Where did the 2.3 kb band go?

Michiaki Masuda masuda at fcs280b.ncifcrf.gov
Fri Apr 15 18:20:34 EST 1994


In article <CnyLF3.IL9 at ncifcrf.gov> masuda at fcs280b.ncifcrf.gov (Michiaki Masuda)[Yes. That's me.] writes:
>    I've been using GIBCO BRL's lambda DNA/HindIII molecular size marker.
>Recently, I realized that the 2.3 kb band of the marker became fainter,
>and now it's almost unvisible. If that happened to the 4.4 kb band,
>I'd assume that it is due to the cos site annealing and heat it at 65 C.
>However, as I understand it, lambda phage DNA 2.3 kb HindIII fragment 
>wouldn't be involved in the cos site annealing. In fact, heating the
>marker at 65 C for 5 min didn't bring my 2.3 kb back. If it were DNA
>degradation, other fragments should be lost as well. However, other
>bands are clearly visible.

   Shortly after I posted this question, I received e-mail from a technical
assistant person of GIBCO BRL, who had read my posting on the net.
He mentioned that the reason for the disappearance of the 2.3 kb band is
most likely due to fragment-specific denaturation. He also pointed out that
this problem was described in the Focus (a technical bulletin of GIBCO?)
and that it is desirable to use a diluent with at least 20 mM NaCl to avoid
the denaturation.

   Actually, this makes sense to me since [1] the particular marker that
has lost the 2.3 kb band is dissolved in TE + glycerol + dye WITHOUT any 
salt and [2] the problem seems to have aggravated when I heated the marker
at 65 degC. Although I haven't looked at the sequence of the 2.3 kb
fragment, I'd assume that it's AT-rich.

   As several netters sent me e-mail to let me know that they had experienced
a similar phenomenon but do not know exactly why that happened, I thought
it might be better to post this piece of information. It may be common
knowledge to some netters, but I hope that it could be of some help to
others.

   Finally, I'd like to thank the thechnical assistant person of GIBCO BRL
for his quick response. And perhaps, I should mention that I am convinced
that the problem I experienced was not related to the quality of a product
of the specific company. I also thank other netters who shared their
experiences of the missing 2.3 kb band with me.  

--Michiaki

P.S. I am not affiliated with GIBCO BRL or any other company.
==============================================================================
Michiaki Masuda, M.D., Ph.D.
Visiting Associate
Laboratory of Molecular Oncology
National Cancer Institute
Frederick, MD 21702-1021
E-mail: Masuda at ncifcrf.gov



More information about the Methods mailing list