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)
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
dH2O to 1L
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.
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.cahttp://www.ucalgary.ca/~dnalab
> 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?