ISHS Acta Horticulturae 217: IV International Workshop on Fire Blight
THE RELEASE AND ELECTRON MICROSCOPE VISUALISATION OF PLASMIDS IN ERWINIA AMYLOVORA USING AN IN SITU LYSIS TECHNIQUE |
| Authors: | D.C. Sigee, M.H. El-Masry |
| Abstract:
A novel microscale technique is described for the electron microscope investigation of bacterial plasmids, involving bursting of lysozyme-treated cells on an electron microscope grid (in situ) with detergent. The resulting preparation contains supercoiled and relaxed (open circle) plasmids. The latter have clear protein and membrane associations. Measurement of contour lengths results in a continuous size-histogram rather than one with discrete size categories. This may arise due to variations in localised coiling or to variations in the degree of extension under the prevailing surface tension forces. Gel electrophoresis resolved a single band of plasmids at about 20Kbp (probably corresponding to a major group of molecules measuring 3–5 μm in the electron microscope preparations), but did not reveal other plasmid sizes - though molecules up to 120 Kbp were observed in the in situ material. | |
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