I know that there are Nuclear Localisation Sequencenes (NLS). They can be taken from endogenous or viral proteins and fused to the N or C terminus of my recombinant protein.

Which is the best one? Which the best short one (so that it can be more easily added with a Primer)? Should it be attached to the N or the C terminus? Do they influence expression?

I have heard that size does matter (Too large wont fit through the nuclear pore, too smalll will diffuse out again). What is the range of protein sizes that can be targeted efficiently?

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    $\begingroup$ To use a N-terminal or a C-terminal tag depends on your protein. I can only say that you can try both and see which one does better. $\endgroup$
    Dec 17 '14 at 11:48
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    $\begingroup$ It also depends if your protein has important domains on one of the sides. If so, I would start by using the other end to avoid interference. $\endgroup$
    – Chris
    Dec 17 '14 at 12:05
  • $\begingroup$ Thanks for the general tip. In my case I have seen people putting stuff (including NLS) to both ends of the protein. Here I am really more concerned about transport efficiency. $\endgroup$
    – jan-glx
    Dec 17 '14 at 13:22
  • $\begingroup$ I've seen people use 3 NLS sequences in a row, I assume this improves transport efficiency, but they never explained their reasoning. $\endgroup$
    – user137
    Dec 17 '14 at 18:15

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