1 year ago
Hi everyone! I am planning on floxing mice with CRISPR but I am having trouble deciding which region to target. Do you have any tips on choosing?
Experiment: CRISPR Mouse - Activation 3T3-L1 C/EBPβ
Hi Mario, I will try to walk you step by step on this. First you need to decide which part of the gene you will delete using loxP or Cre mediated recombination. Then you need to look up the mouse sequence. For this you can use something like Ensembl genome browser. You need to determine where the splice donor and acceptor sites are located. Afterwards, you should align your sequence with that of a different species and check for differences in the untranslated region. If you can identify stretches that are present in the other species but absent in mouse you should use those. You should select a species like rat which is similar to mouse because the stretches you selected are most likely not involved in gene regulation or splicing. If rat and mouse are too much alike then you can compare it to the human sequence and find some differences there. Hope this helps.