Hi!
I would like to know which are the target sites for the PhiC31 Integrase. So far, I couldn't find any precise information about the Orientation of the attB-Sequence and at which position de Double Strand Break occurs.
I want to use the classical pattB-Vector with a white+ marker for transgenesis in Drosophila.
Right now, I'm trying to insert an attP-Site into the Promoter region of my target gene via CRISPR and Homology Directed Repair. Afterwards I want to insert a rescue plasmid so that my transgene would get expressed under the endogenous Promoter. For that I need to know how the attB/attP recombination event occurs exactly and where and in which direction I have to insert my transgene in my future rescue plasmid.
I would be more than glad if someone could recommend a paper or website where I could get precise answers.
For better understanding of my project I attached a PPP.
Thank you!!
Alina
Thank you very much! I couldn't find the sequences that are inserted in my plasmid.
I figured out how I have to design my Rescue Plasmid.
For future reference: What made me stumble was the fact that if you have the attP site in the genome and recomibnate with a plasmid containing attB you get the attR at the 5' end and the attL at the 3' end. If you have your attB site in the genome and recombinate with a plasmid containing the attP site you generate attL at the 5' end and attR at the 3' end. I guess this was quite confusing at the beginning for me.
Furthermore a few publications didn't consider the direction of the inserts and sometimes made mistakes.
A paper that I found after posting this question helped a lot: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0099649
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