Genotyping Protocols Database
Stock No: 004861
Protocol 17752: Standard PCR Assay - Ts(16C-tel)1Cje Alternate1
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

Mutant = ~210 bp
Heterozygote = ~210 bp and 173 bp
Wild type = 173 bp

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
10774 CTC GTG CTT TAC GGT ATC GC Mutant Forward A Neo F
25948 CGG GCA GCT AAC TTT TCA GT Wild type Forward A Sod1
25949 GCC TAC ACA TAA GAG GCA ATT CT Common A Sod1

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
10774 0.50 uM
25948 0.50 uM
25949 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

Stock Number Strain Name
004838 B6.Cg-T(12;16)1Cje/CjeDnJ
004861 B6EiC3Sn-Ts(16C-tel)1Cje/DnJ
2 strains use this protocol