You can find these on the respective product page, section downloads.
Yes, you can order individual primers from the SBTexcellerator® HLA Class I, HLA-DRB1, HLA-DQB1 and HLA-DPB1 Extended kits.
They are stable for over a month and do not require refrigerated transport. For long-term storage however, we recommend storing them at -20°C
Primers are stable for at least 5 months after reconstitution in nuclease-free H2O, when stored at -20°C. Primers can also go through at least 25 freeze/thaw cycles without any negative effects on the results.
You can stop the SBTexcellerator protocol:
- After amplification PCR
Store overnight at 15°C in thermocycler or 4°C in fridge. Continue with ExoI/SAP clean-up preferably within 24 hours but definitely within a week after amplification.
- After clean-up by ExoI/SAP
Store overnight at 15°C in thermocycler or 4°C in fridge. Store for 1-2 weeks at 4°C. Store for >2 weeks at -20°C.
- After sequencing PCR
Store for 1-2 days at 15°C in thermocycler or 4°C in fridge.
- After clean-up by Sephadex
Store for 1-2 days at 4°C. Store for >2 days at -20°C. After a longer period of storage it is recommended to denature the reactions by putting them in a thermocycler for 2 min at 98°C and afterwards directly transfer the reactions to a cooling block or ice before running the plate on the sequencing instrument.
IMPORTANT: During storage, always keep your plates sealed to prevent evaporation!
The PCR primers of the SBTexcellerator HLA-A kit are not multiplexed but are a single set of amplification primers positioned at the UTR regions yielding one large PCR amplicon.
HLA-B *18 is well known for this. It has an InDel in front of exon 2 making it impossible to read through into exon 1. To read exon 1, you need to use the special exon 1 primers.
For DQ-02 dropouts
Use more LongRange PCR enzyme (0.8 µl instead of 0.4 µl/rxn).
For DQ-03 dropouts
Fresh Q solution usually works. However, if this doesn’t work, try 0.8 µl enzyme as above