LOCUS       sequence-357219-        7602 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Kan resistant plasmid derived from pBTK402, a broad-host-range 
            plasmid previously shown to replicate in multiple species of bee gut
            bacteria. It encodes an E2 Crimson promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7602)
  AUTHORS   Li Y, Leonard SP, Powell JE, Moran NA
  TITLE     Species divergence in gut-restricted bacteria of social bees.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 May 3;119(18):e2115013119. doi: 
            10.1073/pnas.2115013119. Epub 2022 Apr 25.
  PUBMED    35467987
REFERENCE   2  (bases 1 to 7602)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2022-05-3"; volume: "119(18):e2115013119. 
            doi"; pages: " 10.1073/pnas.2115013119. Epub 2022 Apr 25"
COMMENT     Sequence Label: pSL1-E2C
FEATURES             Location/Qualifiers
     source          1..7602
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            2118..2205
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      2547..2941
                     /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             3478..4152
                     /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     3876..3895
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             complement(4332..5147)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4332..5147)
                     /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
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5055..5074
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5139..5158
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(5322..6173)
                     /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(6160..6999)
                     /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"
     CDS             complement(join(7510..7602,1..879))
                     /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
                     EYRSEMAGLVKRFGDDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
ORIGIN
        1 cagacccatg accttgctga cggtgcgctc gatgtaatcc gcttcgtggc cgggcttgcg
       61 ctctgccagc gctgggctgg cctcggccat ggccttgccg atttcctcgg cactgcggcc
      121 ccggctggcc agcttctgcg cggcgataaa gtcgcacttg ctgaggtcat caccgaagcg
      181 cttgaccagc ccggccatct cgctgcggta ctcgtccagc gccgtgcgcc ggtggcggct
      241 aagctgccgc tcgggcagtt cgaggctggc cagcctgcgg gccttctcct gctgccgctg
      301 ggcctgctcg atctgctggc cagcctgctg caccagcgcc gggccagcgg tggcggtctt
      361 gcccttggat tcacgcagca gcacccacgg ctgataaccg gcgcgggtgg tgtgcttgtc
      421 cttgcggttg gtgaagcccg ccaagcggcc atagtggcgg ctgtcggcgc tggccgggtc
      481 ggcgtcgtac tcgctggcca gcgtccgggc aatctgcccc cgaagttcac cgcctgcggc
      541 gtcggccacc ttgacccatg cctgatagtt cttcgggctg gtttccacta ccagggcagg
      601 ctcccggccc tcggctttca tgtcatccag gtcaaactcg ctgaggtcgt ccaccagcac
      661 cagaccatgc cgctcctgct cggcgggcct gatatacacg tcattgccct gggcattcat
      721 ccgcttgagc catggcgtgt tctggagcac ttcggcggct gaccattccc ggttcatcat
      781 ctggccggtg gtggcgtccc tgacgccgat atcgaagcgc tcacagccca tggccttgag
      841 ctgtcggcct atggcctgca aagtcctgtc gttcttcatc gggccaccaa gcgcagccag
      901 atcgagccgt cctcggttgt cagtggcgtc aggtcgagca agagcaacga tgcgatcagc
      961 agcaccaccg taggcatcat ggaagccagc atcacggtta gccatagctt ccagtgccac
     1021 ccccgcgacg cgctccgggc gctctgcgcg gcgctgctca cctcggcggc tacctcccgc
     1081 aactctttgg ccagctccac ccatgccgcc cctgtctggc gctgggcttt cagccactcc
     1141 gccgcctgcg cctcgctggc ctgctgggtc tggctcatga cctgccgggc ttcgtcggcc
     1201 agtgtcgcca tgctctgggc cagcggttcg atctgctccg ctaactcgtt gatgcctctg
     1261 gatttcttca ctctgtcgat tgcgttcatg gtctattgcc tcccggtatt cctgtaagtc
     1321 gatgatctgg gcgttggcgg tgtcgatgtt cagggccacg tctgcccggt cggtgcggat
     1381 gccccggcct tccatctcca ccacgttcgg ccccaggtga acaccgggca ggcgctcgat
