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Table 1 Primers and PCR protocols

From: Identification of the vascular plants of Churchill, Manitoba, using a DNA barcode library

PCR recipe for rbc L and ITS2 (total volume of the reaction: 12.5 μL)

Reagents

Final concentration

Volume per reaction (μL)

10% trehalose

5%

6.25

ddH20

 

2.00

10X buffer

1x

1.25

50 mM MgCl2

2.5 mM

0.625

10 μM primer F

0.1

0.125

10 μM primer R

0.1

0.125

10 mM dNTPs

0.05

0.0625

Polymerase (5 U/μl)

0.024 U/μL

0.06

TOTAL

 

10.50

DNA template (20-40 ng/μL)

 

2 .00

PCR recipe for mat K (total volume of the reaction: 7.5 μL)

Reagents

Final concentration

Volume per reaction(μL)

20% trehalose

5%

1.875

ddH20

 

2.60

10X buffer

1x

0.75

50 mM MgCl2

1.5 mM

0.225

10 μM primer F

0.5

0.375

10 μM primer R

0.5

0.375

10 mM dNTPs

0.2

0.15

Polymerase (5 U/μl)

0.1 U/μL

0.15

TOTAL

 

6.50

DNA template (2-4 ng/μL)

 

1.00

Primer sets

 

Sequence

Reference

rbc L primers

rbcLa-F

ATGTCACCACAAACAGAGACTAAAGC

[49] Levin et al. 2003

rbcLa-R

GTAAAATCAAGTCCACCRCG

[16] Kress & Erickson, 2009

mat K primers

MatK-1RKIM-f

ACCCAGTCCATCTGGAAATCTTGGTTC

Ki-Joong Kim, pers. comm.

MatK-3FKIM-r

CGTACAGTACTTTTGTGTTTACGAG

Ki-Joong Kim, pers. comm.

MatK_390f

CGATCTATTCATTCAATATTTC

[51] Cuenoud et al. 2002

MatK_1326r

TCTAGCACACGAAAGTCGAAGT

[51] Cuenoud et al. 2002

ITS2 primers

ITS2-S2F

ATGCGATACTTGGTGTGAAT

[17] Chen et al. 2010

ITS4

TCCTCCGCTTATTGATATGC

[50] White et al. 1990