sequence for gt10 cloning region

Dr. N.A. Affara naffara at crc.ac.uk
Wed Aug 12 06:04:30 EST 1992


In article <1992Aug11.162605.8225 at pollux.lu.se>, Bo Servenius <biogen10 at gemini.ldc.lu.se> writes:
> Dear Netters:
> 
> Anyone that could give me a tip about where to find the
> sequence for the cloning region of lambda gt10 vector.
> Need some 100bp up and down from the EcoRI. 
> 
> Even better would be to get besides the vectorsequence
> the sequencing of some PCR and  sequencing primer sets
> that could be used for solid phase sequencing.
> 
> 
> Happy  for any comment
> 
> BOSSE

We use PCR primers made from the sequence shown in the NEB catalogue. It shows
about 100(?) bases with the RI site in the middle. Instead of using the 
primers they suggest we designed our own with both starting from the first
(and last!) base shown.
I think they were about 20 bases long. Can't remember off hand.
For sequencing, we made primers about 10-15 bases inside the PCR primers.
We use these to sequence cDNA clones. They work perfectly. (well, almost :-) )
If you like, E-mail me and I'll send you the sequence of our oligos.

BTW We use one primer biotinylated in the PCR so we can sequence it directly.
e.g. F+BIO and R. Another tube with F and R+BIO.

Hope this helps,
Phil
(E-mail naffara at uk.ac.crc)

-- 
Kryten (When Cat says why don't we raise the defensive shields?) "A superlative
suggestion sir,with only two drawbacks: one,we don't have any defensive shields
and two,we don't have any defensive shields.I know that,technically,that's only
one drawback,but it was such a big one I thought I'd mention it twice".



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