LOCUS       sequence-479122-        6673 bp    DNA     circular SYN 21-JUL-2025
DEFINITION  MiniTn7 donor plasmid for constructing a ratiometric reporter for P 
            limitation in Pseudomonas synxantha. A weak Anderson promoter 
            (J23116) drives expression of mScarlet-I.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6673)
  AUTHORS   Larsson EM, Murray RM, Newman DK
  TITLE     Engineering the Soil Bacterium Pseudomonas synxantha 2-79 into a 
            Ratiometric Bioreporter for Phosphorus Limitation.
  JOURNAL   ACS Synth Biol. 2024 Jan 19;13(1):384-393. doi: 
            10.1021/acssynbio.3c00642. Epub 2024 Jan 2.
  PUBMED    38165130
REFERENCE   2  (bases 1 to 6673)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 21, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acssynbio.3c00642"; journalName: "ACS Synth Biol"; date: 
            "2024-01-19- 19"; volume: "13"; issue: "1"; pages: "384-393"
COMMENT     Sequence Label: pJM220-PphoA-mNeongreen-J23116-mScarletI
FEATURES             Location/Qualifiers
     source          1..6673
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     584..600
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     585..601
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      629..723
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      826..912
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    958..1005
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             complement(1135..1668)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1857..1862,1863..1879,1880..1885))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    1926..1973
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     RBS             2223..2234
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        3052..3086
                     /label=BBa_J23116
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.16"
     terminator      4007..4093
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     mobile_element  4190..4355
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     oriT            4514..4622
                     /note="incP origin of transfer"
     rep_origin      complement(4854..5442)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4934..4953)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5613..6404,6405..6473))
                     /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     6236..6255
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6474..6578)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6646..6664
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgacgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct agaggaccag ccgcgtaacc tggcaaaatc ggttacggtt gagtaataaa
      361 tggatgccct gcgtaagcag gtgtgggcgg acaataaagt cttaaactga acaaaataga
      421 tctaaactat gacaataaag tcttaaacta gacagaatag ttgtaaactg aaatcagtcc
      481 agttatgctg tgaaaaagca tactggactt ttgttatggc taaagcaaac tcttcatttt
      541 ctgaagtgca aattgcccgt cgtattaaag aggggcgtgg ggttcgaggt cgacggtatc
      601 gataagctag cttaattagc tgagcttgga ctcctgttga tagatccagt aatgacctca
      661 gaactccatc tggatttgtt cagaacgctc ggttgccgcc gggcgttttt tattggtgag
      721 aatccaagct agactgcgat gagtggcagg gcggggcgta atttttttaa ggcagttatt
      781 ggtgccctta aacgcctggg gtaatgactc tctagcttga ggcatcaaat aaaacgaaag
      841 gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg
      901 agtaggacaa atccgccgct aggagcttgc ggcccggacg atcgaattgg ggatcttgaa
      961 gtacctattc cgaagttcct attctctaga aagtatagga acttcagagc gcttttgaag
     1021 ctgatgtgct taaaaactta ctcaatggaa ttctagatcg aattggccgc ggcgttgtga
     1081 caatttaccg aacaactccg cggccgggaa gccgatctcg gcttgaacga attgttaggt
     1141 ggcggtactt gggtcgatat caaagtgcat cacttcttcc cgtatgccca actttgtata
     1201 gagagccact gcgggatcgt caccgtaatc tgcttgcacg tagatcacat aagcaccaag
     1261 cgcgttggcc tcatgcttga ggagattgat gagcgcggtg gcaatgccct gcctccggtg
     1321 ctcgccggag actgcgagat catagatata gatctcacta cgcggctgct caaacttggg
     1381 cagaacgtaa gccgcgagag cgccaacaac cgcttcttgg tcgaaggcag caagcgcgat
     1441 gaatgtctta ctacggagca agttcccgag gtaatcggag tccggctgat gttgggagta
     1501 ggtggctacg tctccgaact cacgaccgaa aagatcaaga gcagcccgca tggatttgac
     1561 ttggtcaggg ccgagcctac atgtgcgaat gatgcccata cttgagccac ctaactttgt
     1621 tttagggcga ctgccctgct gcgtaacatc gttgctgctg cgtaacatcg ttgctgctcc
     1681 ataacatcaa acatcgaccc acggcgtaac gcgcttgctg cttggatgcc cgaggcatag
     1741 actgtacaaa aaaacagtca taacaagcca tgaaaaccgc cactgcgccg ttaccaccgc
     1801 tgcgttcggt caaggttctg gaccagttgc gtgagcgcat acgctacttg cattacagtt
     1861 tacgaaccga acaggcttat gtcaattcga tccattgctg ttgacaaagg gaatcagggg
     1921 atcttgaagt tcctattccg aagttcctat tctctagaaa gtataggaac ttcagagcgc
     1981 ttttgaagct aattcgatca tgcatgagct caaagagtgg aacaatgcag gacgccgtcc
     2041 ggtaacggtg ttgagcctcg gttgtgtggt gattattgtc gccgctaaca tcgtcggcat
     2101 cgagagcgct tttgaagcta attcgatcat gcatgagctc aaaagttaat cttttcaaca
     2161 gctgtcataa agttgtcacg gccgagactt atagtcgctt tgtttttatt ttttaatgta
     2221 ttaaagagga gaaattaagc atggtctcca agggcgagga agataatatg gcgtccctcc
