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size of gene insert wrong?


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#1 eunicycle

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Posted 02 March 2009 - 04:14 PM

I performed a double digestion using BamH1 and HindIII on a strain of E. Coli. accorrding to the protein sequnce, the size of the insert should be 82 bp. But after the double digestion and looking at it on my agarose gel, I calculated that the insert was 0.745 kb. This is a really big difference....what went wrong here?

#2 dormancy

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Posted 02 March 2009 - 04:25 PM

I performed a double digestion using BamH1 and HindIII on a strain of E. Coli. accorrding to the protein sequnce, the size of the insert should be 82 bp. But after the double digestion and looking at it on my agarose gel, I calculated that the insert was 0.745 kb. This is a really big difference....what went wrong here?

Hi
your RE have other restriction sites, not unique to your sequence.

#3 pcrman

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Posted 02 March 2009 - 09:42 PM

82 bp is kind of too small to see on agaros gel after double digestion.

#4 hanming86

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Posted 04 March 2009 - 05:56 AM

82 bp is kind of too small to see on agaros gel after double digestion.



Yea i have to agree with pcrman. and perhaps u should double check on the sequence of the DNA
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