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DNA Smear

| posted 30 May, 2019 19:28
c.sunnen avatar
c.sunnen
Matt,
Did you use our protocol? Your gel looks similar to ours. We also had some issues with sequencing - turns out we had some bacterial DNA contamination - but we assumed that was user error (asking the TAs to add the nuclease instead of doing it ourselves). We're actually going to test this over the summer. Otherwise, our DNA seemed pretty clean.
Nikki
| posted 30 May, 2019 20:07
mastropm avatar
mastropm
Hi Nikki,
Yes we used your protocol. I think our nuclease mix is not 100% correct. We are getting a lot of what is most likely RNA contamination. The one I posted had the most smearing and the DNA concentration was too low. The one that worked had the least smearing of all.
92Kb
| posted 30 May, 2019 20:07
mastropm avatar
mastropm
Hi Nikki,
Yes we used your protocol. I think our nuclease mix is not 100% correct. We are getting a lot of what is most likely RNA contamination. The one I posted had the most smearing and the DNA concentration was too low. The one that worked had the least smearing of all.
92Kb
| posted 03 Jun, 2019 18:29
Thiel avatar
Thiel
Hello,
This was my first time running the phage discovery semester and some of the DNA was smeared. We used M. foliorum as our host. We used the nuclease mix from Sigma-aldrich (#GE80-6501-421018-03) and did the EDTA, proteinase K, and used the wizard kit. We tried it the first time and ran uncut and cut with RE. All of the gels/ phages except one, were smeared. Renzie was the best and still smeared some. We isolated DNA again and just ran uncut. From the uncut DNA gel image, Dan was able to sequence DNA from Renzie and Lenlyn.

Thanks,
Sarah
926Kb
 
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