Table 7 Position of genes analyzed in the reference genome EquCab3.0, primer sequences and PCR annealing temperatures
Gene | Position of the gene analyzed (EquCab3.0) | Strand | Amplification details | Forward primer | Reverse primer | Annealing T (°C) | PCR product lenght (bp) |
|---|---|---|---|---|---|---|---|
TNFA | 32,223,398.. 32,226,182 | + | TNFA_5UTR | CCTTTCAGAAGACCCATCCA | CATCTCGGATCATGCTTTCA | 59.9 | 777 |
| Â | Â | Â | TNFA_1CR | TAAACAGCCAGGCGATTTTCTCCCT | CCTACAACATGGGCTACAGGCTTG | 57.5 | 1144 |
| Â | Â | Â | TNFA_2CR | TGCCTTCCAGTCAATCAACCCTCT | GGTCACACATCCCTGCATTCTAGGTT | 61.5 | 1192 |
| Â | Â | Â | TNFA_3UTR | TGAGCCCATCTACCTGGGAGGAGT | GCAGAGGTTCAGCGATGTAGCGA | 59 | 868 |
DRB1 | 33,625,487.. 33,631,729 | – | 1st round | GGGACGTGTTTAAGATGGGT | AACCACACACCCTCTCCACTG | 7x(62–0,3/cycle) followed by 60 | 812 |
| Â | Â | Â | 2nd round | TGACCGGATCCTTCCTGTAC | GCGCTCACCTCGCCGAC | 60 | 303 |
DRB2 | 34,096,675.. 34,108,525 | + | 1st round | TGTCCTTCAGGTGGAGGCAA | TCACACACTGACAACCACACATT | 65 | 793 |
|  |  |  | 2nd round | TGACCCGATCSTTCCTGTAT | RCGCTCACCTCGCCGAG | 13x(65–0,3/cycle) followed by 61 | 303 |
DRB3 | 34,266,651.. 34,285,281 | + | 1st round | ACTCGCTCACAGTCCTACACAC | GTGCTGGTAGTTCGTGCGTGG | 65 | 532 |
|  |  |  | 2nd round | TGACCGGATCCTTCCTGTAC | GCGCTCACCTCGCCGAT | 13x(65–0,3/cycle) followed by 61 | 303 |
DQB1 | 33,812,679.. 33,820,407 | – |  | CCTCTGGGGTAACGTTCCAG | CGGCCTTGCTTTAGGTTTATC | 4x(63–0,5/cycle) followed by 61 | 590 |
DQB2 | 33,941,932.. 33,956,144 | – |  | AGGTTTCTCCCACTCAACTGCCTGA | GGACGCGCCCACCTCCCTGTCC | 66 | 522 |
DQB3 | 34,031,398.. 34,037,071 | – |  | AGGTTTATCCGATCCAACCGGCTGC | GCCCTCCCAGCTCCGAGACT | 4X(68–0,5/cycle) followed by 66 | 451 |
DQB2-like | unknown | unknown | Â | GCTCTCCTGGCGCAGAGACT | ACAGGGCTCTCATTTCCTTGTA | 65.5 | 603 |
| Â | Â | Â | Â | GGTCAGAGCGGGAGGCGAGT | GCCCCATAAGCTTCGCAGCA | 64 | 902 |