question on carrier salmon sperm DNA - (Oct/02/2005 )

Hi,

im a newbie to this site....dunno if anyone can help:

i have bee using salmon sperm (fragmented i think) in a yeast transformation as a carrier during DNA precipitation with alcohol..... but exactly does it help to precipitate plasmid DNA?

amy.

Precipitation is more efficient from concentrated solutions than from dilute solutions...

I personally haven't used a carrier molecule in years (and never did use one routinely), but on the rare occassions I did, we used RNA or oyster shell glycogen.

hi
homebrew is right. Dna molecules are supposed to precipitates and form aggregates. That's why ssDNA is used to enhances this process and make quite a capture of the wanted DNA. ssDNA is fragmented and don't interfer in tranformation of bacterias. You can use tRNA as a carrier too or glycogen in cases of you have few material and want it remain pure.

fred

thanks for all your help!!!
amy.

tRNA in ligation reactions

The use of yeast tRNA as a carrier for ethanol precipitation of DNA carries an inherent risk of cloning yeast genomic DNA fragments that frequently contaminate tRNA preps. In addition, tRNA inhibits ligation/transformation. Glycogen is safer, but since it is isolated from mussels, it also carries the risk of contamination by DNA. Linear PolyAcrylamide is totally inert and does not carry this risk.

tRNA inhibits ligation

tRNA used as a carrier in ethanol precipitation will inhibit ligation reactions. Slight inhibition of colonies per µg DNA at 500 µg/ml tRNA and significant inhibition at 2500 µg/ml. Glycogen shows no inhibition at 1000 or 5000 µg/ml. (my data 1986).

Presumably, salmon sperm DNA carries a similar risk.