LOCUS       sequence-415397-        5627 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BLaTM-System; Antiparallel negativecontrol EmrE_TMD4_3P_G90V_G97V in
            NBLa 1.2 vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5627)
  AUTHORS   Julius A, Laur L, Schanzenbach C, Langosch D
  TITLE     BLaTM 2.0, a genetic tool revealing preferred antiparallel 
            interaction of transmembrane helix 4 of the dual-topology protein 
            EmrE.
  JOURNAL   J Mol Biol. 2017 Apr 18. pii: S0022-2836(17)30166-3. doi: 
            10.1016/j.jmb.2017.04.003.
  PUBMED    28432015
REFERENCE   2  (bases 1 to 5627)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Mol Biol.
            2017 Apr 18. pii: S0022-2836(17)30166-3. doi: 
            10.1016/j.jmb.2017.04.003."
COMMENT     Sequence Label: NBla_1.2_EmrE_TMD4_3P_G90V_G97V
FEATURES             Location/Qualifiers
     source          1..5627
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      175..720
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     621..640
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     837..854
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        1246..1348
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             1349..2008
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1415..1434)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      complement(3664..3711)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     3707..3725
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(3778..3795)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(4793..5171)
                     /label=ISS
                     /note="immunostimulatory sequence from the AmpR gene; 
                     contains unmethylated CpG dinucleotides in the context of 
                     5'-AACGTT-3' (Sato et al., 1996)"
     primer_bind     5118..5137
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     sig_peptide     complement(5287..5349)
                     /codon_start=1
                     /product="signal peptide from the E. coli outer membrane 
                     protein OmpA"
                     /label=OmpA signal peptide
                     /translation="MKKTAIAIAVALAGFATVAQA"
     promoter        complement(join(5387..5627,1..44))
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     complement(5445..5464)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 gcagtgacgg caatgtctga tgcaatatgg acaattggtt tcttatcgat gataagctgt
       61 caaacatgag aattacaact tatatcgtat ggggctgact tcaggtgcta catttgaaga
      121 gataaattgc actgaaatct agaaatattt tatctgatta ataagatgat cttcttgaga
      181 tcgttttggt ctgcgcgtaa tctcttgctc tgaaaacgaa aaaaccgcct tgcagggcgg
      241 tttttcgaag gttctctgag ctaccaactc tttgaaccga ggtaactggc ttggaggagc
      301 gcagtcacca aaacttgtcc tttcagttta gccttaaccg gcgcatgact tcaagactaa
      361 ctcctctaaa tcaattacca gtggctgctg ccagtggtgc ttttgcatgt ctttccgggt
      421 tggactcaag acgatagtta ccggataagg cgcagcggtc ggactgaacg gggggttcgt
      481 gcatacagtc cagcttggag cgaactgcct acccggaact gagtgtcagg cgtggaatga
      541 gacaaacgcg gccataacag cggaatgaca ccggtaaacc gaaaggcagg aacaggagag
      601 cgcacgaggg agccgccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
      661 caccactgat ttgagcgtca gatttcgtga tgcttgtcag gggggcggag cctatggaaa
      721 aacggctttg ctgcggccct ctcacttccc tgttaagtat cttcctggca tcttccagga
      781 aatctccgcc ccgttcgtaa gccatttccg ctcgccgcag tcgaacgacc gagcgtagcg
      841 agtcagtgag cgaggaagcg gaatatatcc tgtatcacat attctgctga cgcaccggtg
      901 cagccttttt tctcctgcca catgaagcac ttcactgaca ccctcatcag tgccaacata
      961 gtaagccagt atacactccg ccagcgctga tgtccggcgg tgcttttgcc gttacgcacc
     1021 accccgtcag tagctgaaca ggagggacag ctgatagaaa cagaagccac tggagcacct
     1081 caaaaacacc atcatacact aaatcagtaa gttggcagca tcacccgacg cactttgcgc
     1141 cgaataaata cctgtgacgg aagatcactt cgcagaataa ataaatcctg gtgtccctgt
     1201 tgataccggg aagccctggg ccaacttttg gcgaaaatga gacgttgatc ggcacgtaag
     1261 aggttccaac tttcaccata atgaaataag atcactaccg ggcgtatttt ttgagttatc
     1321 gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt
     1381 tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg
     1441 tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa
     1501 taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc
     1561 ggaattccgt atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg
     1621 ttacaccgtt ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga
     1681 cgatttccgg cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct
     1741 ggcctatttc cctaaagggt ttattgagaa tatgtttttc gtctcagcca atccctgggt