     1441 gccctgcgcc tcaagtgttc tgtggtcaat gcgggcgtcg tggccagccc gctctaatgc
     1501 ccggttggca tggtcggccc atgcctcgcg ggtctgctca agccatgcct tgggcttgag
     1561 cgcttcggtc ttctgtgccc cgcccttctc cggggtcttg ccgttgtacc gcttgaacca
     1621 ctgagcggcg ggccgctcga tgccgtcatt gatccgctcg gagatcatca ggtggcagtg
     1681 cgggttctcg ccgccaccgg catggatggc cagcgtatac ggcaggcgct cggcaccggt
     1741 caggtgctgg gcgaactcgg acgccagcgc cttctgctgg tcgagggtca gctcgaccgg
     1801 cagggcaaat tcgacctcct tgaacagccg cccattggcg cgttcataca ggtcggcagc
     1861 atcccagtag tcggcgggcc gctcgacgaa ctccggcatg tgcccggatt cggcgtgcaa
     1921 gacttcatcc atgtcgcggg catacttgcc ttcgcgctgg atgtagtcgg ccttggccct
     1981 ggccgattgg ccgcccgacc tgctgccggt tttcgccgta aggtgataaa tcgccatgct
     2041 gcctcgctgt tgcttttgct tttcggctcc atgcaatggc cctcggagag cgcaccgccc
     2101 gaagggtggc cgttaggcca gtttctcgaa gagaaaccgg taagtgcgcc ctcccctaca
     2161 aagtagggtc gggattgccg ccgctgtgcc tccatgatag cctacgagac agcacattaa
     2221 caatggggtg tcaagatggt taaggggagc aacaaggcgg cggatcggct ggccaagctc
     2281 gaagaacaac gagcgcgaat caatgccgaa attcagcggg tgcgggcaag ggaacagcag
     2341 caagagcgca agaacgaaac aaggcgcaag gtgctggtgg gggccatgat tttggccaag
     2401 gtgaacagca gcgagtggcc ggaggatcgg ctcatggcgg caatggatgc gtaccttgaa
     2461 cgcgaccacg accgcgcctt gttcggtctg ccgccacgcc agaaggatga gccgggctga
     2521 atgatcgacc gagacaggcc ctgcggggct gcacacgcgc ccccaccctt cgggtagggg
     2581 gaaaggccgc taaagcggct aaaagcgctc cagcgtattt ctgcggggtt tggtgtgggg
     2641 tttagcgggc tttgcccgcc tttccccctg ccgcgcagcg gtggggcggt gtgtagccta
     2701 gcgcagcgaa tagaccagct atccggcctc tggccgggca tattgggcaa gggcagcagc
     2761 gccccacaag ggcgctgata accgcgccta gtggattatt cttagataat catggatgga
     2821 tttttccaac accccgccag cccccgcccc tgctgggttt gcaggtttgg gggcgtgaca
     2881 gttattgcag gggttcgtga cagttattgc aggggggcgt gacagttatt gcaggggttc
     2941 gtgacagtta gtacgggagt gacgggcact ggctggcaat gtctagcaac ggcaggcatt
     3001 tcggctgagg gtaaaagaac tttccgctaa gcgatagact gtatgtaaac acagtattgc
     3061 aaggacgcgg aacatgcctc atgtggcggc caggacggcc agccgggatc gggatactgg
     3121 tcgttaccag agccaccgac ccgagcaaac ccttctctat cagatcgttg acgagtatta
     3181 cccggcattc gctgcgctta tggcagagca gggaaaggaa ttgccgggct atgtgcaacg
     3241 ggaatttgaa gaatttctcc aatgcgggcg gctggagcat ggctttctac gggttcgctg
     3301 cgagtcttgc cacgccgagc acctggtcgc tttcagaaat caatctaaag tatatatgag
     3361 taaacttggt ctgacaggcc ccttgagacc agctttggca gtttattctt gacatgtagt
     3421 gagggggctg gtataatcac atagtactgt tatacagaaa cagaggagat attacatatg
     3481 gatagcactg agaacgtcat caagcccttc atgcgcttca aggtgcacat ggagggctcc
     3541 gtgaacggcc acgagttcga gatcgagggc gtgggcgagg gcaagcccta cgagggcacc
     3601 cagaccgcca agctgcaagt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg
     3661 tccccccagt tcttctacgg ctccaaggcg tacatcaagc accccgccga catccccgac
     3721 tacctcaagc agtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac
     3781 ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcaccct catctaccac
     3841 gtgaagttca tcggcgtgaa cttcccctcc gacggccccg taatgcagaa gaagactctg
     3901 ggctgggagc cctccactga gcgcaactac ccccgcgacg gcgtgctgaa gggcgagaac
     3961 cacatggcgc tgaagctgaa gggcggcggc cactacctgt gtgagttcaa gtccatctac
     4021 atggccaaga agcccgtgaa gctgcccggc taccactacg tggactacaa gctcgacatc
     4081 acctcccaca acgaggacta caccgtggtg gagcagtacg agcgcgccga ggcccgccac
     4141 cacctgttcc agactcacgg tatggacgaa ttgtacaagc acgacgaatt gtaaatccgt
     4201 aatcgttaat ccgcaaataa cgtaaaaacc cgcttcggcg ggttttttta tggggggagt
     4261 ttagggaaag agcatttgtc aagtcggtct caccgagcgg ccgcgtgtta caaccaatta
     4321 accaattctg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca
     4381 ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg
     4441 aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca
