making your own size markers

"Jerry Marshall ", Y jerry at PICAN.PI.CSIRO.AU
Wed Feb 8 18:25:25 EST 1995


Here's a method for making your own low MW DNA size markers:

Buy, borrow or otherwise procure a microlitre of Pharmacia's 100 bp 
ladder. This product is made by partial digestion of a plasmid that 
contains lots of 100 bp fragments separated by AvaI sites.  Use your 
microlitre of the ladder to transform your favourite E. coli strain 
(there's still some intact plasmid there since the ladder is only 
partially digested).  Purify the plasmid and do a partial digest with 
AvaI.  It helps to do a pilot run taking timepoints to allow you to 
optimise conditions for production of a nice ladder (overdigestion gives 
a nice plasmid band and a nice band at 100 bp...).

The Pharmacia product is spiked with a DNA fragment of 800 bp to allow 
you to orient yourself after you've run your DNA off the bottom of the gel.  
This can easily be reproduced by adding a restriction fragment from some 
other clone or a PCR product.

The whole procedure is very straightforward and considerably cheaper than 
buying the product from Pharmacia.

Thanks to Tony Ashton for the idea.




More information about the Methods mailing list