Fluidigm Access Array - Prepare Sample Pre-MIX and Samples; Prepare the Sample Pre-MIX Solution; Prepare the Sample MIX Solutions

Access Array System for the Ion Torrent PGM Sequencing System: User Guide
Chapter 5: Bidirectional Amplicon Tagging on the LP 48.48 IFC
Prepare Sample Pre-Mix and Samples
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Prepare Sample Pre-Mix and Samples
All DNA samples need to be added into the sample pre-mix individually, prior to loading the
sample mix solutions into the sample inlets of an IFC.
Prepare the Sample Pre-Mix Solution
1 Working in a DNA-free hood, combine the components listed in the table below. This
protocol prepares enough sample pre-mix for 60 reactions. This is enough reagent to
load one IFC with 12 additional reactions to compensate for dead volume and pipetting
error.
Table 8. Sample pre-mix solution
2 Vortex the sample pre-mix for a minimum of 20 seconds and centrifuge for 30 seconds.
Prepare the Sample Mix Solutions
1 Combine the components listed below in a 96-well plate to prepare 48 individual sample
mix solutions.
Table 9. Sample mix solution
Component Volume
(μL)
Volume
for 60 (μL)
Final
Concentration*
* The final concentration in this sample pre-mix table refers to the amount of each listed component in 5 L of
the final sample mix.
10X FastStart High Fidelity Reaction Buffer
without MgCl
2
(Roche)
0.5 30.0 1X
25 mM MgCl
2
(Roche) 0.9 54.0 4.5 mM
DMSO (Roche) 0.25 15.0 5%
10 mM PCR Grade Nucleotide Mix (Roche) 0.1 6.0 200 M each
5 U/L FastStart High Fidelity Enzyme Blend
(Roche)
0.05 3.0 0.05 U/L
20X Access Array Loading Reagent
(Fluidigm)
0.25 15.0 1X
PCR Certified Water (Teknova) 1.95 117.0
Total 4.0 240.0
Component Volume (μL) Final Concentration
Sample pre-mix 4.0
genomic DNA (50 ng/L) 1.0 10 ng/L
Total 5.0