TTE gels

Glenis Wiebe gwiebe at ucalgary.ca
Thu Jul 13 17:36:10 EST 2000


Hi Kev,

After we tried the TTE gels, we never looked back.  Yes, the results are
that much better. (My thanks to George Grills for getting us started.)

Here are the basics:

GEL (5.3% Long Ranger, 6M urea, 1.2X TTE)
21.6 g urea
6.4mL 50% Long Ranger gel solution
7.2mL 10X TTE
final volume 60uL

30 uL TEMED
300 uL 10% ammonium persulfate

UPPER BUFFER - 2X TTE (700mL)
140mL 10XTTE
560mL dH2O

LOWER BUFFER - 1X TTE (900mL)
90mL 10X TTE
810 mL dH2O

10X TTE (1L)
121.65 g   TAPS (N-tris[hydroxymethyl]methyl-3-aminopropane-sulfonic
acid)  (Sigma)
60.55g   Tris
3.75g   EDTA
dH2O to 1L

RUN CONDITIONS
EP Voltage: 4000V
EP Current: 60mAmp
EP Power: 40W
Gel Temp: 42C
Laser Power: 40mW
Run Time: 16 hrs

We use the long read basecaller, LR-377, from PE (?) Applied Biosystems
3.3.1b2 (available from their web site), with Sequencing Analysis version
3.4.  We aim to have a gel spacing of approx. 14.

I hope this works as well for you as it has for us.  Let me know if you
have any questions.

Sincerely,
Glenis

--
Glenis Wiebe, M.Sc.
University Core DNA & Protein Services
University of Calgary

tel: (403) 220-4503, fax: (403) 283-4907
e-mail: gwiebe at ucalgary.ca
http://www.ucalgary.ca/~dnalab



Kev wrote:

> Can anyone send me the recipe for TTE gels for the ABI377 sequencer.
> It's supposed to be in a user bulletin but I haven't received that one.
>
> Any good or bad experiences with them?
>
> Thanks
>
> Kev
>
> ---







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