Stock No: 000533
Protocol 18274: End Point Analysis Assay - Ghrhr<lit>
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

WT - A/A
Het - A/G
Mut- G/G

 

 

Sequence

tcagaatctcctccctcataggttaccccatgtctcctccttctccacctagGATGCCCTGGGACCTGGGa/gTGGGCTGCTGTGCTGGCCCCCGACAGGCTCTGGCCAGTGGGTCTCTCTCCCCTGCCCTGAATTCTTCTCTCACTTCGGCTCAGACACAGgt

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
17752 TCA GAA TCT CCT CCC TCA TAG GT Forward A
17753 CTG TGT CTG AGC CGA AGT GA Reverse A
17754 Fluorophore-1 GAC CTG GGA TGG GCT GC Quencher-1 WT Probe A
17755 Fluorophore-2 GAC CTG GGG TGG GCT GC Quencher-2 MUT Probe A

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
17752 0.40 uM
17753 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 -- repeat steps 2-3 for 40 cycles
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
010816 B6;C-Ghrhrlit Prkdcscid/BmJ
005764 C3.B6-Ghrhrlit/J
000533 C57BL/6J-Ghrhrlit/J
3 strains use this protocol