Step 4: Sub-Cloning the Sheared Fragment

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The solution from the cuvette is diluted and then plated on a large 9x9-inch agar plate that contains ampicillin, an antibiotic. This is incubated at 37°C for 18-20 hours. Only those cells that have taken in the plasmid with the ampicillin resistance gene are able to survive and reproduce. Wherever a transformed cell lands on the agar plate, it survives and begins dividing once every 20 minutes. Each colony contains about a million bacteria.

An agar plate with many transformed colonies.

Some of the colonies are blue and some are white, depending on whether they were successfully transformed (white) or not (blue).

So, if each white colony were like a single encyclopedia, this plate of hundreds of colonies would be like an encyclopedia set with hundreds of volumes, each volume containing overlapping information. All the plates containing fragments of the original 150-200-kb piece of DNA now are referred to as a "library."

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