Ethanol precipitation without using salts
Posted 29 December 2010 - 10:02 PM
Posted 30 December 2010 - 01:16 AM
Also, additional advice: if you are doing "home-made mini prep" (not the one with commercial kit and silica membrane), be sure to dry the EtOH before you add TE-buffer(or water) for storing the plasmid, so it would not interfere as well..
Hope I helped a bit.
Posted 30 December 2010 - 04:50 AM
Posted 31 December 2010 - 01:26 AM
I see... I want to sequence the insert.
I think it depends what you will do with your plasmid later 'cause salt can interfere with later steps. I am using the ice-cold EtoH+centrifugation for precipitation of the plasmid.
If it is the supernatant from spinning down neutralized cell lysate, then you don't need to add salt. If it comes from (for example) elution of a column with water or TE, then you do need to add salt.
I am using plasmid eluted with TE from column. And i didn't add salts as the miniprep supplier told me i don't have to Can i add the salts after abs EtOH is added? Or is there anyway to retrieve my plasmids from abs EtOh?
I read in some online articles that say vortex when wash DNA with EtOH, but some ppl say vortex only if RNA. Should i vortex or not? IF DNA can break in vortex, why can't RNA?
Edited by hianghao, 31 December 2010 - 01:35 AM.
Posted 31 December 2010 - 06:08 AM
Vortexing plasmid DNA makes little difference, since it is physically small. Vortexing genomic DNA, which can be centimeters long if stretched out, will break up the DNA. RNA is usually quite small, so there is little issue with vortexing RNA.