*Why are you just beating about the bush? I had mentioned that I told optimal vector: insert ratio additionally, as your supporters found nothing new in what I said.
And please dont explain to me all that mumbo jumbo of ligation etc.
If it doesn't help you, just don't follow it.. It better to start with something that you know than just making a wild guess.
Secondly, if you are really interested on knowing about Avantageand long Pol, just give a google search.
Now, Reitaliating what you said:
If you used proofreading polymerase, it
would have to be A-tailed first. If Taq and the like, no need to.
These days, enzyme mix come withproof reading enzymes in them. therefore, just after PCR,if you want to continue just gel purify the product or if, to be used later, it should be quickly kept in -20, so that proof reading enzyme in that will polish away 3'A left by Taq.
I hope I made myself clear now.
On Sun, 28 Dec 2008 DK wrote :
Protein science Lab
Dept. of Biological sciences
National University of Singapore