Stock No: 005304
Protocol 21988: End Point Analysis Assay - Kif1a
Version 3.0

Notes

The genotyping protocol(s) presented here have been optimized for reagents and conditions used by The Jackson Laboratory (JAX). To genotype animals, JAX recommends researchers validate the assay independently upon receipt of animals into their facility. Reaction cycling temperature and times may require additional optimization based on the specific genotyping reagents used.

Expected Results

Mutant = T
Heterozygote =C/T
Wild type = C

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
15616 Fluorophore-1 CCA CGT GTT CTG GTT TAA CCA Quencher-1 MUT Probe
18219 TAG GTT GGT TTG CCT CCA TC Forward A
18220 CCA TCT CCT GCT TCA TGT CA Reverse A
18221 Fluorophore-2 CAC GTG TTC CGG TTT AAC C Quencher-2 WT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
18219 0.40 uM
18220 0.40 uM
Wt Probe 0.15 uM
Mutant Probe 0.15 uM
DNA

Cycling

Step Temp °C Time Note
1 95.0 --
2 95.0 --
3 60.0 --
4 -- repeat steps 2-3 for 40 cycles
5 4.0 -- Forever
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.

Strains Using This Protocol

Stock Number Strain Name
017473 C57BL/6J-Kif1algdg-2J/CxJ
005304 C57BL/6NJ
2 strains use this protocol