LOCUS       sequence-442534-        7801 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Honey bee gut symbiont broad-host range vector with constitutive 
            E2-crimson, ampR and RSF1010 replicon.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7801)
  AUTHORS   Chhun A, Moriano-Gutierrez S, Zoppi F, Cabirol A, Engel P, Schaerli 
            Y
  TITLE     An engineered bacterial symbiont allows noninvasive biosensing of 
            the honey bee gut environment.
  JOURNAL   PLoS Biol. 2024 Mar 5;22(3):e3002523. doi: 
            10.1371/journal.pbio.3002523. eCollection 2024 Mar.
  PUBMED    38442124
REFERENCE   2  (bases 1 to 7801)
  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: "PLoS 
            Biol."; date: "2024-03-5"; volume: "22(3):e3002523. doi"; pages: " 
            10.1371/journal.pbio.3002523. eCollection 2024 Mar"
COMMENT     Sequence Label: pAC09
FEATURES             Location/Qualifiers
     source          1..7801
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(347..1318)
                     /codon_start=1
                     /gene="repB"
                     /product="replication protein B of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepB
                     /translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDATTGQMMNREWSAAEVL
                     QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
                     YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
                     PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELPERQLSRHRRTALD
                     EYRSEMAGLVKRFGHDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
     oriT            2557..2644
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      2986..3380
                     /label=RSF1010 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     RSF1010; requires the RSF1010 RepA/B/C proteins for 
                     replication (Scholz et al., 1989)"
     CDS             4004..4678
                     /codon_start=1
                     /product="far-red noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2009)"
                     /label=E2-Crimson
                     /note="mammalian codon-optimized"
                     /translation="MDSTENVIKPFMRFKVHMEGSVNGHEFEIEGVGEGKPYEGTQTAK
                     LQVTKGGPLPFAWDILSPQFFYGSKAYIKHPADIPDYLKQSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGTLIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERNYPRDGVLKGENHM
                     ALKLKGGGHYLCEFKSIYMAKKPVKLPGYHYVDYKLDITSHNEDYTVVEQYERAEARHH
                     LFQ"
     primer_bind     4402..4421
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             complement(join(4942..5733,5734..5802))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5565..5584
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5803..5907)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(5960..6811)
                     /codon_start=1
                     /gene="repC"
                     /product="replication protein C of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepC
                     /translation="MVKPKNKHSLSHVRHDPAHCLAPGLFRALKRGERKRSKLDVTYDY
                     GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKWEAV
                     TADAMVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRQRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     CDS             complement(6798..7637)
