PCR in ten minutes!

Ted M. tedm at darkwing.uoregon.edu
Tue Jul 25 00:28:55 EST 1995


In article <jpcd0-2207951135550001 at mje-mac3.welc.cam.ac.uk>,
jpcd0 at mole.bio.cam.ac.uk (John Dixon) wrote:

> Hi there,
> 
> has anybody out there tried the 'Rapidcycler' from Idaho technology?
> 
> John Dixon 

Hi John,
   Yes I have tried the machine.  My last Lab at the University of
Washington purchased a Rapidcycler from Idaho technology and I was the
primary user. It is indeed true that 30 cycles take ~10-20min. The machine
heats with a 300watt halogen lamp and cools via venting to ambient air.
There is no capability for temps lower than ambient, but with that speed
there is little need. The trap door which opens with a "POP" when it cools
scared the pants off new users! and the pulsing glow of the lamp within
the machine is very eery in a darkened lab... 
   The main advantage of the machine is the amazing specificity of priming
that can be acheived with very short anneal and denature times; reactions
from genomic that are messy smears will become tight bands. Another
advantage is the short times means several serial experiments exploring
optimal anneal temps are posssible in one day, and with one pooled
reaction mix. The machine is optimized for 10ul volumes although it was
scaled to larger tubes (50ul) in our lab. The sample handling requires
practice and patience. The sample prep may take 1hr, as the samples are
loaded into the capillaries and heat sealed. Using 32P is nerve racking as
there is a very real possibility of breaking a tube and contaminating the
machines interior chamber.The use of the capillaries requires either a
very high BSA content in the buffer, to avoid surface denaturation of the
enzyme, or the use of siliconized capillaries, which negate the helpful
capillary action of the tubes. The company even includes a buffer with
BSA/ficoll and tartrazine dye to facilitate loading the reaction directly
into a gel well, I liked this method..
   There exists a ref. (Biotechniques? TIBS?) which is the origin of the
idea of an air cycler. It would not be impossible to make your own
machine. The trick would be to tune the incredibly fast PID to hit the
temperatures and to program the EEPROM, many undergrads in physics could
probably do this with a budget of ~$1000. The Idaho machine cost ~$4000, I
belive and was the size of a typical Perkin Elmer PCR machine (not a
9600!)  Feel free to Email me with any specific questions.
                                                Ted Michelini
                                                Institute of Molecular Biology
                                                University of Oregon
                                                tedm at darkwing.uoregon.edu



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