Protocol 33700: Probe Assay - 1600014C10Rik<em1(IMPC)J>
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

>chr7:38197750+38197838 89bp CCCAGTCCTGTGTAGGCAGT TTAGACCAGCACCTCCCTCA

Mut= 104 bp

Wt= 89 bp

Fam=Mut

Hex=Wt

Sequence

Wt Sequence (deletion in lower case):

 CCTTTCCAGGCAAATGGCTTTCCCCAGAAGATGCACGTCCCTCTCGGCTTCTCCTTTATGGAGACAAATTGCTCTGGGAGCCACAGCGGGTGGGGGTGGGGGGGTGTTCCTCAGTCATGAAGGGGATCTTAGTTGGCacagttctgaggtagggagggtgtaagtgaggggcattgggaacaaccaatttgagtatcaattatgtggaccaaatggactagagaccacatcaacctttgccctgcagagtgcctggcagaagctgggcgtgctgtgacctggtgaatgtccccacttgtcatgactctcagcaagaaacaaaggccattgttggggaatgaggctgactaattgagcccttccttgttttccaggggggaccgttgggggcctgttaggtgcctggatgacgagtggacagttcaagcccgttcctcagatcttaatggagctgcccccccgctgaacagcggaaacttgttaacgaagccatggccatcatcgggaacttggactggacagacgcagtgcagctgactgctctggtcatgagcaaccaggccatgcagcagaggctcttggccatgctaacgacctatgtcaccaaggagctgcaggctgagatccgctatgaggactaggccccgccccctggggagtggggctctcattttagatgattctatggctcattgaaggtgaccagggaatgggggaagcctgacttctgggaaacttgtctaataccagcattgattgtaagccgccatgctagtgcattctgttggaccgtcctgctggagaggcatcagccatgcctcttgggctgtgtgtgctatggaagggtacaggcgtgcctgctgtaggcattcacggacagcttggagtcctcccgatgaccatgtgggtctaaaaggttctcatgttgttggctggccgaccatagcagatgcttctctttctgtgaagtgtacctccaggtacctgtcacaggatcgaatccccttagactctggggtcttggggttcagagccagctcttgtcaggattgtgctaacctgctgagtgtttccttcccttggctaaggacttcccagtggtctcatggtgacaactgtccctcctttgagtgaagccacagatgccaggcaggttctattctctgctacaggctatgccctagacgctcttgagtaactttagtttagcttgtaggagcttcttctgcctccttaagaaaaatcatggcacttcattatattttattttccatgtcgattctggactctttttgttctctccactttcgtagcctaaattcatttagctctgttttctttttcttcttactatgtttccttgggctagccttgaagtcctgggctcaagctacatagctgggactgtaggcttgtgccaccatacctagctttacctcaaagactgttgtccactctacttttcctaagccctcccgtgttctcctgtctgctggctgcaatctataattcatttgtgtgaaaagacaaaattaccaatgacttgagttcagagatcctaacatagctaggtgctatggtgcagacccatacccacggtcccagataattgaaaagctgaagggggaacatccttgggcccaggaatccaaggacaaccggacattgtggaagccgggagataggtggggacacacacatcttcttagttagcttttatttgcagtcacggaatcaagaggcagcaagtgccctggtgagacggagagagagggtggacaagggcgagcaagggaaagtggaaacagcaagaaaactagattaacctcccgtgacactggctgtttatggattgggctagcctcgcctccatttccctaagtgacggaggctcaaatctgacattagcttgtttgttttccttgtgttgtggatggaagccagggccttgcataggctaagtgccacccctgagcgctgctcctagtcctcctctctgctgattcccgcccagcctggtcccagcccttgagtgactcagctttggtctggtcggttggtcccagtgctggagctcaacatctaacattgtatccatttgattaagtatttgatacttcctccttatccctccttggaggtgctcactcactgagggcaggagctatgtgtctcggtcacttctcctgcctccagcttgctcctgcctggcaccatggcttttgtggggtagacactcaataaatatatatttttaaagctatgtatttcatatctttctgtttgtttgtttatttatttatttatatctttacttatttattttacatgttttgtctgcatttatataagtacgatgtatgtgtctggttcccacagaggccacaggaggcactggacctcctggaaacgatgttactgacaattgtgagccgccatgtgggtgctaggaactgaacccaggccctccattagggcatcaatggctaaacaatctccctcctctaggtttgggtttttaaacctgggtttctccatgtgattctggttggcctcaaactcctggtctttcttcctgcctcagccccttgaactcggagactacaggcaggcatttccaagccaagacaaatacgttttgttgttgttagaagtctttaagttcttattgtttgccttttctttgtttagttttttgtttgtgtttgtgttttgttttgttttgagacaggatctcatgtagcccaggctagcccccaactctatgtagccaaagacgaccttgaactcttgattctccagcctccatcactgccaagtgttgggaattacagagataatacctttatgtgagacttcagtaggtatttttggaagttattgttctgaaatcacttcctgtggaattcctccaggaaggaaacatgtgacaggagagacctagaatccagttttaagctttttgaaaccactgagctgctagccacggaagctggagtttggcctggtgtccctagaacccagagcctcgcttgagcaccatgtggtccctcagttttcacaaacaggtttgggacatgttttcacatgaaatagttttagtcaaatgtatgctgtctctgaagtggggtatcccaacctcagatagttccaccccaaggtatcccccatgctatatcctttcagaactactacccgactatcatcaaattactaataaattacttttccctgaaaaagtcactctctgtgtattttgtggaaaccacttctgaagcctttaacatcaaaaggaagattgctccatctgaggcttgacaggacaggtaaaagctccctggcccggttcgtggcggccatttctggccagcatgcgccatgttactgcaagccatgtttaacctccaccagccctccagccccagtcctgtgtaggcagtgctgggaccggggctctctgtgtggcggcagcccagACTTTCTGCAGTCTGAGGGAGGTGCTGGTCTAAGGGCCAGCTTTGCAGTACAGATCTCTCTGTGTAAAGGAAGCCTCCTGTGGACTTCACCCCATGGGGATAAGGCTTTGTCTTACAGTCTGCCTCAGTGA

This mutation is a 3,339 bp deletion beginning at Chromosome 7 position 38,194,467 bp and ending after 38,197,805 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
42000 CCC AGT CCT GTG TAG GCA GT Wild type Forward A
42001 TTA GAC CAG CAC CTC CCT CA Common A
42002 AAT TGC TCT GGG AGC CAC A Mutant Forward A
42003 Fluorophore-1 CTC TCT GTG TGG CGG CA Quencher-1 WT Probe
42004 Fluorophore-2 ATC TTA GTT GGC ACT TTC TGC AGT Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
42000 0.40 uM
42001 0.40 uM
42002 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.