Stock No: 033668
Protocol 36808: Standard PCR Assay - Tc(HSA17)1Mdk MAPT
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

>chr11:104265226+104265523 298bp TTCTTAACCGCTGAGCCATC CCTTTGGGATTTTAACCATGTG

 

Mutant = 151 bp
Heterozygote = 151 bp and 298 bp
Wild type = 298 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
49773 TTC TTA ACC GCT GAG CCA TC Wild type Forward A Mapt F
49774 CCT TTG GGA TTT TAA CCA TGT G Wild type Reverse A Mapt R
49775 TCA CTT ACA CGA GGC ATC CA Mutant Forward A MAPT F
49776 AAC TGA CCA CGG TTC TCC AC Mutant Reverse A MAPT R

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
49773 0.50 uM
49774 0.50 uM
49775 0.50 uM
49776 0.50 uM
Glycerol 6.50 %
Dye 1.00 X
Kapa 2G HS taq polym 0.03 U/ul
DNA

Cycling

Step Temp °C Time Note
1 94.0 --
2 94.0 --
3 65.0 -- -0.5 C per cycle decrease
4 68.0 --
5 -- repeat steps 2-4 for 10 cycles (Touchdown)
6 94.0 --
7 60.0 --
8 72.0 --
9 -- repeat steps 6-8 for 28 cycles
10 72.0 --
11 10.0 -- hold
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.
JAX uses a 'touchdown' cycling protocol and therefore has not calculated the optimal annealing temperature for each set of primers.

Strains Using This Protocol

This is the only strain that uses this protocol.