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Problems in screening genomic DNA Library with antibody

Problems in screening genomic DNA Library with antibody - Molecular Cloning Forum

Problems in screening genomic DNA Library with antibody - DNA cloning forum. Discuss the cloning of plasmids, vectors, DNA, RNA, cDNA, proteins, and other molecules for expression, sub-cloning, and other methods. Also discuss digestion, ligation, transformation, plasmid preparation, cDNA library screening, and competent cells here.


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Old 01-17-2010, 04:42 PM
Pipette Filler
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Default Problems in screening genomic DNA Library with antibody



I constructed a genomic DNA library of a bacterium using Partial DNA digestion employing Lambda ZAP II Kit (Stratagene). I did everything alright as shown in kit. I used 2-4 kb fragments for ligation with pre-digested vector supplied in kit. I used 1 uL of ligate for packaging and then amplified the library. In Blue-white screening, all the plaques showed transperant or clear with blue rings at the edges. There was no blue colony. I thought all are recombinant clones. Then, I started screening the library using antibodies. But I am not getting any positive clones. Please any one help me and tell the reason? My antibodies are good and I checked them several times.
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