Stock No: 002810
Protocol 3204: Fluorescent PCR Assay - Tg(HDexon1)62Gpb STD PCR
Version 1.0

Notes

AM Bufer:  670 mM TrisHCl pH 8.8, 166 mM (NH4)2SO4, 20 mM MgCl2, 1.7 mg/ml BSA, 10 mM 2-mercaptoethanol (Add 2-mercaptoethanol to 10 mM just prior to use. Do not store with 2-mercaptoethanol in buffer.)

Size on ABI 3730.
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

Transgene = ~671-731 bp (160 +/- 10 CAG repeats)
C57BL/6J = no band
Human = 250 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
oIMR6533 GGC TGA GGA AGC TGA GGA G
TmoIMR1594 Fluorophore CCG CTC AGG TTC TGC TTT TA

Reaction A

Component Final Concentration
ddH2O
AM Buffer 1.09 X
dNTP 0.22 mM
TmoIMR1594 0.13 uM
oIMR6533 0.13 uM
DMSO
5prime HotMaster Taq 0.03 U/ul
DNA (crude prep dilu

Cycling

Step Temp °C Time Note
0 94.0 -- Put samples into preheated block.
1 94.0 --
2 94.0 --
3 58.0 --
4 70.0 -- repeat steps 2-4 for 38 cycles
5 70.0 --
6 10.0 -- hold
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
002810 B6CBA-Tg(HDexon1)62Gpb/1J
027421 B6J.Cg-Tg(HDexon1)62Gpb/160ChdiJ
2 strains use this protocol