Montys

Hi everybody,

I should start with a new project in a new field. I should analyze an Protein. Therefore we want to look of different truncated versions of this protein (if it interacts with other, can fold properly aso.)

Now I have to make Constructs/Plasmids for each truncated version we want to test.

Questions are how to achieve this in the best way. I know that I can use specific primer, that map the region of the wanted truncated protein or full length protein, but how important is the 3'UTR region. The polyA I can get from plasmids, but should I if I take out a specific domain in the end of the gene/protein as in (Construct B) add the 3'UTR.

Example:
Construct A: 1---------2------------3----------4-------------5--------UTR----polyA
Construct B: 1---------2------------3----------4-------------
Construct C: 1-------------------5--------UTR----polyA

Do you have a good idea how to obtain the Construct C easily. I have to cut out right in the middle of the gene several domains. Of course you can try using enzymes but I am not sure if I can that way cut out ONLY the specific domains.