Genomic DNA Isolation - Method Required

Andre Hamel hamel at ccu.umanitoba.ca
Tue Mar 30 10:37:11 EST 1993


There's recent articles in Biotechniques describing use of formamide to
"dissolve" whole blood/clotted blood, tissues, etc ... simply take about 1
to 10 uL worth of sample (blood/tissue/culture cells, etc), add to 100 uL
formamide, mix, heat 95oC for few minutes, then add aliquots (up to 20 uL)
to 100 uL pcr rx'n mix. Taq pol WON't work for this though, must use Tth
pol (Perkin Elmer, US Biochemical plus several other vendors), Pfu pol
(Stratagene) or Vent polymerases (New England Biolabds) ... remember to
DECREASE the step temperatures according to formamide concentration ...
for example, if final formamide conc. in pcr rx'n is, say 10%, then reduce
each pcr step temp by about 5oC to 10oC.

If you're after the genomic DNA of the host from it's plasma I doubt if
this would be adequate starting material ... one needs enough nucleated
cells (lymphocytes from whole blood or from the blood clot, tissue, cells,
etc) in order to provide source of genomic DNA.

Andre Hamel
Manitoba Vet. Virol.
Winnipeg, Manitoba
CANADA

email: hamel at ccu.umanitoba.ca



In article <1993Mar24.153536.1 at vcp1.vcp.monash.edu.au> michael_m at vcp1.vcp.monash.edu.au writes:
>Dear Netters
>
>I would like to make a genomic DNA preparation from blood and would like to
>know if anyone has a foolproof (necessary!) method.  I would also like to
>know if it is possible to use plasma (such as that obtained from the Red Cross)
>for the isolation.  I hope to use PCR to amplify my gene from this DNA.
>
>Thanks in advance.
>
>Lukim yu
>
>Mike
>--------------------------------------------------------------------------------
>Michael J McLeish Ph.D.  		
>School of Pharmaceutical Chemistry	   Phone  61  3  3899540
>Victorian College of Pharmacy     	   FAX    61  3  3899582
>381 Royal Parade Parkville 3052		   Email  michael_m at vcp.monash.edu.au
>AUSTRALIA
>--------------------------------------------------------------------------------



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