I used Trans1-T1 phage resistant chemically competent cells to extracted lentiviral gRNA expression vectors. And there were bands near 2000bp and smear on 1% argrose gel. Did the smear come from too much DNA I loaded(600-700ng)? And what are the bands near 2000bp?
This is a well-known problem, sometimes due to prolonged alkaline lysis step, the plasmid DNA become permanently denatured and an additional band of denatured supercoiled DNA migrates below the supercoiled form.
Also, you may check this link:
https://bitesizebio.com/13524/how-to-identify-supercoils-nicks-and-circles-in-plasmid-preps/
Hello Yuxin Liu ,
To answer the first question, the smearing is indeed due to increased DNA concentration which is quite normal in such cases. The band at approx. 2000 bp is very faint which could be a degraded fragment of the same plasmid. Since it is of very low intensity, it should not cause any problems with your planned experiments. If in case it does, then try utilizing a purification kit.
- Badges
- Science topic
- Similar topics
- Gels