Protocol 21878: End Point Analysis Assay - Gt(ROSA)26Sor<tm6(CAG-ZsGreen1)Hze>-Probe
Version 1.0

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 = ZsGreen1-WPRE

Sequence

Mut Seq:
AACCAGAAGTGGCACCTGACCGAGCACGCCATCGCCTCCGGCTCCGCCTTGCCCtgataagatat
cggccggccctgcaggaattcgatatcaagcttatcgATAATCAACCTCTGGATTACAAAATTTGTG
AAAGATTGACTGGTATTCTTAACTATGTTGCTCCTTTTACGCTATGTGGATACGCTGCTTTAATGCC

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
21306 CTG GCT TCT GAG GAC CG Wild type Forward A
21309 AAT CTG TGG GAA GTC TTG TCC Wild type Reverse A
21310 Fluorophore-1 TAA CCT GGT GTG TGG GCG TTG T Quencher-1 WT Probe
33414 ACC AGA AGT GGC ACC TGA C Mutant Forward A ZsGreen1
33415 CAA ATT TTG TAA TCC AGA GGT TGA Mutant Reverse A WPRE
33416 Fluorophore-2 CGC CTT GCC CTG ATA AGA TA Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
21306 0.40 uM
21309 0.40 uM
33414 0.40 uM
33415 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

This is the only strain that uses this protocol.