Table 1 PCR conditions for quantitative real-time PCR analysis of hamster and mouse models of TSE disease.
Hamster | ||||||||
|---|---|---|---|---|---|---|---|---|
Gene | Primer Pairs | Size (bp) | Denature 95°C (sec) | Anneal (°C) | Anneal Time (sec) | Extension 72°C (sec) | Mg2+ (mM) | Number Cycles |
Mx2 | 5'-TGGCGGTAGGCATTCAGG-3' 5'-TGCCAGGACCAAGTTTACAGG-3' | 161 | 5 | 54 | 5 | 7 | 4 | 40 |
2'5'-OAS | 5'-AGCCGCTGCCCCCACTGTA-3' 5'-TCCGAGACTGCCCTGAAGC-3' | 109 | 5 | 57 | 5 | 5 | 3 | 50 |
Eif2ak2 | 5'-TAGGCCTTGTCAACAGTTATGCTCA-3' 5'-GCTGCTTTGCCTCCTGCTTGGTAG-3' | 162 | 5 | 58 | 5 | 7 | 3 | 45 |
GAPdH | 5'-ATGGCAAGTTCAAAGGCACAGTCA-3' 5'-TGGGGGCATCAGCAGAAGG-3' | 231 | 5 | 60 | 5 | 11 | 3 | 40 |
GFAP | 5'-GGACATCGAGATTGCCACCTAT-3' 5'-CATCCCTCATCTCCACTGTCTTTA-3' | 176 | 5 | 60 | 5 | 8 | 3 | 45 |
Mouse | ||||||||
Gene | Primer Pairs | Size (bp) | Denature 95°C (sec) | Anneal (°C) | Anneal Time (sec) | Extension 72°C (sec) | Mg2+ (mM) | Number Cycles |
Mx2 | 5'-CCTGCCTGCCATCGCTGTC-3' 5'-GCCTCTCCACTCCTCTCCCTCATT-3' | 160 | 5 | 60 | 5 | 11 | 3 | 40 |
2'5'-OAS | 5'-GAGGCGGTTGGCTGAAGAGG-3' 5'-GAGGAAGGCTGGCTGTGATTGG-3' | 312 | 5 | 60 | 5 | 11 | 3 | 40 |
Eif2ak2 | 5'-GTACAAGCGCTGGCAGAACTCAAT-3' 5'-AAGAGGCACCGGGTTTTGTAT-3' | 124 | 5 | 60 | 5 | 11 | 4 | 40 |
GAPdH | 5'-CACGGCAAATTCAACGGCACAGT-3' 5'-TGGGGGCATCGGCAGAAGG-3' | 232 | 5 | 60 | 5 | 11 | 3 | 40 |
GFAP | 5'-GAGCGAGCGTGCAGAGATGATGG-3' 5'-CTCCCGAAGCTCCGCCTGGTAGA-3' | 166 | 5 | 60 | 5 | 11 | 3 | 45 |
Caspase 8 | 5'-TCTGCTGGGAATGGCTACGGTGAA-3' 5'-GTGTGAAGGTGGGCTGTGGCATCT-3' | 212 | 5 | 60 | 5 | 11 | 3 | 45 |
TNFα | 5'-TGCTCTGTGAAGGGAATGGGTGTT-3' 5'-AGTCCTTGATGGTGGTGCATGAGA-3' | 241 | 5 | 60 | 5 | 11 | 3 | 45 |
TNFα Receptor | 5'-AACCAGTTCCAACGCTACCTGAGT-3' 5'-AAGGGACGCACTCACTTTCTCTCA-3' | 157 | 5 | 60 | 5 | 8 | 3 | 45 |
Fas Ligand | 5'-ATCCCTCTGGAATGGGAAGACACA-3' 5'-ACCCAGTTTCGTTGATCACAAGGC-3' | 91 | 5 | 63 | 5 | 6 | 3 | 45 |
Fas/ApoI Receptor | 5'-TCGCCTATGGTTGTTGACCATCCT-3' 5'-TGGTATGGTTTCACGACTGGAGGT-3' | 136 | 5 | 57 | 5 | 7 | 3 | 45 |