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Preparation of megabase-sized tomato DNA and separation of large restriction fragments by field inversion gel electrophoresis (FIGE) by Raymond A. J. van Daelen; Jos J. Jonkers; Pim Zabel is a Agricultural and Biological Sciences article available to read on EtoBox.

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The Schwartz and Cantor technique for releasing and fractionating megabase-sized DNA from agaroseembedded cells is beginning to bridge the gap in resoluation between classical genetics and current molecular DNA techniques, particularly in mammalian systems. As yet no conditions have been described for preparing plant DNA that is of sufficient length to allow similar long-range restriction mapping experiments in plant systems. In this report, we describe the application of the Schwartz and Cantor technique for preparing high molecular weight DNA from embedded tomato leaf protoplasts, as well as conditions for generating and fractionating large restriction fragments, by field inversion gel electrophoresis (FIGE). The bulk of DNA released from lysed protoplasts was at least 2 Mb in size and amenable to restriction digestion as shown by hybridizing Southern blots with, among others, a probe for the Adh-2 gene of tomato. Restriction fragments as large as 700 kb were detected. Chloroplast DNA is isolated intact, amenable to restriction analysis and, in its native form, not mobile in FIGE.

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Author

Raymond A. J. van Daelen; Jos J. Jonkers; Pim Zabel

Publisher

Springer; Springer-Verlag; Kluwer Academic Publishers; Springer Science and Business Media LLC; Society for Mining, Metallurgy and Exploration Inc. (ISSN 0167-4412)

Published

1989

Language

EN

Field

Agricultural and Biological Sciences (Life Sciences)