LOCUS       sequence-469017-        5158 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Quantitative bacterial two-hybrid (qB2H). Expresses IP1..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5158)
  AUTHORS   Guyot A, Maillard E, Ferreira-Pinto K, Plancon-Arnould L, 
            Nadaradjane AA, Ochsenbein F, Guerois R, Martin L, Ramos OHP
  TITLE     Optimized quantitative bacterial two-hybrid (qB2H) for 
            protein-protein interaction assessment
  JOURNAL   bioRxiv 2025.10.29.680610
REFERENCE   2  (bases 1 to 5158)
  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: "bioRxiv 
            2025.10.29.680610"
COMMENT     Sequence Label: VN515 pBR-rpoA-IP1
FEATURES             Location/Qualifiers
     source          1..5158
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(235..240,241..258,259..265)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    273..289
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     278..300
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     297..313
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     297..313
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1235..1254)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             2712..2903
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2897..2919)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3005..3145
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3060..3079
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3160..3177)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3331..3919)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3411..3430)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4090..4881,4882..4950))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4713..4732
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4951..5055)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5123..5141
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcaataaaaa atcaaatcgg atttcactat ataatctcac tttatctaag atgaatccga
       61 tggaagcatc ctgttttctc tcaatttttt tatctaaaac ccagcgttcg atgcttcttt
      121 gagcgaacga tcaaaaataa gtgccttccc atcaaaaaaa tattgacaac ataaaaaact
      181 ttgtgttata cttgtaacgc tacatggaga ttaactcaat ctagctagag aggctttaca
      241 ctttatgctt ccggctcgta taatgtgtgg aattgtgagc ggataacaat ttcacacagg
      301 aaacagctat gaccatgatt acggattcac tggaactcta gaccaaagag aggacacaat
      361 gcagggttct gtgacagagt ttctaaaacc gcgcctggtt gatatcgagc aagtgagttc
      421 gacgcacgcc aaggtgaccc ttgagccttt agagcgtggc tttggccata ctctgggtaa
      481 cgcactgcgc cgtattctgc tctcatcgat gccgggttgc gcggtgaccg aggttgagat
      541 tgatggtgta ctacatgagt acagcaccaa agaaggcgtt caggaagata tcctggaaat
      601 cctgctcaac ctgaaagggc tggcggtgag agttcagggc aaagatgaag ttattcttac
      661 cttgaataaa tctggcattg gccctgtgac tgcagccgat atcacccacg acggtgatgt
      721 cgaaatcgtc aagccgcagc acgtgatctg ccacctgacc gatgagaacg cgtctattag
      781 catgcgtatc aaagttcagc gcggtcgtgg ttatgtgccg gcttctaccc gaattcattc
      841 ggaagaagat gagcgcccaa tcggccgtct gctggtcgac gcatgctaca gccctgtgga
      901 gcgtattgcc tacaatgttg aagcagcgcg tgtagaacag cgtaccgacc tggacaagct
      961 ggtcatcgaa atggaaacca acggcacaat cgatcctgaa gaggcgattc gtcgtgcggc
     1021 aaccattctg gctgaacaac tggaagcttt cgttgactta cgtgatgtac gtcagcctga
     1081 agtgaaagaa gagaaaccag aggcggccgc aggcgcgatg ggcaccatca tgccgaaaga
     1141 tatccagctg gcgcgtcgta tccgtggctg aggatcctct acgccggacg catcgtggcc
     1201 ggcatcaccg gcgccacagg tgcggttgct ggcgcctata tcgccgacat caccgatggg
     1261 gaagatcggg ctcgccactt cgggctcatg agcgcttgtt tcggcgtggg tatggtggca
     1321 ggccccgtgg ccgggggact gttgggcgcc atctccttgc atgcaccatt ccttgcggcg
     1381 gcggtgctca acggcctcaa cctactactg ggctgcttcc taatgcagga gtcgcataag
     1441 ggagagcgtc gaccgatgcc cttgagagcc ttcaacccag tcagctcctt ccggtgggcg
     1501 cggggcatga ctatcgtcgc cgcacttatg actgtcttct ttatcatgca actcgtagga
     1561 caggtgccgg cagcgctctg ggtcattttc ggcgaggacc gctttcgctg gagcgcgacg
     1621 atgatcggcc tgtcgcttgc ggtattcgga atcttgcacg ccctcgctca agccttcgtc
     1681 actggtcccg ccaccaaacg tttcggcgag aagcaggcca ttatcgccgg catggcggcc
     1741 gacgcgctgg gctacgtctt gctggcgttc gcgacgcgag gctggatggc cttccccatt
     1801 atgattcttc tcgcttccgg cggcatcggg atgcccgcgt tgcaggccat gctgtccagg
     1861 caggtagatg acgaccatca gggacagctt caaggatcgc tcgcggctct taccagccta
     1921 acttcgatca ttggaccgct gatcgtcacg gcgatttatg ccgcctcggc gagcacatgg
     1981 aacgggttgg catggattgt aggcgccgcc ctataccttg tctgcctccc cgcgttgcgt
     2041 cgcggtgcat ggagccgggc cacctcgacc tgaatggaag ccggcggcac ctcgctaacg
     2101 gattcaccac tccaagaatt ggagccaatc aattcttgcg gagaactgtg aatgcgcaaa
     2161 ccaacccttg gcagaacata tccatcgcgt ccgccatctc cagcagccgc acgcggcgca
     2221 tctcgggcag cgttgggtcc tggccacggg tgcgcatgat cgtgctcctg tcgttgagga
     2281 cccggctagg ctggcggggt tgccttactg gttagcagaa tgaatcaccg atacgcgagc
     2341 gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca tgaatggtct
     2401 tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc agcgccctgc accattatgt
     2461 tccggatctg catcgcagga tgctgctggc taccctgtgg aacacctaca tctgtattaa
     2521 cgaagcgctg gcattgaccc tgagtgattt ttctctggtc ccgccgcatc cataccgcca
     2581 gttgtttacc ctcacaacgt tccagtaacc gggcatgttc atcatcagta acccgtatcg
     2641 tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat cccccttaca
     2701 cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc tttatcagaa
     2761 gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa caggcagaca
     2821 tctgtgaatc gcttcacgac cacgctgatg agctttaccg cagctgcctc gcgcgtttcg
     2881 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     2941 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     3001 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     3061 ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg
     3121 cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg
     3181 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     3241 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     3301 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3361 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3421 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3481 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     3541 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     3601 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     3661 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     3721 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     3781 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     3841 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     3901 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     3961 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     4021 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     4081 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     4141 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     4201 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     4261 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     4321 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     4381 tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat
     4441 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     4501 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     4561 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     4621 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     4681 accgagttgc tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt
     4741 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     4801 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     4861 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     4921 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     4981 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     5041 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat
     5101 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttggat aaccagaa
//
