PCR components:
PCR amplifications were performed in 30-μL volumes containing 1X PCR buffer (ABM Inc.), 1.5 mM MgCl2 (ABM Inc.), 2.5 mM dNTP, 0.5 mM each primer, 1.0 unit of Taq polymerase (ABM Inc.), and 1–2 μL (50 ng DNA) of template DNA. The 5′ ends of forward primer were labeled with fluorophores (FAM and HEX).
PCR conditions:
| |
|
Temp. (ºC) |
Time |
| Initial denaturation |
|
95 |
5 mins |
| |
Denaturation |
95 |
30 sec |
| 35 cycle |
Annealing |
55-60 |
30 sec |
| |
Elongation |
72 |
45 sec |
| Final Elongation |
|
72 |
7 mins |
Primers:
| Locus |
Forward primer (5'-3') |
Reverse primer (5'-3') |
| BH16.1 |
GAGATGCTGCAATCGCACTT |
TCGTGCAGTTAAGCAAACCG |
| BH108 |
TGTGAATCCATTGTTAAATCGG |
ATTGATTGAATGGTGAATCCAA |
| BH26 |
AGTGGATGACCAGCACGATC |
CCGTCAGTTTATGTACCGCTA |
| BH137.1 |
TTTGTGAATCCATTGTTAAATCG |
TTGGATTGATTGAATGGTGAA |
| BH36 |
CGAAGCAATCAGTTTCCTATC |
ACCAATGACCCAATGATCAC |
| BH155.2 |
CCGAACTGCTTTCCTTGTTC |
GGAGAGAAGGCCGATGCATT |
| BH18.1 |
CGACAGCATCATCAAATCTC |
CTCCTATTCCTTCGTTGACG |
| BH63.1 |
ACCAATGACCCAATGATCAC |
CGAAGCAAGCAGTTTCCTAT |
