1. How do you purify wt virus - that could give the lead. 2. How do you expext the mutant virus to be different than the wt virus. 3, Can those differtence be exploited. 4. Randomly puirify some clones/preps, characterize them by whatever assays are applicable.5. If plaque purfication possible, do so and characterize, even sequence.

I'm currently trying to purify by plaque picking up.

The mutation would contribute to the cell tropism of virus but since mutant virus is still mixed, there is no was I can demonstrate. 

I could verify mutation by PCR and already done with sequencing.

Any other tips from experiences?

Hi Sungseok, 

Despite reading your question multiple times, I am afraid to say that I could not understand it. If you can elaborate it better, I am sure many people on this forum will be able to help. 

As far as I understand, you added a 20 bp DNA stretch to a non-structural protein region of your virus, and confirmed it with sequencing. As a next step, you transfected cells with this DNA. Here I am confused: did you transfect your cells with a mixture of wild-type and mutated DNA? If so, the wild-type and mutated DNA would make virus particles, which are then released in the culture medium. Are you interested in separating the wild-type and mutants from this culture medium?

Please confirm, and we can then take it from here.