Stock No: 040180
Protocol 40106: End Point Analysis Assay - Hdac8<em1Lutzy>
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

>chrX:102504962-102505067 106bp GAGATGGAGATGCCAGAGGA GGACCTTCACAAGGGAGTCG

Mutant = G, G, C

Wild type = T, T, T

X-Linked

Sequence

AGATGGAGATGCCAGAGGAACCCGCCAACAGTGGGCATTCACTACCCCCGGT(t/g)TATA(tt/gc)TACAGTCCCGAGTATGTCAGCATCTGCGACTCCCTTGTGAAGGTC

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
55258 GAG ATG GAG ATG CCA GAG GA Forward A
55259 GGA CCT TCA CAA GGG AGT CG Reverse A
55260 Fluorophore-1 CCG GTT TAT ATT TAC AGT CCC G Quencher-1 WT Probe
55261 Fluorophore-2 CCG GTG TAT AGC TAC AGT CCC G Quencher-2 MUT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
55258 0.40 uM
55259 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.