     4501 tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca
     4561 tgagtgacga ctgaatccgg tgagaatggc aaaagcttat gcatttcttt ccagacttgt
     4621 tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc
     4681 attcgtgatt gcgcctgagc gaggcgaaat acgcgatcgc tgttaaaagg acaattacaa
     4741 acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct
     4801 gaatcaggat attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt
     4861 aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc
     4921 gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca
     4981 tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct
     5041 gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa
     5101 tttaatcgcg gcctggagca agacgtttcc cgttgaatat ggctcataac accccttgta
     5161 ttactgttta tgtaagcaga cagttttatt gttcatgatg atatattttt atcttgtgca
     5221 atgtaacatc agagattttg agacacaacg tggctttgtt gaataaatcg aacttttgct
     5281 gagttgaagg atcagtcatg accaaaatcc cttaacgtga gtcagcctgc cgccttgggc
     5341 cgggtgatgt cgtacttgcc cgccgcgaac tcggttaccg tccagcccag cgcgaccagc
     5401 tccggcaacg cctcgcgcac ccgctggcgg cgcttgcgca tggtcgaacc actggcctct
     5461 gacggccaga catagccgca caaggtatct atggaagcct tgccggtttt gccggggtcg
     5521 atccagccac acagccgctg gtgcagcagg cgggcggttt cgctgtccag cgcccgcacc
     5581 tcgtccatgc tgatgcgcac atgctggccg ccacccatga cggcctgcgc gatcaagggg
     5641 ttcagggcca cgtacaggcg cccgtccgcc tcgtcgctgg cgtactccga cagcagccga
     5701 aacccctgcc gcttgcggcc attctgggcg atgatggata ccttccaaag gcgctcgatg
     5761 cagtcctgta tgtgcttgag cgccccacca ctatcgacct ctgccccgat ttcctttgcc
     5821 agcgcccgat agctaccttt gaccaccatg gcatcagcgg tgacggcctc ccacttgggt
     5881 tccaggaaca gccggagctg ccgtccgcct tcggtcttgg gttccgggcc aagcactagg
     5941 ccattaggcc cagccatggc caccagccct tgcaggatgc gcagatcatc agcgcccagc
     6001 ggctccgggc cgctgaactc gatccgcttg ccgtcgccgt agtcatacgt cacgtccagc
     6061 ttgctgcgct tgcgctcgcc ccgcttgagg gcacggaaca ggccgggggc cagacagtgc
     6121 gccgggtcgt gccggacgtg gctgaggctg tgcttgttct taggcttcac cacggggcac
     6181 ccccttgctc ttgcgctgcc tctccagcac ggcgggcttg agcaccccgc cgtcatgccg
     6241 cctgaaccac cgatcagcga acggtgcgcc atagttggcc ttgctcacac cgaagcggac
     6301 gaagaaccgg cgctggtcgt cgtccacacc ccattcctcg gcctcggcgc tggtcatgct
     6361 cgacaggtag gactgccagc ggatgttatc gaccagtacc gagctgcccc ggctggcctg
     6421 ctgctggtcg cctgcgccca tcatggccgc gcccttgctg gcatggtgca ggaacacgat
     6481 agagcacccg gtatcggcgg cgatggcctc catgcgaccg atgacctggg ccatggggcc
     6541 gctggcgttt tcttcctcga tgtggaaccg gcgcagcgtg tccagcacca tcaggcggcg
     6601 gccctcggcg gcgcgcttga ggccgtcgaa ccactccggg gccatgatgt tgggcaggct
     6661 gccgatcagc ggctggatca gcaggccgtc agccacggct tgccgttcct cggcgctgag
     6721 gtgcgcccca agggcgtgca ggcggtgatg aatggcggtg ggcgggtctt cggcgggcag
     6781 gtagatcacc gggccggtgg gcagttcgcc cacctccagc agatccggcc cgcctgcaat
     6841 ctgtgcggcc agttgcaggg ccagcatgga tttaccggca ccaccgggcg acaccagcgc
     6901 cccgaccgta ccggccacca tgttgggcaa aacgtagtcc agcggtggcg gcgctgctgc
     6961 gaacgcctcc agaatattga taggcttatg ggtagccatt gattgcctcc tttgcaggca
     7021 gttggtggtt aggcgctggc ggggtcacta cccccgccct gcgccgctct gagttcttcc
     7081 aggcactcgc gcagcgcctc gtattcgtcg tcggtcagcc agaacttgcg ctgacgcatc
     7141 cctttggcct tcatgcgctc ggcatatcgc gcttggcgta cagcgtcagg gctggccagc
     7201 aggtcgccgg tctgcttgtc cttttggtct ttcatatcag tcaccgagaa acttgccggg
     7261 gccgaaaggc ttgtcttcgc ggaacaagga caaggtgcag ccgtcaaggt taaggctggc
     7321 catatcagcg actgaaaagc ggccagcctc ggccttgttt gacgtataac caaagccacc
     7381 gggcaaccaa tagcccttgt cacttttgat caggtagacc gaccctgaag cgcttttttc
     7441 gtattccata aaaccccctt ctgtgcgtga gtactcatag tataacaggc gtgagtacca
     7501 acgcaagcac tacatgctga aatctggccc gcccctgtcc atgcctcgct ggcggggtgc
     7561 cggtgcccgt gccagctcgg cccgcgcaag ctggacgctg gg
//
