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missing 500 bp band w/ lambda hind3

Song Tan tan at aeolus.vmsmail.ethz.ch
Fri Jul 8 07:05:49 EST 1994

In article <1994Jul7.155930.11658 at leeds.ac.uk> futers at biovax.leeds.ac.uk
>In article <2vh39l$k3r at mserv1.dl.ac.uk>, (David Johnston) daj <daj at nhm.ac.uk> writes:
>>On Thu, 7 Jul 1994 13:03:27 GMT,
>>>We need some help.  We're running 1% and 2% gels in our lab using Lambda hin3 
>>>as a molecular weight marker.  We load 500 ng of lambda in our marker lane and 
>>>we don't get resolution of the 500 bp band when prestaining the gel with etbr.
>>>Do we need more DNA in the marker lane?  More etbr (we now use 2ul/50ml of 
>>>gel, this used to be enough).  Any comments/suggestions will be appreciated.
>>If my memory serves me correctly, the 564 base fragment and the 23130 base 
>>fragment contain the 2 cos sites/sticky ends of the phage DNA and anneal 
>>together. Heat marker to 65C for 5 mins and cool rapidly on ice before 
>>loading to dissassociate the 2 fragments.
>It is the 23130 and the 4361 bands that have the cos sites.  The 564 band is
>weak due to being small.  If you need to have a size reference around 500, I
>would suggest using a different marker such as the 100bp ladder.
>               Simon Futers (futers at biovax.leeds.ac.uk)

This is only tangential to the above discussion, but I'd like to mention that
HindIII+EcoRI doubly digested lambda makes a very nice molecular weight marker
for < 5000 bp fragments.

HindIII lambda is 23130 + 9416 + 6557 + 4361+ 2322 + 2027 + 564 + 125 bp.

HindIII+EcoRI lambda is 23130 + 5148 + 4973 + 4268 + 3530 + 2027 + 1904 + 1584
+ 1375 + 947 + 831 + 564 + 125 bp.

Having the 1584/1375 bp and 947/831 bp pairs is quite useful.  

Song Tan
Institute for Molecular Biology and Biophysics
ETH-Honggerberg (Swiss Federal Institute of Technology)
8093 Zurich, Switzerland
email:  tan at aeolus.vmsmail.ethz.ch

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