Enhanced efficiency of cloning FACS-sorted mammalian cells.

ethidium bromide, and nucleic acids are visualized on a UV transilluminator. Typically, about 50–100 ng of supercoiled pUC119-derived plasmid DNA, without any limitation concerning the length of the insert, can be detected using this method (Figure 1). This is sufficient to discriminate between transformants containing inserts of various lengths and the control DNA. Although some chromosomal DNA and RNA are also extracted, this does not interfere with the correct interpretation of the results. When properly organized, one person can easily process up to 96 samples in less than 1 h. After examination of the gel, only the clone(s) of interest can be submitted to an isolation procedure that generates high-quality DNA suitable for subsequent experiments. In summary, we have developed an extremely fast, convenient and economical method to screen recombinant plasmids. This procedure avoids virtually all incubation, precipitation/elution, washing and drying steps as used in standard plasmid preparations (1), commercial kits and colony screening by PCR (2). It can be applied in a variety of projects such as DNA sequencing, recombinant protein expression, nested deletion mapping or whenever a distinction in size between different plasmids can be made. REFERENCES