I have been using the NEB Hifi Assembly master mix and following the same protocol. I used three different constructs but the same vector. Whenever I do the colony PCR for screening, I always get two bands instead of just one. The size of the bands is exactly what it should be, but it looks like a mixed clone. I am wondering how every cell can take up two plasmids (one wildtype and one having an extra copy of EYFP) during the transformation. Can anybody please suggest what should I do to resolve that issue or any other technique that should I try. Thank You!

Bilal

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