                     /codon_start=1
                     /gene="repA"
                     /product="replication protein A of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepA
                     /translation="MATHKPINILEAFAAAPPPLDYVLPNMVAGTVGALVSPGGAGKSM
                     LALQLAAQIAGGPDLLEVGELPTGPVIYLPAEDPPTAIHHRLHALGAHLSAEERQAVAD
                     GLLIQPLIGSLPNIMAPEWFDGLKRAAEGRRLMVLDTLRRFHIEEENASGPMAQVIGRM
                     EAIAADTGCSIVFLHHASKGAAMMGAGDQQQASRGSSVLVDNIRWQSYLSSMTSAEAEE
                     WGVDDDQRRFFVRFGVSKANYGAPFADRWFRRHDGGVLKPAVLERQRKSKGVPRGEA"
ORIGIN
        1 atatcgcgct tggcgtacag cgtcagggct ggccagcagg tcgccggtct gcttgtcctt
       61 ttggtctttc atatcagtca ccgagaaact tgccggggcc gaaaggcttg tcttcgcgga
      121 acaaggacaa ggtgcagccg tcaaggttaa ggctggccat atcagcgact gaaaagcggc
      181 cagcctcggc cttgtttgac gtataaccaa agccaccggg caaccaatag cccttgtcac
      241 ttttgatcag gtagaccgac cctgaagcgc ttttttcgta ttccataaaa cccccttctg
      301 tgcgtgagta ctcatagtat aacaggcgtg agtaccaacg caagcactac atgctgaaat
      361 ctggcccgcc cctgtccatg cctcgctggc ggggtgccgg tgcccgtgcc agctcggccc
      421 gcgcaagctg gacgctgggc agacccatga ccttgctgac ggtgcgctcg atgtaatccg
      481 cttcgtggcc gggcttgcgc tctgccagcg ctgggctggc ctcggccatg gccttgccga
      541 tttcctcggc actgcggccc cggctggcca gcttctgcgc ggcgataaag tcgcacttgc
      601 tgaggtcatg accgaagcgc ttgaccagcc cggccatctc gctgcggtac tcgtccagcg
      661 ccgtgcgccg gtggcggcta agctgccgct cgggcagttc gaggctggcc agcctgcggg
      721 ccttctcctg ctgccgctgg gcctgctcga tctgctggcc agcctgctgc accagcgccg
      781 ggccagcggt ggcggtcttg cccttggatt cacgcagcag cacccacggc tgataaccgg
      841 cgcgggtggt gtgcttgtcc ttgcggttgg tgaagcccgc caagcggcca tagtggcggc
      901 tgtcggcgct ggccgggtcg gcgtcgtact cgctggccag cgtccgggca atctgccccc
      961 gaagttcacc gcctgcggcg tcggccacct tgacccatgc ctgatagttc ttcgggctgg
     1021 tttccactac cagggcaggc tcccggccct cggctttcat gtcatccagg tcaaactcgc
     1081 tgaggtcgtc caccagcacc agaccatgcc gctcctgctc ggcgggcctg atatacacgt
     1141 cattgccctg ggcattcatc cgcttgagcc atggcgtgtt ctggagcact tcggcggctg
     1201 accattcccg gttcatcatc tggccggtgg tggcgtccct gacgccgata tcgaagcgct
     1261 cacagcccat ggccttgagc tgtcggccta tggcctgcaa agtcctgtcg ttcttcatcg
     1321 ggccaccaag cgcagccaga tcgagccgtc ctcggttgtc agtggcgtca ggtcgagcaa
     1381 gagcaacgat gcgatcagca gcaccaccgt aggcatcatg gaagccagca tcacggttag
     1441 ccatagcttc cagtgccacc cccgcgacgc gctccgggcg ctctgcgcgg cgctgctcac
     1501 ctcggcggct acctcccgca actctttggc cagctccacc catgccgccc ctgtctggcg
     1561 ctgggctttc agccactccg ccgcctgcgc ctcgctggcc tgctgggtct ggctcatgac
     1621 ctgccgggct tcgtcggcca gtgtcgccat gctctgggcc agcggttcga tctgctccgc
     1681 taactcgttg atgcctctgg atttcttcac tctgtcgatt gcgttcatgg tctattgcct
     1741 cccggtattc ctgtaagtcg atgatctggg cgttggcggt gtcgatgttc agggccacgt
     1801 ctgcccggtc ggtgcggatg ccccggcctt ccatctccac cacgttcggc cccaggtgaa
     1861 caccgggcag gcgctcgatg ccctgcgcct caagtgttct gtggtcaatg cgggcgtcgt
     1921 ggccagcccg ctctaatgcc cggttggcat ggtcggccca tgcctcgcgg gtctgctcaa
     1981 gccatgcctt gggcttgagc gcttcggtct tctgtgcccc gcccttctcc ggggtcttgc
     2041 cgttgtaccg cttgaaccac tgagcggcgg gccgctcgat gccgtcattg atccgctcgg
     2101 agatcatcag gtggcagtgc gggttctcgc cgccaccggc atggatggcc agcgtatacg
     2161 gcaggcgctc ggcaccggtc aggtgctggg cgaactcgga cgccagcgcc ttctgctggt