     2281 cagcgacgca tgaactccat atcttcgggt cgatcaacgg ggtggacttt gatatggtgg
     2341 ggcaagggac gggcaacccc aatgatggct acgaagagct caacctcaag agcacgaagg
     2401 gcgacctgca gtttagcccc tggatcctgg tgccgcacat cgggtatggc ttccatcaat
     2461 atctgcccta ccccgacggg atgtccccgt ttcaggcggc catggtcgac ggctcggggt
     2521 accaagtgca tcgcacgatg cagtttgagg atggcgcctc cctgaccgtg aactatcgct
     2581 acacgtacga aggcagccat atcaaggggg aagcgcaggt gaagggcacc ggctttccgg
     2641 cggacgggcc ggtgatgacc aattccctga cggccgcgga ttggtgtcgg tccaagaaga
     2701 cgtacccgaa cgataagacc atcatcagca cgttcaagtg gtcgtatacc accggcaacg
     2761 gcaagcgcta ccgttcgacg gcccgcacga cgtacacctt tgccaagccg atggcggcga
     2821 actatctgaa gaaccaaccc atgtatgtgt tccggaagac cgagctcaag cattcgaaga
     2881 cggagctgaa ttttaaggaa tggcagaagg cgtttaccga tgtgatgggg atggatgagc
     2941 tctataagta agactcctgt tgatagatcc agtaatgacc tcagaactcc atctggattt
     3001 gttcagaacg ctcggttgca ttggtgagaa tagactagtg ggtctcagga gttgacagct
     3061 agctcagtcc tagggactat gctagctact agagacctac tagtagcgga tgaaattcaa
     3121 ctagtgctct gcagtaagga ggaacagcta tgagtaaagg agaagctgtg attaaagagt
     3181 tcatgcgctt caaagttcac atggagggtt ctatgaacgg tcacgagttc gagatcgaag
     3241 gcgaaggcga gggccgtccg tatgaaggca cccagaccgc caaactgaaa gtgactaaag
     3301 gcggcccgct gcctttttcc tgggacatcc tgagcccgca atttatgtac ggttctaggg
     3361 cgttcatcaa acacccagcg gatatcccgg actattataa gcagtctttt ccggaaggtt
     3421 tcaagtggga acgcgtaatg aattttgaag atggtggtgc cgtgaccgtc actcaggaca
     3481 cctccctgga ggatggcacc ctgatctata aagttaaact gcgtggtact aattttccac
     3541 ctgatggccc ggtgatgcag aaaaagacga tgggttggga ggcgtctacc gaacgcttgt
     3601 atccggaaga tggtgtgctg aaaggcgaca ttaaaatggc cctgcgcctg aaagatggcg
     3661 gccgctatct ggctgacttc aaaaccacgt acaaagccaa gaaacctgtg cagatgcctg
     3721 gcgcgtacaa tgtggaccgc aaactggaca tcacctctca taatgaagat tatacggtgg
     3781 tagagcaata tgagcgctcc gagggtcgtc attctaccgg tggcatggat gaactataca
     3841 aataatcgcg aatcagaacg cagaagcggt ctgataaaac agaatttgcc tggcggcagt
     3901 agcgcggtgg tcccacctga ccccatgccg aactcagaag tgaaacgccg tagcgccgat
     3961 ggtagtgtgg ggtctcccca tgcgagagta gggaactgcc aggcatcaaa taaaacgaaa
     4021 ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcct
     4081 gagtaggaca aatccgccgg gagcggattt gaacgttgcg aagcaacggc ccggagggtg
     4141 gcgggcagga cgcccgccat aaactgccag gcataaggcc ttgcaggcca accagataag
     4201 tgaaatctag ttccaaacta ttttgtcatt tttaattttc gtattagctt acgacgctac
     4261 acccagttcc catctatttt gtcactcttc cctaaataat ccttaaaaac tccatttcca
     4321 cccctcccag ttcccaacta ttttgtccgc ccacagcggg gcatttttct tcctgttatg
     4381 tttgggcgct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4441 gctcttccgc ttcctcgctc actgacccgc tgcgctcggt cgttcggctg cggcgagcgg
     4501 tatcagagct tatcggccag cctcgcagag caggattccc gttgagcacc gccaggtgcg
     4561 aataagggac agtgaagaag gaacacccgc tcgcgggtgg gcctacttca cctatcctgc
     4621 ccggctgacg ccgttggata caccaaggaa agtctacacg aaccctttgg caaaatcctg
     4681 tatatcgtgc gaaaaaggat ggatataccg aaaaaatcgc tataatgacc ccgaagcagg
     4741 gttatgcagc ggaaagtata ccttaaggaa tcccctgata acgcaggaaa gaacatgtga
     4801 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4861 aggctccgcc cccctgacga gcatcacaaa aatcgacgct cgagtcagag gtggcgaaac
     4921 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4981 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     5041 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     5101 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     5161 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     5221 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     5281 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     5341 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     5401 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5461 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     5521 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     5581 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     5641 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     5701 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     5761 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5821 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5881 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5941 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     6001 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     6061 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     6121 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     6181 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     6241 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6301 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     6361 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     6421 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6481 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     6541 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     6601 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     6661 aggccctttc gtc
//
