Stock No: 021830
Protocol 21998: End Point Analysis Assay - Lrrk2<tm4.1Mjff>
Version 2.0

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 = C/C
Heterozygote = A/C
Wild type = A/A

Sequence

GTTTTTTGATAAGACCTCCTGTGTACTCTCTGGCAGGTATCTCCACTCAGCCATGATTATTTACCGTG[a/c]CCTGAAGCCCCACAATGTGCTGCTTTTTACCCTGTATCCCAATGCTGCC

ATCATTGCGAAGATTGCGGACTACGGGATCGCACAGTACTGCTGCAGGATGGGAATAAAGACATCAGAGGGCACCCCAGGTAGGAGAACAAGTTTACTTTTTTTGCAGTGGACCAT

CAGTGCCCTCAAGTGT

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
17631 ACT TGA GGG CAC TGA TGG TC Reverse A
20130 GGC AGG TAT CTC CAC TCA GC Forward A
20131 Fluorophore-1 TTT ACC GTG ACC TGA AGC C Quencher-1 WT Probe
20132 Fluorophore-2 TTA CCG TGC CCT GAA GC Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
17631 0.40 uM
20130 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 40.0 -- Forever
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.

Strains Using This Protocol

This is the only strain that uses this protocol.