vary dramatically between different anatomical muscles and myofiber types. The differences in promoter activity can be due to a myofiber’s developmental history, innervation, response to normal or experimental physiological signals, and its disease state. It is thus important to identify the individual fiber types within muscle cross-sections and to correlate these with transgene expression signals. Here, we provide a detailed description of how to process and analyze muscle tissues to determine the fiber-type composition and β-gal transgene expression within cryosections.
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Acknowledgments
Miki Haraguchi and Paul Gregorevic are thanked for their initial technical assistance and very helpful advice; Joel R. Chamberlain is thanked for providing a ROSA26 mouse; and Robert E. Welikson, Charis L. Himeda, and Joel R. Chamberlain are thanked for their critical comments on earlier versions of the manuscript. This research was supported by grants from the NIH RO1-AR18860 and 1P01-NS046788 to SDH and by an NIH Developmental Biology Training Grant 5732-HD07183 to PWLT.
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Tai, P.W.L., Smith, C.L., Angello, J.C., Hauschka, S.D. (2012). Analysis of Fiber-Type Differences in Reporter Gene Expression of β-Gal Transgenic Muscle. In: DiMario, J. (eds) Myogenesis. Methods in Molecular Biology, vol 798. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-343-1_26
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DOI: https://doi.org/10.1007/978-1-61779-343-1_26
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