Access Array System for the Ion Torrent PGM Sequencing System: User Guide
Chapter 5: Bidirectional Amplicon Tagging on the LP 48.48 IFC
Attach Sequence Tags and Sample Barcodes
34
Table 11. Sample mix solution - SET A
Table 12. Sample mix solution - SET B
IMPORTANT
It is essential to vortex all components to ensure complete mixing.
NOTE
•
The final concentrations of the forward and reverse barcode primers are 400 nM per
well.
•
Each well should receive a unique barcode primer pair.
2
Vortex the sample mix solutions for a minimum of 20 seconds and centrifuge for
30 seconds.
Thermal-Cycle the 96-Well PCR Plate
Place the PCR plates on two PCR thermal cyclers and run the following PCR protocol:
Table 13. PCR protocol for attaching sequence tags and sample barcodes
Component
Volume
(
μ
L)
Sample pre-mix
15.0
Access Array Barcode Library for Ion Torrent PGM
Sequencer Plate A1
4.0
Diluted Harvested PCR Product Pool
1.0
Total
20.0
Component
Volume
(
μ
L)
Sample pre-mix
15.0
Access Array Barcode Library for Ion Torrent PGM
Sequencer Plate B1
4.0
Diluted Harvested PCR Product Pool
1.0
Total
20.0
PCR Stages
Cycle
95°C 10 min
1
95°C 15 sec
60°C 30 sec
72°C 1 min
15
72°C 3 min
1
