Standard List for AFLP® Primer Nomenclature

Principle of the AFLP Method
The AFLP® technique is based on the amplification of subsets of genomic restriction fragments using PCR. DNA is cut with restriction enzymes, and double-stranded (ds) adapters are ligated to the ends of the DNA-fragments to generate template DNA for amplification. The sequence of the adapters and the adjacent restriction site serve as primer binding sites for subsequent amplification of the restriction fragments. Selective nucleotides are included at the 3' ends of the PCR primers, which therefore can only prime DNA synthesis from a subset of the restriction sites. Only restriction fragments in which the nucleotides flanking the restriction site match the selective nucleotides will be amplified. (Vos, et al., 1995)

Core sequences for the primer sets (with selective nucleotides shown as N)
EcoRI 5'-GACTGCGTACCAATTCNNN-3'
 MseI 5'-GATGAGTCCTGAGTAANNN-3'
 PstI 5'-GACTGCGTACATGCAGNNN-3'

Click here to see a flowchart of the AFLP protocol from KeyGene.

References

This list was generously provided by KeyGene. AFLP® is a registered trademark of Keygene N.V.

EcoRI

Primers +1        ¦A-3   E01
                  ¦C-3   E02
                  ¦G-3   E03
                  ¦T-3   E04


Primers +2        ¦AA-3   E11
                  ¦AC-3   E12
                  ¦AG-3   E13
                  ¦AT-3   E14
                  ¦CA-3   E15
                  ¦CC-3   E16
                  ¦CG-3   E17
                  ¦CT-3   E18
                  ¦GA-3   E19
                  ¦GC-3   E20
                  ¦GG-3   E21
                  ¦GT-3   E22
                  ¦TA-3   E23
                  ¦TC-3   E24
                  ¦TG-3   E25
                  ¦TT-3   E26

Primers +3        ¦AAA-3  E31
                  ¦AAC-3  E32
                  ¦AAG-3  E33
                  ¦AAT-3  E34
                  ¦ACA-3  E35
                  ¦ACC-3  E36
                  ¦ACG-3  E37
                  ¦ACT-3  E38
                  ¦AGA-3  E39
                  ¦AGC-3  E40
                  ¦AGG-3  E41
                  ¦AGT-3  E42
                  ¦ATA-3  E43
                  ¦ATC-3  E44
                  ¦ATG-3  E45
                  ¦ATT-3  E46
                  ¦CAA-3  E47
                  ¦CAC-3  E48
                  ¦CAG-3  E49
                  ¦CAT-3  E50
                  ¦CCA-3  E51
                  ¦CCC-3  E52
                  ¦CCG-3  E53
                  ¦CCT-3  E54
                  ¦CGA-3  E55
                  ¦CGC-3  E56
                  ¦CGG-3  E57
                  ¦CGT-3  E58
                  ¦CTA-3  E59
                  ¦CTC-3  E60
                  ¦CTG-3  E61
                  ¦CTT-3  E62
                  ¦GAA-3  E63
                  ¦GAC-3  E64
                  ¦GAG-3  E65
                  ¦GAT-3  E66
                  ¦GCA-3  E67
                  ¦GCC-3  E68
                  ¦GCG-3  E69
                  ¦GCT-3  E70
                  ¦GGA-3  E71
                  ¦GGC-3  E72
                  ¦GGG-3  E73
                  ¦GGT-3  E74
                  ¦GTA-3  E75
                  ¦GTC-3  E76
                  ¦GTG-3  E77
                  ¦GTT-3  E78
                  ¦TAA-3  E79
                  ¦TAC-3  E80
                  ¦TAG-3  E81
                  ¦TAT-3  E82
                  ¦TCA-3  E83
                  ¦TCC-3  E84
                  ¦TCG-3  E85
                  ¦TCT-3  E86
                  ¦TGA-3  E87
                  ¦TGC-3  E88
                  ¦TGG-3  E89
                  ¦TGT-3  E90
                  ¦TTA-3  E91
                  ¦TTC-3  E92
                  ¦TTG-3  E93
                  ¦TTT-3  E94

MseI


Primers +1  ¦A-3    M01
            ¦C-3    M02
            ¦G-3    M03
            ¦T-3    M04

MseI+2:     ¦AA-3   M11
            ¦AC-3   M12
            ¦AG-3   M13
            ¦AT-3   M14
            ¦CA-3   M15
            ¦CC-3   M16
            ¦CG-3   M17
            ¦CT-3   M18
            ¦GA-3   M19
            ¦GC-3   M20
            ¦GG-3   M21
            ¦GT-3   M22
            ¦TA-3   M23
            ¦TC-3   M24
            ¦TG-3   M25
            ¦TT-3   M26

