Recombinant DNA techniques: storage and screening of cDNA libraries with large numbers of individual colonies from initial transformations.
A typical cDNA library with a large number of initial transformants, plated in soft agarose, can be stored and shipped in 12% glycerol at -70 degrees C. To prepare the library for storage, the soft agarose layer is made into a paste and the agarose is removed by Sephadex G-25 filtration. This method of cDNA library storage does not alter the relative representation of the plasmids carried in the library. To achieve a very uniform distribution of colonies at high colony density, an aliquot of the cDNA library is diluted to 3000 to 10,000 colonies/ml. One milliliter of this suspension is evenly distributed on a nitrocellulose filter on an agar plate and air-dried. Filter copies are made and screened by published methods.
D003001:Cloning, Molecular; D004247:DNA / Q000302:isolation & purification*; D004274:DNA, Recombinant / Q000302:isolation & purification*; D004926:Escherichia coli / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D014169:Transformation, Bacterial
D004274:DNA, Recombinant; D004247:DNA