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. 2012 Dec 13;2(1):35–53. doi: 10.1002/mbo3.50

Table 1.

Primers used for genome walking, primer walking, and mRNA expression studies

Primer name Sequence1 Organism/source Gene2 Application
I ESP GSP1 up AAATCCATCTTCTCTAGGTACACCGTT Clostridium ultunense strain Esp fhs1 Upstream walk
II ESP GSP1 down CTGAAGAACTAGGTTCAGTAGCAGTAC C. ultunense strain Esp fhs1 Downstream walk
III ESP GSP2 down GAGACGGTGGAATCGAATTAGCTAAGA C. ultunense strain Esp fhs1 Downstream walk
I 2.fhs ESP GSP1 up CTCCTCCATGGGAACCACCTGTGAGTA C. ultunense strain Esp fhs2 Upstream walk
II 2.fhs ESP GSP1-3 up GGTGGATCTTCGCTTTATATTTTCCGT C. ultunense strain Esp fhs2 Upstream walk
III 2.fhs ESP GSP1 down CGGAAGGGGCCATGGCCCTTTTGTTAA C. ultunense strain Esp fhs2 Downstream walk
I Sp3 GSP1 up TGTGTGCGTTCATGATGTCATTGATGT Syntrophaceticus schinkii strain Sp3 fhs Upstream walk
II Sp3 GSP2 up AGCGGCAGACCCTTTAATGTTCATAGT S. schinkii strain Sp3 fhs Upstream walk
IV Sp3 GSP1 down TTGATGGCCATTTTGGCTGTGGCAAAA S. schinkii strain Sp3 fhs Downstream walk
V Sp3 GSP2 down AACAAGAAGGGTGAGCCGGTTACAACA S. schinkii strain Sp3 fhs Downstream walk
III Sp3 GSP1-1 up ATAACACCAGAGGAAGGCAAGACCTAT S. schinkii strain Sp3 fhs Upstream walk
VI Sp3 GSP1-1 down TGCACTGGAACTGGCAGATGCTGTTAT S. schinkii strain Sp3 fhs Downstream walk
VII Sp3 A5 downstream primer TTCTGCAAGCATAACCTGCAGCACTTT S. schinkii strain Sp3 fhs Primer walking
I first primer upstream GSP1 TTTTCCATCATATGTATAACCAATTAT Tepidanaerobacter acetatoxydans strain ReI fhs1 Upstream walk
II nested primer upstr GSP2 GCAAGCCTATTTTTCAAATCTTCCAT T. acetatoxydans strain ReI fhs1 Upstream walk
III first primer upstr GSP1-2 TTACCGAGCCTTCTGAGTGCATCACCT T. acetatoxydans strain ReI fhs1 Upstream walk
IV nested primer up GSP2-2 TTTGCCTTCACCTGCAGGAGTTGGATT T. acetatoxydans strain ReI fhs1 Upstream walk
V DL2 upstream primer ATAGCAGGTGCTGCATCAATGCCCTC T. acetatoxydans strain ReI fhs1 Upstream walk
VI GSP1-3up GTCTGCATCACAGCCGATATGTGTTGA T. acetatoxydans strain ReI fhs1 Upstream walk
VII nested GSP2-3up TGGCCCGAGGCACCACATTCATCTTTT T. acetatoxydans strain ReI fhs1 Upstream walk
VIII DL1 2.3-kb up primer TTGTGCAAAGCATGCACTGCTCTAATT T. acetatoxydans strain ReI fhs1 Primer walking
VIV GSP1-4up CAAAGATAACCGATTCGGCAATCCCAA T. acetatoxydans strain ReI fhs1 Upstream walk
I 2.fhs ReI GSP1 down CATATGATCGGGAGGGTAAGCCCGTTA T. acetatoxydans strain ReI fhs2 Downstream walk
II 2.fhs ReI nested GSP2 down CAAGGTGCCATGGCGGCTCTACTCAAA T. acetatoxydans strain ReI fhs2 Downstream walk
III 2.fhs downstr pr (A1/A2) GCCCTGAAATAACACCCTTTTCATCCA T. acetatoxydans strain ReI fhs2 Primer walking
IV 2.fhs GSP1-2 down CGCGGAGCCCTTAAAGCAATGGCAAAT T. acetatoxydans strain ReI fhs2 Downstream walk
V DL1 (A1) 2.fhs downstr pr5' CCCTTTGAAGAAATAACTGAGTATTTA T. acetatoxydans strain ReI fhs2 Primer walking
VI DL1 (A1) 2.fhs downstr pr3' GTGCCAATTCGGCAGTAACTGCAAACT T. acetatoxydans strain ReI fhs2 Primer walking
