PLASMID DNA yield - (Mar/22/2007 )
i did a miniprep for 2 different plasmids* the same gene inserted in 2 different vectors* using promegs kit , one yield 0.8 ug and the other 0.13ug??????
can anyone explain to me why is that difference?
one collegue here said may be because the difference between the 2 vectors?
i just want to know is this something wrong i did?
after miniprep by the way i usulallt do phenol,chloroform and ethanol precipitation then resuspend in TE and measure my DNA.
your colleague is right.
Yields depend on the vector backbone, high copy vs low copy plasmids.
Small amounts of phenol contamination can throw off spec readings of DNA concentration rather dramatically. Make sure your DNA has had the phenol removed. Extract an extra time with chloroform and then wash the pellet an extra time with 70% EtOH.
The phenol also (more importantly) is an inhibitor of most enzymatic reactions you want to do with your DNA.
You might want to check DNA concentrations by comparing band intensities on a gel.
thankx for you replies, it really helped.