so many claims for so many kits -- how can we tell what works?
vando005 at mc.duke.edu
Fri Dec 8 10:50:40 EST 1995
Excellent question. I guess you don'nt know what works until you
try and waste a lot of your valuable time. That is the reason why
most people stick with a method for a long time once they developed
it.Since I had my share of frustration I can give you a few kits that
are routinely used without any magic tricks in our lab.
Whizzard preps from Promega gives you excellent DNA, consistent yield
and fine for sequencing.
Genomed has a kit called Jetsorb which we use to purify out fragments,
length does not matter and it is excellent for ligationreactions. This
in contrast with Geneclean which in our hands gave variable results
with purification and ligation.
Qiagen sells a kit for total RNA isolation called RNAeasy which works
fine to get total RNA from tissueculturecells. The yield is not that
great in our hands but the RNA works fine in downstream applications.
Be carefull if you buy it though you have to buy a Qiashredder kit as
well to go with it so the price of the kit higher than you think.
Have had great succes with both Promega's 5 prime extension kits
and in vitro translation kits. The 5 prime extension kit has a positive
control included that really works (not always the case) so you can
optimize your own reactions.
TA cloningsystem from Invitrogen of course never fails it is expensive
and if I ever are going to have time we develop our own T vector.
Large scale plasmidpreps we do manually (ie no kit) these days after
trying out a lot of kits and not getting consistent results. And, yes
I read all the troubleshooting sections (sigh).Too expensive for too
little DNA and if a salesrep wants me to buy their kit I need to try
it out for free otherwise no chance!!
Gibco's gelsystem for acrylamide gels for western blots works fine, cheap
enough so you do not have to make your own buffers.
Promega mRNA kit Poly A track magnetic beads gave me fine mRNA but
one thing I noticed is that your starting material has to be a lot.
It does not work very well with little RNA even if you follow their
small scale protocol.
While I'm at it I also would like to mention a problem with oligo's
for PCR, not a kit but worth knowing. We have switched companies 3
times before we found one we liked.(and from what I hear this is not
uncommon for other labs) Gibco sells excellent oligo's at a good price.
Have used a lot and not found any mistakes. Mistakes
from other companies included missing bases, short oligo's, customer
service that did not believe their oligo was wrong even if you told
them you sequenced over it.(Which took you weeks of frustration and still
had to start over)
Hope this helps an I have no affiliations with any of these companies.
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