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Cloning Large Fragments in a pool of DNA fragments


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#1 Samantha

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Posted 01 March 2005 - 08:00 PM

hello

In my experiment, genomic DNA was digested into fragments with different sizes from ~200-1000bp and then amplified by PCR. After that, I cloned the pool of fragments into a vector by TA cloning (TOPO2.1) and then the clones were sequenced. I found that most of the fragments cloned into the vector were small in size, about 200-300bp. I want to ask how to improve the experiment, so the large fragments, >300bp can be cloned. I know that I can add more vector or use some gel filtration column to eliminate the small fragments. However, are there any other methods? or have you faced this problem?

#2 sharath

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Posted 01 March 2005 - 08:34 PM

Run the mixture on a low melting agarose gel and extract the bands of required size and use them for cloning.
Sharath B.




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