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miketibs

Member Since 05 Mar 2012
Offline Last Active Mar 13 2013 04:14 PM
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Topics I've Started

Restriction digest band shift?

13 March 2013 - 04:15 PM

Hi Everyone,

Having trouble interpreting these results. I amplified a gene through PCR (right lane). The top band was excised and the fragment was purified. I then performed a restriction digest with HINDIII and Xho1 for 1 hour and ran the reaction on the gel (left lane). I'm having problems interpreting why there is a band shift. Is it due to maintained association of the enzymes with the DNA fragment? Would the higher band be undigested DNA and the lower one successfully digested?


Thanks!

Problems with PMA treatment

04 June 2012 - 04:39 PM

Hi Everyone,

I was hoping I could get some tips on treating cells with PMA. I got a really robust effect in our neuron cell culture with treatment with PMA a couple of times. However, I'm not getting this effect anymore. I tried new PMA, higher and lower concentrations, longer time points and different culture batches. I was wondering if PMA is sensitive to things like temperature and medium composition. Is PMA poorly soluble at low temperatures and does it need to be warmed up and mixed very well before diluted in solution? Any tips on using PMA will be helpful.

Thanks,

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