FW: DNA fingerprinting for yeast

Deanne Bell dbell at fresno.ars.usda.gov
Fri Feb 6 10:48:10 EST 2004



-----Original Message-----
From: owner-methods at hgmp.mrc.ac.uk [mailto:owner-methods at hgmp.mrc.ac.uk]
On Behalf Of dav668 at yahoo.com
 and followed their recommended PCR conditions. But after running the
gel electrophoresis, i can only obtain smear for each lane, which were
loaded with the amplicons, instead of getting some bands. 



SMEARS USUALLY ARE DUE TO NON-SPECIFIC ANNEALING OF THE PRIMERS DURING
PCR.
SOME IDEAS TO START YOU OFF:
1)	 OPTIMIZE MgCl BY RUNNING A GRADIENT OF REACTIONS RANGING FROM
1mM - 6mM FINAL CONCENTRATION.

2)	OPTIMIZE ANNEALING TEMPERATURE OF PCR

3)	USE A 'HOT START' PCR METHOD 

4)	MAKE SURE YOU ARE USING GEL % THAT IS OPTIMAL FOR THE LENGTH OF
YOUR PCR PRODUCT
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