Technique / Molecular Biology / DNA analysis techniques


Separation of specific DNA fragment



Separation of specific DNA fragment

What I wish to do is to look at the repertoire of mutations in a
specific gene in a population of cells. So ideally I would like to
isolate the same DNA fragment from each cell and sequence it.


Obviously PCR on the population of cells won't work, because ... ...

Last update 14-Oct-2001, Rating of 0 votes.

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By Antonio Alcochete on 28-Sep-2002
Dear Sir,

I am runing gel agarose 3,5% electropohorese in TBE 1X and 100 V for70 min to separate genomic DNA of rice amplified with microsatellite. What happens is that some gel show smeared bands and others show mix of faint bands with smeared, and when it shows bands they are so light. PCR conditions are: 94C/4 min, 30 cicles of 94C/1 min; 56C/1min; 72C/1min; and final extension of 72C/7min. The EtBr 1mg/ul is 80 ul for 600 ml gel agarose 3,5%.

I would be very pleased if you can help to get the right results.

ThankYou

Alcochete
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By Devin Hendricks on 14-Oct-2001
Ok, I am a high school student attending Tremper High School in Kenosha, WI. Some friends of mine and I decided to run an experiment on DNA mutation of fruit flies in a toxic environment(using naphthalene or para-dichlorobenzene). I was wondering how we would run the PCR procedure on one fly within one generation of fruit flies? Would we have to run PCR on EACH CELL or would we just worry about running one PCR procedure for the whole fly? Bear with me, I'm learning this is as fast as possible, much of this is sketchy at best. Thank you very much for your time.



By the way, would cloning a fly (by nuclear cell transfer) be out of the conventional arena for a High School laboratory, or would you suggest using a corporate lab?
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