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Table 1 Characteristics of Melt-MAMA primers designed for six major branches containing Italian Brucella abortus isolates. Genome position is in the reference strain 2308, all in chromosome I

From: Origins and global context of Brucella abortus in Italy

Branch

Location in B. abortus 2308

SNP allele

Melt-MAMA primer

Melt-MAMA Primer sequences (5′ to 3′)

PCR mix Concentration

Annealing Temp.

Melting Temp.

Branch 1

West Italia

951620

NC_007618.1

C/T

A

GCGACCGCATCCTCTAaTC

0.4 μM

 

80.8 ° C

D

cggggcggggcggggGCGACCGCATCCTCTAcTT

0.1 μM

56 ° C

83.4 ° C

C

CATGGTAGGCAGGCTGGTT

0.5 μM

  

Branch 2

1336380

A/G

A

CGAAGTTTCCCGACACCcA

0.4 μM

 

81.2 ° C

West Italia

NC_007618.1

 

D

cggggcggggcggggCGAAGTTTCCCGACACCtG

0.1 μM

56 ° C

83.9 ° C

   

C

GATGAAGAAATCGATGAGCG

0.5 μM

  

Branch 1

104052

G/T

A

TGGAGGACGAAGTCTTCGaTG

0.25 μM

 

78 ° C

East Italia

NC_007618.1

 

D

cggggcggggcggggGGAGGACGAAGTCTTCGcTT

0.25 μM

56 ° C

82.9 ° C

   

C

ATCCTCGACAGTTGCGGC

0.5 μM

  

Branch 2

360724

C/A

A

TACCGCGGCCCGTCTAtC

0.25 μM

 

81.6 ° C

East Italia

NC_007618.1

 

D

cggggcggggcggggTACCGCGGCCCGTCTAaA

0.25 μM

56 ° C

83.7 ° C

   

C

AATCAAAGCGGCTTGCCTC

0.5 μM

  

Branch 1

740090

T/C

A

CAAAAGCGGACGGTCAgAT

0.4 μM

 

78.9 ° C

Trans Italia

NC_007618.1

 

D

cggggcggggcggggTCAAAAGCGGACGGTCAtAC

0.1 μM

56 ° C

80.9 ° C

   

C

CCCTCAACCCAGACGTTCG

0.5 μM

  

Branch 2

864308

C/T

A

AGGCATTACCGTTTCAGATTtC

0.4 μM

 

78.5 ° C

Trans Italia

NC_007618.1

 

D

cggggcggggcggggAGGCATTACCGTTTCAGATTaT

0.1 μM

56 ° C

81.9 ° C

   

C

TCTTTCCCGGCTCCGAAT

0.5 μM

  
  1. Melt-MAMA primer abbreviations: A ancestral, D derived, C common
  2. Details of primers concentration used in each reaction with proper annealing temperature is given. Melting temperatures detected for both the amplicons derived by the specific primer binding is given according to the experiments run on an Applied Biosystems 7500 Real-Time PCR System