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Sequencing PCR problem

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Dear friends,

I am a new member of this forum. Actually i was thinking about problem with my sequencing PCR. I did successfully this simple experiment many times with various plasmids extracted from E. coli. Recently I need to sequence the new cloned gene but I have NNNNNNN in my sequencing sheet. I changed everything that you may think! but the results are the same.

It would be really appreciated if somebody shears his/ her experiences with me.

Thank you in advance,

M. Dadashi

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