Protocol 40042: Probe Assay - Lilr4b<em1(IMPC)J> Alternate4
Version 1.0

Notes

Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.

 

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

>chr10:51491479+51491581 103bp TCGCTGCGTATACATCTGTG ACTTCAGTGTCCCAAGGATTTAC

Mut = 122 bp

Wt = 103 bp

Fam = Mut

Hex = Wt

Sequence

Wt Sequence:

 

Mutant Sequence:

This mutation is a 137 bp deletion beginning at Chromosome 10 position 51,481,213 bp and ending after 51,481,349 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
52948 TCG CTG CGT ATA CAT CTG TG Common A
52949 ACT TCA GTG TCC CAA GGA TTT AC Wild type Reverse A
52952 Fluorophore-1 AGA GTG CTG CTG GCT TCT CA Quencher-1 MUT Probe
54744 TTC CCT GAT GTC CTC TCA CC Mutant Reverse A
55122 Fluorophore-2 ATC TGG TGT TGG GGT TCC TG Quencher-2 WT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
52948 0.40 uM
52949 0.40 uM
54744 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.