     1801 gagtttcacc agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt
     1861 caccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt
     1921 tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta
     1981 ctgcgatgag tggcagggcg gggcgtaatt tttttaaggc agttattggt gcccttaaac
     2041 gcctggttgc tacgcctgaa taagtgataa taagcggatg aatggcagaa attcgaaagc
     2101 aaattcgacc cggtcgtcgg ttcagggcag ggtcgttaaa tagccgctta tgtctattgc
     2161 tggtttaccg gtttattgac taccggaagc agtgtgaccg tgtgcttctc aaatgcctga
     2221 ggccagtttg ctcaggctct ccccgtggag gtaataattg acgatatgat catttattct
     2281 gcctcccaga gcctgataaa aacggttagc gcttcgttaa tacagatgta ggtgttccac
     2341 agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgcttgtt
     2401 tcggcgtggg tatggtggca ggccccgtgg ccgggggact gttgggcgct gccggcacct
     2461 gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc
     2521 gcgcccaccg gaaggagcta ccggcagcgg tgcggactgt tgtaactcag aataagaaat
     2581 gaggccgctc atggcgttga ctctcagtca tagtatcgtg gtatcaccgg ttggttccac
     2641 tctctgttgc gggcaacttc agcagcacgt aggggacttc cgcgtttcca gactttacga
     2701 aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca
     2761 gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg
     2821 ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg gccaggaccc
     2881 aacgctgccc gagatgcgcc gcgtgcggct gctggagatg gcggacgcga tggatatgtt
     2941 ctgccaaggg ttggtttgcg cattcacagt tctccgcaag aattgattgg ctccaattct
     3001 tggagtggtg aatccgttag cgaggtgccg ccggcttcca ttcaggtcga ggtggcccgg
     3061 ctccatgcac cgcgacgcaa cgcggggagg cagacaaggt atagggcggc gcctacaatc
     3121 catgccaacc cgttccatgt gctcgccgag gcggcataaa tcgccgtgac gatcagcggt
     3181 ccaatgatcg aagttaggct ggtaagagcc gcgagcgatc cttgaagctg tccctgatgg
     3241 tcgtcatcta cctgcctgga cagcatggcc tgcaacgcgg gcatcccgat gccgccggaa
     3301 gcgagaagaa tcataatggg gaaggccatc cagcctcgcg tcgcgaacgc cagcaagacg
     3361 tagcccagcg cgtcggccgc catgccggcg ataatggcct gcttctcgcc gaaacgtttg
     3421 gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca agattccgaa taccgcaagc
     3481 gacaggccga tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat gacccagagc
     3541 gctgccggca cctgtcctac gagttgcatg ataaagaaga cagtcataag tgcggcgacg
     3601 atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc
     3661 ggtcaaaaaa cccctcaaga cccgtttaga ggccccaagg ggttatgcta gttattgctc
     3721 agcggtggca gcagccaact cagcttcctt tcgggctttg ttagcagccg gatctcagtg
     3781 gtggtggtgg tggtgctcga gttagccgtc gtgatcttta taatcgcctt tatacagttc
     3841 atccatacca tgggtaatac ctgctgcggt aacaaattcc agcagcacca tgtgatcacg
     3901 tttctcattc ggatctttgc tcagaacgct ctgggtgctc agataatgat tatccggcag
     3961 cagaaccgga ccatcaccaa tcggtgtatt ctgctgataa tgatctgcca gctgaacgct
     4021 gccatcttca acattatggc ggatcttaaa gttggctttg atgccattct tctgtttatc
     4081 tgcggtgata tacacgttgt ggctgttgaa attatattcc agtttatggc ccaggatatt
     4141 accatcttct ttgaaatcga tgcctttcag ttcaatgcga ttcaccaggg tatcaccttc
     4201 aaatttaact tcggcacggg tcttataggt gccatcatct ttaaaggaga tggtacgttc
     4261 ttgaacataa ccttccggca ttgcagattt gaagaaatcg tgctgtttca tatgatccgg
     4321 ataacggctg aaacactgaa caccataggt cagggtggta accagggtcg gccacggaac
     4381 cggcagtttg ccggtggtac agataaattt cagggtcagt ttaccattgg ttgcatcacc
     4441 ttcgccttca ccacgaactg agaatttgtg gccattaaca tcaccatcca gttcaaccag
     4501 aatcggaaca acaccggtaa acagttctcc acctttgctt gcggctttag ctgctgcctc
     4561 tttcgcggca gcctctttcg ctgccgcttc tttggcagcg gcttctttcg ctgctgcttc
     4621 ggcaggcagc tgaacattaa ctgccagttt gtggatcccc aggttaataa tcagcaccac
     4681 cgcgcaaatc agcatcatca caataatcgc cggcaggcta gctcgattag cagaaccgct
     4741 gccgccgccg ctgccgccgc cgctgccact agtgccagtt aatagtttgc gcaacgttgt
     4801 tgccattgct acaggcgtcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     4861 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     4921 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     4981 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     5041 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     5101 cccggcgtca acacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     5161 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     5221 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     5281 tggcgccgcc tgcgccacgg tcgcaaagcc cgccagcgcc accgcaatcg caatcgcggt
     5341 ctttttcatt ttttgccctc gttatctaga gcccaaaaaa acgggtatgg agaaacagta
     5401 gagagttgcg ataaaaagcg tcaggtagca tccgctaatc ttatggataa aaatgctatg
     5461 gcatagcaaa gtgtgacgcc gtgcaaataa tcaatgtgga cttttctgcc gtgattatag
     5521 acacttttgt tacgcgtttt tgtcatggct ttggtcccgc tttgttacag aatgctttta
     5581 ataagcgggg ttaccggttt ggttagcgag aagagccagt aaaagac
//
