Megabase-size DNA isolation from plants

Megabase-size DNA isolation from plants

To construct large insert DNA libraries in BAC and YAC vectors, methods must be developed to isolate very high molecular weight DNA - megabase-size DNA - from plants. To isolate such DNA, protoplasts or nuclei must first be embedded in agarose plugs or microbeads. The agarose acts as a solid yet porous matrix which allows for the diffusion of various reagents for DNA purification and subsequent manipulations while preventing the DNA from being sheared (Schwartz and Cantor, 1984). Microbeads are preferred over plugs because the use of beads increases the surface area surrounding the tissue sample by approximately 1000 fold thereby allowing for more efficient and rapid diffusion of chemicals and enzymes into and out of the agarose beads (Cook, 1984, Overhauser and Radic, 1987, Wing..........

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http://www.genome.clemson.edu/groups/bac/protocols/megabasedna.html