Sequencing

Jens Tornoe imjt at pop.dtu.dk
Tue Nov 16 17:52:40 EST 1999


In our hands, automated (fluorescence) sequencing can cover up to 700-800 bp
per primer. Still, you would have to be lucky to be able to sequence your
entire 1,5 kb fragment. You would need two entirely perfect reactions to do
that.

Manual (radioactive labeling+film) sequencing only yields the mentioned ~400
bp per primer. And it is a drag to read...

My advice would be to send off your vector to sequencing with two flanking
primers, see how far you get and then buy an additional internal primer to
get the last bit. It is not that expensive. You can buy a usable sequencing
primer at Life Tech. for ~$10. If the fragment has never been sequenced
before, you would need to sequence both strands anyway, to be completely
confident in the result.

Regards,

Jens



Dr Raja Kota <rkota at scu.edu.au> wrote in message
news:rkota-1611991232290001 at ip3956.scu.edu.au...
> Hi Netters,
>
>    Whats the best way to sequence a fragment which is over 1500 bp in
> length and is cloned into a vector.  Any references shall be appreciated.
>
> Thanks in advance,
>
> Raja






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