From GenoSeq

Cycle Sequencing Reaction for PCR Template

template	x1 µl	<200 bp	20 ng
		200-300 bp	50 ng
		>300 bp	90 ng
BigDye Terminator Mix	2 µl
Sequencing Buffer	2 µl
primer (10 µM)	2 µl	(20 pmol)
water	x2 µl
Total volume	10 µl

The above guidelines are from the manufacturer.

In actual practice, using an aliquot

from your cleaned PCR reaction usually works fine.

There is no need to dilute the sample further.

1 µl of BigDye is more than adequate for each reaction.

Adjust the volume with water.

Cycle Sequencing Temperature Profile for PCR templates

repeat the following for 25 cycles
96 °C	10 sec
50 °C	5 sec	(50 °C -58 °C depending on your primer's Tm)
60 °C	4 min
rapid thermal ramp to
4 °C

primers the core provides:

M13F primer: -21 M13F : TGT AAA ACG ACG GCC AGT

M13 R primer: -28 M13R : AGG AAA CAG CTA TGA CCA T

T7 primer: TAA TAC GAC TCA CTA TAG GG

T3 primer: ATT AAC CCT CAC TAA AGG GA

SP6 primer: ATT TAG GTG ACA CTA TAG

BGHR primer: TAG AAG GCA CAG TCG AGG