Table 3 |
|||
|
Primers used for specific PCR and direct sequencing, amplification conditions and temperature profiles. |
|||
|
Primer |
Sequence |
amplification conditions |
temperature profile |
|
|
|||
|
COI universal [43] |
5'-GGTCAACAATCATAAAGATATTGG-3' 5'-TAAACTTCAGGGTGACCAAAAAATCA-3' |
total volume 25 μl with: 0.17 mM dNTPs 3 mM MgCl2 in 1 × PCR buffer 0.13 μM of each primer 1 unit Taq polymerase (Invitrogen) |
1 cycle of 2.5 min at 94°C 40 cycle 30s at 90°C 1 min at 48°C 1 min at 72°C 1 cycle of 10 min at 72°C |
|
16S universal [44] |
5'-CGGCCGCCTGTTT ATCAAAAACAT-3' 5'-GGAGCTCCGGTTTGAACTCAGATC-3' |
total volume 15 μl with: 0.1 mM dNTPs 2.5 mM MgCl2 in 1 × PCR buffer 0.2 μM of each primer 0.5 unit Taq polymerase (Invitrogen) |
1 cycle of 2.5 min at 90°C 10 cycles of 50s at 92°C 30s at 44°C 40s at 72°C 36 cycles of 30s at 92°C 40s at 48°C 40s at 72°C 1 cycle of 3 min at 72°C |
|
ITS-1 mollusc specific [45] |
5'-TAACAAGGTTTCCGTAGGTGAA-3' 5'GCTGCGTTCTTCATCGATGC-3' |
total volume 15 μl with: 0.3 mM dNTPs 2.5 mM MgCl2 in 1 × PCR buffer 0.18 μM of each primer 0.5 unit Taq polymerase (Invitrogen) |
1 cycle of 3 min at 94°C 40 cycles of 30s at 92°C 30s at 52°C 1 min at 72°C 1 cycle of 5 min at 72°C |
|
|
|||
|
Pfenninger et al. BMC Evolutionary Biology 2005 5:59 doi:10.1186/1471-2148-5-59 |
|||