     2221 cgagggtcag ctcgaccggc agggcaaatt cgacctcctt gaacagccgc ccattggcgc
     2281 gttcatacag gtcggcagca tcccagtagt cggcgggccg ctcgacgaac tccggcatgt
     2341 gcccggattc ggcgtgcaag acttcatcca tgtcgcgggc atacttgcct tcgcgctgga
     2401 tgtagtcggc cttggccctg gccgattggc cgcccgacct gctgccggtt ttcgccgtaa
     2461 ggtgataaat cgccatgctg cctcgctgtt gcttttgctt ttcggctcca tgcaatggcc
     2521 ctcggagagc gcaccgcccg aagggtggcc gttaggccag tttctcgaag agaaaccggt
     2581 aagtgcgccc tcccctacaa agtagggtcg ggattgccgc cgctgtgcct ccatgatagc
     2641 ctacgagaca gcacattaac aatggggtgt caagatggtt aaggggagca acaaggcggc
     2701 ggatcggctg gccaagctcg aagaacaacg agcgcgaatc aatgccgaaa ttcagcgggt
     2761 gcgggcaagg gaacagcagc aagagcgcaa gaacgaaaca aggcgcaagg tgctggtggg
     2821 ggccatgatt ttggccaagg tgaacagcag cgagtggccg gaggatcggc tcatggcggc
     2881 aatggatgcg taccttgaac gcgaccacga ccgcgccttg ttcggtctgc cgccacgcca
     2941 gaaggatgag ccgggctgaa tgatcgaccg agacaggccc tgcggggctg cacacgcgcc
     3001 cccacccttc gggtaggggg aaaggccgct aaagcggcta aaagcgctcc agcgtatttc
     3061 tgcggggttt ggtgtggggt ttagcgggct ttgcccgcct ttccccctgc cgcgcagcgg
     3121 tggggcggtg tgtagcctag cgcagcgaat agaccagcta tccggcctct ggccgggcat
     3181 attgggcaag ggcagcagcg ccccacaagg gcgctgataa ccgcgcctag tggattattc
     3241 ttagataatc atggatggat ttttccaaca ccccgccagc ccccgcccct gctgggtttg
     3301 caggtttggg ggcgtgacag ttattgcagg ggttcgtgac agttattgca ggggggcgtg
     3361 acagttattg caggggttcg tgacagttag tacgggagtg acgggcactg gctggcaatg
     3421 tctagcaacg gcaggcattt cggctgaggg taaaagaact ttccgctaag cgatagactg
     3481 tatgtaaaca cagtattgca aggacgcgga acatgcctca tgtggcggcc aggacggcca
     3541 gccgggatcg ggatactggt cgttaccaga gccaccgacc cgagcaaacc cttctctatc
     3601 agatcgttga cgagtattac ccggcattcg ctgcgcttat ggcagagcag ggaaaggaat
     3661 tgccgggcta tgtgcaacgg gaatttgaag aatttctcca atgcgggcgg ctggagcatg
     3721 gctttctacg ggttcgctgc gagtcttgcc acgccgagca cctggtcgct ttcagaaatc
     3781 aatctaaagt atatatgagt aaacttggtc tgacaggccc ctgaattcgc atctagatgg
     3841 tagagccaca aacagccggt acaagcaacg atctccagga ccatctgaat catgcgcgga
     3901 tgacacgaac tcacgacgct ttggcagttt attcttgaca tgtagtgagg gggctggtat
     3961 aatcacatag tactgttata cagaaacaga ggagatatta catatggata gcactgagaa
     4021 cgtcatcaag cccttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga
     4081 gttcgagatc gagggcgtgg gcgagggcaa gccctacgag ggcacccaga ccgccaagct
     4141 gcaagtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc cccagttctt
     4201 ctacggctcc aaggcgtaca tcaagcaccc cgccgacatc cccgactacc tcaagcagtc
     4261 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac
     4321 cgtgacccag gactcctccc tgcaggacgg caccctcatc taccacgtga agttcatcgg
     4381 cgtgaacttc ccctccgacg gccccgtaat gcagaagaag actctgggct gggagccctc
     4441 cactgagcgc aactaccccc gcgacggcgt gctgaagggc gagaaccaca tggcgctgaa
     4501 gctgaagggc ggcggccact acctgtgtga gttcaagtcc atctacatgg ccaagaagcc
     4561 cgtgaagctg cccggctacc actacgtgga ctacaagctc gacatcacct cccacaacga
     4621 ggactacacc gtggtggagc agtacgagcg cgccgaggcc cgccaccacc tgttccagac
     4681 tcacggtatg gacgaattgt acaagcacga cgaattgtaa atccgtaatc gttaatccgc
     4741 aaataacgta aaaacccgct tcggcgggtt tttttatggg gggagtttag ggaaagagca
     4801 tttgtcagct ggatcagcag atgggaaccg gatactcgag cggccgcgat tatcaaaaag
