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Inserting Multiple Amplicons into a plasmid

Inserting Multiple Amplicons into a plasmid - Molecular Cloning Forum

Inserting Multiple Amplicons into a plasmid - 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 11-30-2012, 08:31 PM
Pipette Filler
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Question Inserting Multiple Amplicons into a plasmid



Hi all,


So I need to insert multiple genes (a peptide coding gene and the two genes which code for its transport to the outside of the cell, which were amplified via PCR) into a plasmid vector. The genes are not located next to each other, meaning I have three different amplicons, one of each gene. The only plasmid vector we have access to is the one included in a TA TOPO kit i.e. only has one amplicon insertion site.

I wanted to know if there as I a way I could connect the three gene amplicons together to make one large amplicon, which I would then insert into the plasmid. I can think of numerous reasons why this wouldn't work orwould give me unintended results, but I'm just trying anything at this point to insert these three genes, in order, into one vector.
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amplicon , amplicons , inserting , multiple , pcr , peptide , plasmid


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