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#1 radicalteh

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Posted 14 May 2010 - 05:29 AM

I digested a Enhanced Cyan Fluorescent Protein (ECFP) fragment from it's plasmid. When I purify it, and try to read the amount by spectrophometry, it came up negative. I then did ethanol precipitation, where I can clearly see the DNA, but when i did spec again, came up negative. I double check my spec machine with other plasmid, its working. I just dont know why...anyone any adivce? HELP?

#2 HomeBrew

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Posted 14 May 2010 - 06:45 AM

Hi radicalteh -- welcome to the BioForums!

When I purify it, and try to read the amount by spectrophometry, it came up negative.


How did you purify the fragment? Was it visible on an ethidium bromide stained gel? What do you mean by "negative"? What reading did you get on the spectrophotometer?

I then did ethanol precipitation, where I can clearly see the DNA...


What do you mean by "I can clearly see the DNA"? Do you mean you can see a pellet (the presence of a pellet does not necessarily mean there's DNA there...)? How did you do the ethanol precipitation?




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