MseI+3:     ¦AAA-3  M31
            ¦AAC-3  M32
            ¦AAG-3  M33
            ¦AAT-3  M34
            ¦ACA-3  M35
            ¦ACC-3  M36
            ¦ACG-3  M37
            ¦ACT-3  M38
            ¦AGA-3  M39
            ¦AGC-3  M40
            ¦AGG-3  M41
            ¦AGT-3  M42
            ¦ATA-3  M43
            ¦ATC-3  M44
            ¦ATG-3  M45
            ¦ATT-3  M46
            ¦CAA-3  M47
            ¦CAC-3  M48
            ¦CAG-3  M49
            ¦CAT-3  M50
            ¦CCA-3  M51
            ¦CCC-3  M52
            ¦CCG-3  M53
            ¦CCT-3  M54
            ¦CGA-3  M55
            ¦CGC-3  M56
            ¦CGG-3  M57
            ¦CGT-3  M58
            ¦CTA-3  M59
            ¦CTC-3  M60
            ¦CTG-3  M61
            ¦CTT-3  M62
            ¦GAA-3  M63
            ¦GAC-3  M64
            ¦GAG-3  M65
            ¦GAT-3  M66
            ¦GCA-3  M67
            ¦GCC-3  M68
            ¦GCG-3  M69
            ¦GCT-3  M70
            ¦GGA-3  M71
            ¦GGC-3  M72
            ¦GGG-3  M73
            ¦GGT-3  M74
            ¦GTA-3  M75
            ¦GTC-3  M76
            ¦GTG-3  M77
            ¦GTT-3  M78
            ¦TAA-3  M79
            ¦TAC-3  M80
            ¦TAG-3  M81
            ¦TAT-3  M82
            ¦TCA-3  M83
            ¦TCC-3  M84
            ¦TCG-3  M85
            ¦TCT-3  M86
            ¦TGA-3  M87
            ¦TGC-3  M88
            ¦TGG-3  M89
            ¦TGT-3  M90
            ¦TTA-3  M91
            ¦TTC-3  M92
            ¦TTG-3  M93
            ¦TTT-3  M94

PstI/SseI

PstI +1     ¦A    :P01
            ¦C    :P02
            ¦G    :P03
            ¦T    :P04

PstI +2     ¦AA   :P11
            ¦AC   :P12
            ¦AG   :P13
            ¦AT   :P14
            ¦CA   :P15
            ¦CC   :P16
            ¦CG   :P17 
            ¦CT   :P18
            ¦GA   :P19¦ S01
            ¦GC   :P20¦ S02  Sse+1
            ¦GG   :P21¦ S03
            ¦GT   :P22¦ S04
            ¦TA   :P23
            ¦TC   :P24
            ¦TG   :P25
            ¦TT   :P26

PstI+3:     ¦AAA  :P31
            ¦AAC  :P32
            ¦AAG  :P33
            ¦AAT  :P34
            ¦ACA  :P35
            ¦ACC  :P36
            ¦ACG  :P37
            ¦ACT  :P38
            ¦AGA  :P39
            ¦AGC  :P40
            ¦AGG  :P41
            ¦AGT  :P42
            ¦ATA  :P43
            ¦ATC  :P44
            ¦ATG  :P45
            ¦ATT  :P46
            ¦CAA  :P47
            ¦CAC  :P48
            ¦CAG  :P49
            ¦CAT  :P50
            ¦CCA  :P51
            ¦CCC  :P52
            ¦CCG  :P53
            ¦CCT  :P54
            ¦CGA  :P55
            ¦CGC  :P56
            ¦CGG  :P57
            ¦CGT  :P58
            ¦CTA  :P59
            ¦CTC  :P60
            ¦CTG  :P61
            ¦CTT  :P62
            ¦GAA  :P63¦ S11
            ¦GAC  :P64¦ S12
            ¦GAG  :P65¦ S13
            ¦GAT  :P66¦ S14
            ¦GCA  :P67¦ S15
            ¦GCC  :P68¦ S16
            ¦GCG  :P69¦ S17
            ¦GCT  :P70¦ S18  Sse+2
            ¦GGA  :P71¦ S19
            ¦GGC  :P72¦ S20
            ¦GGG  :P73¦ S21
            ¦GGT  :P74¦ S22
            ¦GTA  :P75¦ S23
            ¦GTC  :P76¦ S24
            ¦GTG  :P77¦ S25
            ¦GTT  :P78¦ S26
            ¦TAA  :P79
            ¦TAC  :P80
            ¦TAG  :P81
            ¦TAT  :P82
            ¦TCA  :P83
            ¦TCC  :P84
            ¦TCG  :P85
            ¦TCT  :P86
            ¦TGA  :P87
            ¦TGC  :P88
            ¦TGG  :P89
            ¦TGT  :P90
            ¦TTA  :P91
            ¦TTC  :P92
            ¦TTG  :P93
            ¦TTT  :P94


PstI+4/     ¦GACC     ¦ S36
SseI+3      ¦GACG     ¦ S37
            ¦GACT     ¦ S38
            ¦GAGC     ¦ S40
            ¦GAGG     ¦ S41
            ¦GCAA     ¦ S47
            ¦GCAT     ¦ S50
            ¦GGTA     ¦ S75
            ¦GGTC     ¦ S76
            ¦GGTG     ¦ S77
            ¦GGTT     ¦ S78
            ¦GTGA     ¦ S87
            ¦GTGC     ¦ S88  SseI+3
            ¦GTGG     ¦ S89
            ¦GTGT     ¦ S90
            ¦GAAC     ¦ S32
            ¦GAAG     ¦ S33
            ¦GACA     ¦ S35
            ¦GAGA     ¦ S39
            ¦GCAC     ¦ S48
            ¦GCAG     ¦ S49
            ¦GGAC     ¦ S64
            ¦GGAG     ¦ S65
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