LOCUS       sequence-469020-        5188 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Quantitative bacterial two-hybrid (qB2H). Expresses IP2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5188)
  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 5188)
  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: VN516 pBR-rpoA-IP2
FEATURES             Location/Qualifiers
     source          1..5188
                     /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(1265..1284)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             2742..2933
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2927..2949)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3035..3175
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3090..3109
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3190..3207)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3361..3949)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3441..3460)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4120..4911,4912..4980))
                     /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     4743..4762
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4981..5085)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5153..5171
                     /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 tccgtggcgc gggcggccgt accctgtacg gctttggctg
     1201 aggatcctct acgccggacg catcgtggcc ggcatcaccg gcgccacagg tgcggttgct
     1261 ggcgcctata tcgccgacat caccgatggg gaagatcggg ctcgccactt cgggctcatg
     1321 agcgcttgtt tcggcgtggg tatggtggca ggccccgtgg ccgggggact gttgggcgcc
     1381 atctccttgc atgcaccatt ccttgcggcg gcggtgctca acggcctcaa cctactactg
     1441 ggctgcttcc taatgcagga gtcgcataag ggagagcgtc gaccgatgcc cttgagagcc
     1501 ttcaacccag tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc cgcacttatg
     1561 actgtcttct ttatcatgca actcgtagga caggtgccgg cagcgctctg ggtcattttc
     1621 ggcgaggacc gctttcgctg gagcgcgacg atgatcggcc tgtcgcttgc ggtattcgga
     1681 atcttgcacg ccctcgctca agccttcgtc actggtcccg ccaccaaacg tttcggcgag
     1741 aagcaggcca ttatcgccgg catggcggcc gacgcgctgg gctacgtctt gctggcgttc
     1801 gcgacgcgag gctggatggc cttccccatt atgattcttc tcgcttccgg cggcatcggg
     1861 atgcccgcgt tgcaggccat gctgtccagg caggtagatg acgaccatca gggacagctt
     1921 caaggatcgc tcgcggctct taccagccta acttcgatca ttggaccgct gatcgtcacg
     1981 gcgatttatg ccgcctcggc gagcacatgg aacgggttgg catggattgt aggcgccgcc
     2041 ctataccttg tctgcctccc cgcgttgcgt cgcggtgcat ggagccgggc cacctcgacc
     2101 tgaatggaag ccggcggcac ctcgctaacg gattcaccac tccaagaatt ggagccaatc
     2161 aattcttgcg gagaactgtg aatgcgcaaa ccaacccttg gcagaacata tccatcgcgt
     2221 ccgccatctc cagcagccgc acgcggcgca tctcgggcag cgttgggtcc tggccacggg
     2281 tgcgcatgat cgtgctcctg tcgttgagga cccggctagg ctggcggggt tgccttactg
     2341 gttagcagaa tgaatcaccg atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt
     2401 ctgcgacctg agcaacaaca tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa
     2461 cgcggaagtc agcgccctgc accattatgt tccggatctg catcgcagga tgctgctggc
     2521 taccctgtgg aacacctaca tctgtattaa cgaagcgctg gcattgaccc tgagtgattt
     2581 ttctctggtc ccgccgcatc cataccgcca gttgtttacc ctcacaacgt tccagtaacc
     2641 gggcatgttc atcatcagta acccgtatcg tgagcatcct ctctcgtttc atcggtatca
     2701 ttacccccat gaacagaaat cccccttaca cggaggcatc agtgaccaaa caggaaaaaa
     2761 ccgcccttaa catggcccgc tttatcagaa gccagacatt aacgcttctg gagaaactca
     2821 acgagctgga cgcggatgaa caggcagaca tctgtgaatc gcttcacgac cacgctgatg
     2881 agctttaccg cagctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc
     2941 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
     3001 agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg
     3061 atagcggagt gtatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca
     3121 ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgctc
     3181 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     3241 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     3301 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     3361 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     3421 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     3481 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     3541 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     3601 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     3661 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     3721 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     3781 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     3841 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     3901 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     3961 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4021 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     4081 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     4141 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     4201 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     4261 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     4321 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     4381 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctgcagg
     4441 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     4501 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     4561 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     4621 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     4681 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg
     4741 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     4801 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     4861 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     4921 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     4981 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     5041 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     5101 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     5161 tatcacgagg ccctttggat aaccagaa
//
