Anything and everything cloning: Go...
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I used the protocol described in: Methods Mol Biol. 2014;1116:73-87 with minor changes.
Give me an Email and I'll send it to you so you'll be able to upload it to the website.
Steve,
The simultaneous RF cloning work fine. I'v inserted two fragments into pBABE-Puro on the first try (mCherry instead of puromycin and another gene at the MCS).
Gal
Take a look at that:
Application of the Restriction-Free (RF) Cloning
for Multicomponents Assembly
Yoav Peleg and Tamar Unger
(Methods Mol Biol. 2014;1116:73-87)
or at: http://www.ncbi.nlm.nih.gov/pubmed/24395358
Thanks.
Gal
Hi Steve,
First, thats really a great site and it helps a lot in designing RF.
I'm trying to design primers for inserting 2 genes into pBABe-puro plasmid (one into the MCS and second instead of Puro) .
The trouble is that in both genes the first 21 nucleotides are simmilar.
Do you have any suggestions to overcome it?
Maybe to design a longer primer for the gene?
Thanks
Gal
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