Table 1 of Mabuchi, Mol Vis 2009; 15:1045-1049.
Allele specific primer PCR method |
Annealing temperature |
Product size |
---|---|---|
1st PCR | ||
Forward Primer: 5’-TGGTCCTCTGACTGCTCTTT-3’ | 55 °C | 531 bp |
Reverse Primer: 5’-AGGGTGTGATGGGATGGATAA-3’ | ||
2nd PCR for G allele genotyping | ||
Forward Primer: 5’-ATGCCAGAGGCTGCTCGCCG-3’ | 65 °C | 398 bp |
Reverse Primer: 5’-AGGGTGTGATGGGATGGATAA-3’ | ||
2nd PCR for C allele genotyping | ||
Forward Primer: 5’-TGGTCCTCTGACTGCTCTTT-3’ | 62 °C | 172 bp |
Reverse Primer: 5’-TGCTGGTGCAGGGGCCTCGG-3’ | ||
Pyrosequencing analysis | ||
PCR forward primer: 5’-AATTCCATGGGACTGACTTTCTGC-3’ | 60 °C | 370 bp |
PCR reverse primer: biotin labeled 5’-Bio-GGGAAGGGACAGAAGATGACAG-3’ | ||
Sequencing reverse primer: 5’-GGACAAGGGTTGGGC-3’ |