LOCUS       sequence-417781-        5269 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BLaTM-System; Antiparallel negativecontrol EmrE_TMD4_3P_G90V_G97V in
            CBLa 2.0 vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5269)
  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 5269)
  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: CBla_2.0_EmrE_TMD4_3P_G90V_G97V
FEATURES             Location/Qualifiers
     source          1..5269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(94..972)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        999..1283
                     /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     1206..1225
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             2309..2338
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2342..2359
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2425..2442)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2595..2681
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2773..2800
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2968..2987)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2979..3794
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPSIKHFIRTPDEAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3052..3071)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3876..4464
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4365..4384
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4618..4635
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4650..4790
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4716..4735)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4876..4898
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4892..5083)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 cgatgcataa tgtgcctgtc aaatggacga agcagggatt ctgcaaaccc tatgctactc
       61 cgtcaagccg tcaattgtct gattcgttac caattatgac aacttgacgg ctacatcatt
      121 cactttttct tcacaaccgg cacggaactc gctcgggctg gccccggtgc attttttaaa
      181 tacccgcgag aaatagagtt gatcgtcaaa accaacattg cgaccgacgg tggcgatagg
      241 catccgggtg gtgctcaaaa gcagcttcgc ctggctgata cgttggtcct cgcgccagct
      301 taagacgcta atccctaact gctggcggaa aagatgtgac agacgcgacg gcgacaagca
      361 aacatgctgt gcgacgctgg cgatatcaaa attgctgtct gccaggtgat cgctgatgta
      421 ctgacaagcc tcgcgtaccc gattatccat cggtggatgg agcgactcgt taatcgcttc
      481 catgcgccgc agtaacaatt gctcaagcag atttatcgcc agcagctccg aatagcgccc
      541 ttccccttgc ccggcgttaa tgatttgccc aaacaggtcg ctgaaatgcg gctggtgcgc
      601 ttcatccggg cgaaagaacc ccgtattggc aaatattgac ggccagttaa gccattcatg
      661 ccagtaggcg cgcggacgaa agtaaaccca ctggtgatac cattcgcgag cctccggatg
      721 acgaccgtag tgatgaatct ctcctggcgg gaacagcaaa atatcacccg gtcggcaaac
      781 aaattctcgt ccctgatttt tcaccacccc ctgaccgcga atggtgagat tgagaatata
      841 acctttcatt cccagcggtc ggtcgataaa aaaatcgaga taaccgttgg cctcaatcgg
      901 cgttaaaccc gccaccagat gggcattaaa cgagtatccc ggcagcaggg gatcattttg
      961 cgcttcagcc atacttttca tactcccgcc attcagagaa gaaaccaatt gtccatattg
     1021 catcagacat tgccgtcact gcgtctttta ctggctcttc tcgctaacca aaccggtaac
     1081 cccgcttatt aaaagcattc tgtaacaaag cgggaccaaa gccatgacaa aaacgcgtaa
     1141 caaaagtgtc tataatcacg gcagaaaagt ccacattgat tatttgcacg gcgtcacact
     1201 ttgctatgcc atagcatttt tatccataag attagcggat gctacctgac gctttttatc
     1261 gcaactctct actgtttctc catacccgtt tttttgggct ctagataacg agggcaaaaa
     1321 atgttcggat taggacacaa ctcaaaagag atttcgatga gtcatattgg tactaaattc
     1381 attcttgctg aaaaatttac cttcgatccc ctaagcaata ctctgattga caaagaagat
     1441 agtgaagaga tcattcgatt aggcagcaac gaaagccgaa ttctttggct gctggcccaa
     1501 cgtccaaacg aggtgatttc tcgcaatgat ttgcatgact ttgtttggcg agagcaaggt
     1561 tttgaagtcg atgattccag cttaacccaa gccatttcga ctctgcgcaa aatgctcaaa
     1621 gattcgacaa agtccccaca atacgtcaaa acggttccga agcgcggtta ccaattgatc
     1681 gcccgagtgg aaacggttga agaagagatg gctcgcgaaa acgaagctgc tcatgacatc
     1741 tctcagccag aatctgtcaa tgaatacgca gaatcaagca gtgtgccttc atcagccact
     1801 gtagtgaaca caccgcagcc agccaatgtc gtggcgaata aatcggctcc aaacttgggg
     1861 aatcgagcta gcctgccggc gattattgtg atgatgctga tttgcgcggt ggtgctgatt
     1921 attaacctgg ggatcctgat caacccaagc caatccagct ttaaacccct aacggatggt
     1981 ggtagcggcg gcagcctact tactctagct tcccggcaac aattaataga ctggatggag
     2041 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     2101 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     2161 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     2221 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattgggg cgattataaa
     2281 gatcacgacg gctaagagct cggtacccga acagaagctg atcagcgaag aagatctgaa
     2341 ccaccaccac caccaccact aactcgaggt cgacctgcag gcatgcaagc ttggctgttt
     2401 tggcggatga gagaagattt tcagcctgat acagattaaa tcagaacgca gaagcggtct
     2461 gataaaacag aatttgcctg gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa
     2521 ctcagaagtg aaacgccgta gcgccgatgg tagtgtgggg tctccccatg cgagagtagg
     2581 gaactgccag gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta
     2641 tctgttgttt gtcggtgaac gctctcctga gtaggacaaa tccgccggga gcggatttga
     2701 acgttgcgaa gcaacggccc ggagggtggc gggcaggacg cccgccataa actgccaggc
     2761 atcaaattaa gcagaaggcc atcctgacgg atggcctttt tgcgtttcta caaactcttt
     2821 tgtttatttt tctaaataca ttcaaatatg tatccgctca tggagctcaa agccacgttg
     2881 tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca tgaacaataa
     2941 aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa
     3001 cgtcttgctc caggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     3061 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgtttgtat gggaagcccg
     3121 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     3181 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgtccatc aagcatttta
     3241 tccggactcc tgatgaagca tggttactca ccactgcgat ccccggcaaa acagcattcc
     3301 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     3361 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc
     3421 gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     3481 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcacaaa cttttgccat
     3541 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     3601 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     3661 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     3721 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     3781 atgagttttt ctaagagctc catgaccaaa atcccttaac gtgagttttc gttccactga
     3841 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3901 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3961 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     4021 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     4081 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     4141 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     4201 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     4261 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     4321 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     4381 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     4441 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     4501 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     4561 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     4621 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     4681 tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag
     4741 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
     4801 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     4861 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     4921 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg
     4981 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
     5041 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag
     5101 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
     5161 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
     5221 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcg
//
