PCR Lab Design

Robert Horton horton at biosci.cbs.umn.edu
Wed Aug 30 12:02:11 EST 1995


Jeremy Carson (carsonj at ozemail.com.au) wrote:
: <description of really cool PCR contamination-resistant setup deleted>

According to my calculations: For a 400 base pair PCR product, the
molecular weight should be about 400nt * ~330 g/mol*nt = ~1.3 * 10^5 g/mol

Assuming we can detect 1ng on a gel (I think that's conservative), we'd
need (1 x 10^-9 g) * (1mol/(1.3X10^5g)) * (6.02 * 10^23 molecules/mol) =
~1.6 * 10^10 molecules

(For a 200bp band, you need twice as many molecules.)

10^10 =~ 2^33, so you would have to do at least 33 cycles of PCR to have
even a theoretical chance of detecting a single molecule. (If you can
actually see a band on an ethidium stained gel amplified from a single
molecule with fewer than 40 cycles, let me know. I'll buy you a beer.) 

This is an important point (so I hope people check my so-called math):
Contamination that you detect with fewer than 33 cycles does not come from
single molecules of DNA floating around in the air. Dust particles,
flakes, or droplets maybe, but not single molecules. So unless your air is
loaded with DNA vapors, I think you're relatively safe with HEPA filters. 


Robert M. Horton (PhD!) /\ "Crash programs fail because of the theory that
U of M Dermatology Dept || with nine women pregnant you get a baby a month"
Box 98 UMHC, 4-154 PWB /||\ -W. von Braun.   Disclaimer: "Bob who?"
Minneapolis, MN 55455   ^^   horton at biosci.cbs.umn.edu       (612)625-8941





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