VII 2.fhs ReI GSP1 up GGGCTTACCCTCCCGATCATATGCTAC T. acetatoxydans strain ReI fhs2 Upstream walk
VIII 2.fhs ReI GSP2 up CTTCGCCAGCCGTTCTTTCAAATCCAT T. acetatoxydans strain ReI fhs2 Upstream walk
VIV 2.fhs ReI GSP2-2up CGACCTTTGCTTTATATTTGCCGTACA T. acetatoxydans strain ReI fhs2 Upstream walk
X 2.fhs DL4 upstream pr TGCCTGACAGCAAGATCATGAATAATT T. acetatoxydans strain ReI fhs2 Primer walking
X first primer down GSP1 AAAGAGAGCTGGCACTGGTTCAAGAAG T. acetatoxydans strain ReI fhs1 Downstream walk
XI nested primer down GSP2 GCAGTGCTTTCCGAGGTTTGGGCAAAA T. acetatoxydans strain ReI fhs1 Downstream walk
XII DL3 downstream primer AGCACTTACCGGTGCTATCATGACAAT T. acetatoxydans strain ReI fhs1 Primer walking
XIII DL3 downstream primer2 ATGTTCATCGCCCTGTGGTGCAACATA T. acetatoxydans strain ReI fhs1 Primer walking
1.fhs locus rev CCATTACTATGCACAAACCTAAAGCCT T. acetatoxydans strain ReI fhs1 Locus verification
2.fhs locus rev TCAATGGGATCATATTCCGGAGTTACA T. acetatoxydans strain ReI fhs2 Locus verification
1.locus Oxido fw TTATTTGCGATTATCCGCAGGAAAGAC T. acetatoxydans strain ReI fhs1 Locus verification
2.locus Methyltr fw ATGATTATTATTGGGGAAAAGATTAAC T. acetatoxydans strain ReI fhs2 Locus verification
Esp 1.fhs RNA fw TAGACCAAACGGTGTACCTA C. ultunense strain Esp fhs1 mRNA expression
Esp 1.fhs RNA rev AAGTAAAGCTACTGCTCCCT C. ultunense strain Esp fhs1 mRNA expression
Esp 2.fhs RNA fw AAAGAAAAACGGAGTGCCTC C. ultunense strain Esp fhs2 mRNA expression
Esp 2.fhs RNA rev CTTTAACAAAAGGGCCATGG C. ultunense strain Esp fhs2 mRNA expression
ReI 1.fhs RNA fw GGCTGCAACAGTATAATAGC T. acetatoxydans strain ReI fhs1 mRNA expression
ReI 1.fhs RNA rev TCTTGACCCCGCCATTATAT T. acetatoxydans strain ReI fhs1 mRNA expression
ReI 2.fhs RNA fw TGTTGTAGCATATGATCGGG T. acetatoxydans strain ReI fhs2 mRNA expression
ReI 2.fhs RNA rev TTACCGAGTTACATCCGTGT T. acetatoxydans strain ReI fhs2 mRNA expression
Sp3 fhs RNA fw CTCGAAAGACGGTTTCTTGA S. schinkii strain Sp3 fhs mRNA expression
Sp3 fhs RNA rev TTGATCGTGTTACGCATCCA S. schinkii strain Sp3 fhs mRNA expression
T. phaeum this study fw CGGACCCACCATGAACATTAAGGGTA Thermacetogenium phaeum strain PB fhs Authenticity check
T. phaeum this study rev AAGTCGATAACCCAGCCCATTTCCAC T. phaeum strain PB fhs Authenticity check
T. phaeum AB523739 fw ATCTTGGCTCTAACTACCGGCCTCAA T. phaeum strain PB fhs Authenticity check
T. phaeum AB523739 rev CTCGTTGCCATCATATCGGCCAGTAT T. phaeum strain PB fhs Authenticity check
M13 fw GTAAAACGACGGCCAGTG Invitrogen Sequencing
M13 rev GGAAACAGCTATGACCATG Invitrogen Sequencing
Adaptor primer 1 (AP1) GTAATACGACTCACTATAGGGC GenomeWalker Kit Sequencing/walk
Nested Adaptor primer 2 AP2) ACTATAGGGCACGCGTGGT GenomeWalker Kit Sequencing/walk

Primers were designed using Geneious v4.5 and synthesized by Invitrogen (Carlsbad, CA).

1

Sequences are indicated as 5′ to 3′.

2

Gene used to start genome walking or targeted in mRNA expression studies.