Table 2 PCR amplification conditions for COI gene in each research institute.

From: A taxonomically reliable DNA barcode reference library for North Sea macrobenthos

PCR recipe (μL)

DZMB

Naturalis

ILVO

template DNA

2

 

2

 

2

 

Master Mix

12.5*

 

6**

 

20***

 

Forward primer, 10 pmol/μL

0.5

 

1.3

 

4

 

Reverse primer, 10 pmol/μL

0.5

 

1.3

 

4

 

ddH2O

to 25

 

to 25

 

to 40

 

PCR program (steps)

T, °C

Time

T, °C

Time

T, °C

Time

Initial denaturation

94

3′

98

30″

94

2′

35–40 cycles

Denaturation

94

30″

98

5″

94

1′

Annealing

47

60″

50

5″

50

1′

Elongation

72

1′

72

15″

72

1′

Final elongation

72

5′

72

5′

72

7′

  1. *AccuStart PCR SuperMix (ThermoFisher Scientific).
  2. **0.5 μL Phire II Hotstart polymerase, 5.0 μL Phire buffer, 0.5 µL dNTP (2.5 mM).
  3. ***20 µl Red Taq DNA-polymerase: 2x MasterMix, 1,5 mM MgCl2 (VWR).
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