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Genomic DNA Library


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#1 bigvels

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Posted 15 January 2010 - 07:39 AM

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.

#2 pDNA

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Posted 15 January 2010 - 12:47 PM

isolate plasmid DNA from your library and check the plasmids for inserts by PCR or restriction digest.

Maybe your X-Gal is not working ...you can check that easily by transforming empty vector as a control!

Regards,
p




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