     4861 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     4921 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     4981 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5041 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgggacccac gctcaccggc
     5101 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5161 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5221 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5281 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5341 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5401 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5461 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5521 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     5581 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     5641 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     5701 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     5761 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     5821 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     5881 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgtcatgac
     5941 caaaatccct taacgtgagt cagcctgccg ccttgggccg ggtgatgtcg tacttgcccg
     6001 ccgcgaactc ggttaccgtc cagcccagcg cgaccagctc cggcaacgcc tcgcgcaccc
     6061 gctggcggcg cttgcgcatg gtcgaaccac tggcctctga cggccagaca tagccgcaca
     6121 aggtatctat ggaagccttg ccggttttgc cggggtcgat ccagccacac agccgctggt
     6181 gcagcaggcg ggcggtttcg ctgtccagcg cccgcacctc gtccatgctg atgcgcacat
     6241 gctggccgcc acccatgacg gcctgcgcga tcaaggggtt cagggccacg tacaggcgcc
     6301 cgtccgcctc gtcgctggcg tactccgaca gcagccgaaa cccctgccgc ttgcggccat
     6361 tctgggcgat gatggatacc ttccaaaggc gctcgatgca gtcctgtatg tgcttgagcg
     6421 ccccaccact atcgacctct gccccgattt cctttgccag cgcccgatag ctacctttga
     6481 ccaccatggc atcagcggtg acggcctccc acttgggttc caggaacagc cggagctgcc
     6541 gtccgccttc ggtcttgggt tccgggccaa gcactaggcc attaggccca gccatggcca
     6601 ccagcccttg caggatgcgc agatcatcag cgcccagcgg ctccgggccg ctgaactcga
     6661 tccgcttgcc gtcgccgtag tcatacgtca cgtccagctt gctgcgcttg cgctcgcccc
     6721 gcttgagggc acggaacagg ccgggggcca gacagtgcgc cgggtcgtgc cggacgtggc
     6781 tgaggctgtg cttgttctta ggcttcacca cggggcaccc ccttgctctt gcgctgcctc
     6841 tccagcacgg cgggcttgag caccccgccg tcatgccgcc tgaaccaccg atcagcgaac
     6901 ggtgcgccat agttggcctt gctcacaccg aagcggacga agaaccggcg ctggtcgtcg
     6961 tccacacccc attcctcggc ctcggcgctg gtcatgctcg acaggtagga ctgccagcgg
     7021 atgttatcga ccagtaccga gctgccccgg ctggcctgct gctggtcgcc tgcgcccatc
     7081 atggccgcgc ccttgctggc atggtgcagg aacacgatag agcacccggt atcggcggcg
     7141 atggcctcca tgcgaccgat gacctgggcc atggggccgc tggcgttttc ttcctcgatg
     7201 tggaaccggc gcagcgtgtc cagcaccatc aggcggcggc cctcggcggc gcgcttgagg
     7261 ccgtcgaacc actccggggc catgatgttg ggcaggctgc cgatcagcgg ctggatcagc
     7321 aggccgtcag ccacggcttg ccgttcctcg gcgctgaggt gcgccccaag ggcgtgcagg
     7381 cggtgatgaa tggcggtggg cgggtcttcg gcgggcaggt agatcaccgg gccggtgggc
     7441 agttcgccca cctccagcag atccggcccg cctgcaatct gtgcggccag ttgcagggcc
     7501 agcatggatt taccggcacc accgggcgac accagcgccc cgaccgtacc ggccaccatg
     7561 ttgggcaaaa cgtagtccag cggtggcggc gctgctgcga acgcctccag aatattgata
     7621 ggcttatggg tagccattga ttgcctcctt tgcaggcagt tggtggttag gcgctggcgg
     7681 ggtcactacc cccgccctgc gccgctctga gttcttccag gcactcgcgc agcgcctcgt
     7741 attcgtcgtc ggtcagccag aacttgcgct gacgcatccc tttggccttc atgcgctcgg
     7801 c
//
