DNA size standards, odd bands, & polyA gels
dbell at qnis.net
Thu Aug 9 13:05:09 EST 2001
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
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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
: 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,
: bp, 100 bp, but the 300 and 200 bp bands may be *closer* together than
: 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:
: Can anyone explain this?
: Thanks in advance for any insight,
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