DNA size standards, odd bands, & polyA gels

[dbell]

Hi Russell

I started thinking about this several months ago when I was looking into 
running microsats and AFLPs on PAG.  All of the markers I used for DNA were 
ran on agarose and I wanted to know how they would react on PAG - 
especially on denaturing type of gels (6-7M Urea).

I think the bands get all funky because of how they are made (some ladders 
are dsDNA cut by enzymes, some are constructed . . . etc)

Through all of my research I decided to use a sequncing reaction as a 
ladder for my microsatellites (Silver Sequence DNA Sequencing Sytem by 
Promega is the one I found).  And I am staying with my automated AFLP 
system.

Deanne Bell
USDA Agricultural Research Service
2021 South Peach Avenue
Fresno, CA 93727
(559) 453-3170
(559) 453-3088 fax

On Thursday, August 09, 2001 9:06 AM, "Russell S. Pfau" 
[SMTP:pfau at tarleton.edu] wrote:
: I've recently begun using a 20cm x 20cm x 1.5mm polyacrylamide gel format
: for SSCP and microsatellite analyses. Two things have happened regarding
: the appearance of the DNA size standards that I can't explain, and I'm
: wondering if anyone else can.
:
: One size standard, a 100 bp ladder from New England Biolabs, exhibits 
bands
: that are not separating as they normally should. For example, with this
: size standard there are bands of the following lengths: 400 bp, 300 bp, 
200
: bp, 100 bp, but the 300 and 200 bp bands may be *closer* together than 
the
: 400 and 300 bp bands, the opposite of what should be true. Also, some of
: the bands are missing! From what I can tell, bands 1517 & 1200 are not
: there at all!
:
: The other size standard I'm using, a 25 bp ladder, exhibits quadruplets!
: Four bands for each size category.
:
: Images of my gels can be viewed here:
:
: http://www.tarleton.edu/~biology/pfau/images/odd_bands.htm
:
: Can anyone explain this?
:
: Thanks in advance for any insight,
:
: Russell
:
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: 

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