Rapid purification of transfected porcine muscle cells
Blanton, J. R. ; Robinson, J. P. ; Gerrard, D. E. ; Bidwell, C. A. ; Grant, Alan L.
Springer
Published 2000
Springer
Published 2000
| ISSN: |
1573-0603
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|---|---|
| Keywords: |
Flow cytometry ; Muscle cell ; Porcine ; Transfection
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| Source: |
Springer Online Journal Archives 1860-2000
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| Topics: |
Biology
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| Notes: |
Abstract A detailed methodology is described for fluorescence-activated cell sorting (FACS) of porcine muscle cells that have been transfected to express green fluorescent protein (GFP). Cells are liberated from porcine skeletal muscle and primary cultures are transfected with DNA encoding GFP. Primary cultures are subjected to immunocytochemistry using a primary muscle-specific monoclonal antibody followed by incubation with a phycoerythrin-conjugated second antibody. Transfected myoblasts aresorted from fibroblasts using forward angle light scatter and ninety degree light scatter, phycoerythrin fluorescence, and GFP fluorescence. These procedures allow for isolation of genetically-engineered porcine muscle cells more rapidly than traditional clonal selection procedures. Consequently, FACS provides porcine myoblast populations that retain the majority of their replicative capacity and are not contaminated with non-myogenic cells.
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| Type of Medium: |
Electronic Resource
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| URL: |