Literature on making vectors

Glenn Jenkins gjenkins at facstaff.wisc.edu
Fri Sep 6 21:44:46 EST 1996


I don't think you will find a reference on this specific subject. If you
do, then post it so we can all see it. Most of this information is picked
up through experience and practice.

Generally speaking, stickys ends are the way to go. PCR is useful but like
all tools, it has its limits and should only when fitting. An ATG is
required since it tells the translation enzymes where to begin. And the
start site does not need to be 'in frame' with gene but the sequence
following the start site does. The length between the promoter and the
start site varies but there is an optimal length (anybody know what it
is?)

If you are unsure about your stratagy, then post it and we can offer
specific suggestions.


Glenn Jenkins



In article <randyg-0609961626150001 at crl228-200-217.crl.umn.edu>,
randyg at puccini.crl.umn.edu (Dr. Randal W. Giroux) wrote:

> We are in the process of starting to make vectors for plant
> transformation, to express particular genes of interest. We have some
> backbone vectors, some really great promotors and some interesting genes
> that we would like to transiently express. However, we are short on
> information on the pitfalls and tricks for vector construction. Could
> someone tell us what papers or publications exist about constructing
> vectors. 
> We are concerned about things like, blunt end vs Sticky end ligations, To
> PCR or not to PCR, if we have the whole gene (ie the ATG start site) do we
> need to be 'in frame' with the promotor? These are some of the questions
> that we are concerned about.
> 
> Any help or pointing in the right direction would be much appreciated. 
> 
> 
> Randy



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