LOCUS       sequence-479123-        6688 bp    DNA     circular SYN 21-JUL-2025
DEFINITION  MiniTn7 donor plasmid for constructing a ratiometric reporter for P 
            limitation in Pseudomonas synxantha. A strong Anderson promoter 
            (J23101) drives expression of mScarlet-I.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6688)
  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 6688)
  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-PhoA-mNeongreen-J23101-mScarletI
FEATURES             Location/Qualifiers
     source          1..6688
                     /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)"
     terminator      2952..3046
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        3067..3101
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     terminator      4022..4108
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     mobile_element  4205..4370
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     oriT            4529..4637
                     /note="incP origin of transfer"
     rep_origin      complement(4869..5457)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4949..4968)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5628..6419,6420..6488))
                     /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     6251..6270
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6489..6593)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6661..6679
                     /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 ctcggttgcc gccgggcgtt ttttattggt gagaatagac tagtgggtct
     3061 caggagttta cagctagctc agtcctaggt attatgctag ctactagaga cctactagta
     3121 gcggatgaaa ttcaactagt gctctgcagt aaggaggaac agctatgagt aaaggagaag
     3181 ctgtgattaa agagttcatg cgcttcaaag ttcacatgga gggttctatg aacggtcacg
     3241 agttcgagat cgaaggcgaa ggcgagggcc gtccgtatga aggcacccag accgccaaac
     3301 tgaaagtgac taaaggcggc ccgctgcctt tttcctggga catcctgagc ccgcaattta
     3361 tgtacggttc tagggcgttc atcaaacacc cagcggatat cccggactat tataagcagt
     3421 cttttccgga aggtttcaag tgggaacgcg taatgaattt tgaagatggt ggtgccgtga
     3481 ccgtcactca ggacacctcc ctggaggatg gcaccctgat ctataaagtt aaactgcgtg
     3541 gtactaattt tccacctgat ggcccggtga tgcagaaaaa gacgatgggt tgggaggcgt
     3601 ctaccgaacg cttgtatccg gaagatggtg tgctgaaagg cgacattaaa atggccctgc
     3661 gcctgaaaga tggcggccgc tatctggctg acttcaaaac cacgtacaaa gccaagaaac
     3721 ctgtgcagat gcctggcgcg tacaatgtgg accgcaaact ggacatcacc tctcataatg
     3781 aagattatac ggtggtagag caatatgagc gctccgaggg tcgtcattct accggtggca
     3841 tggatgaact atacaaataa tcgcgaatca gaacgcagaa gcggtctgat aaaacagaat
     3901 ttgcctggcg gcagtagcgc ggtggtccca cctgacccca tgccgaactc agaagtgaaa
     3961 cgccgtagcg ccgatggtag tgtggggtct ccccatgcga gagtagggaa ctgccaggca
     4021 tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct gttgtttgtc
     4081 ggtgaacgct ctcctgagta ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca
     4141 acggcccgga gggtggcggg caggacgccc gccataaact gccaggcata aggccttgca
     4201 ggccaaccag ataagtgaaa tctagttcca aactattttg tcatttttaa ttttcgtatt
     4261 agcttacgac gctacaccca gttcccatct attttgtcac tcttccctaa ataatcctta
     4321 aaaactccat ttccacccct cccagttccc aactattttg tccgcccaca gcggggcatt
     4381 tttcttcctg ttatgtttgg gcgctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     4441 gtttgcgtat tgggcgctct tccgcttcct cgctcactga cccgctgcgc tcggtcgttc
     4501 ggctgcggcg agcggtatca gagcttatcg gccagcctcg cagagcagga ttcccgttga
     4561 gcaccgccag gtgcgaataa gggacagtga agaaggaaca cccgctcgcg ggtgggccta
     4621 cttcacctat cctgcccggc tgacgccgtt ggatacacca aggaaagtct acacgaaccc
     4681 tttggcaaaa tcctgtatat cgtgcgaaaa aggatggata taccgaaaaa atcgctataa
     4741 tgaccccgaa gcagggttat gcagcggaaa gtatacctta aggaatcccc tgataacgca
     4801 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4861 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcgagt
     4921 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4981 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5041 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5101 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5161 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5221 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5281 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     5341 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5401 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     5461 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     5521 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     5581 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     5641 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     5701 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     5761 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     5821 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     5881 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     5941 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     6001 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     6061 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     6121 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6181 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6241 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     6301 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     6361 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     6421 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     6481 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     6541 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     6601 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
     6661 aataggcgta tcacgaggcc ctttcgtc
//
