LOCUS       sequence-128615-        5430 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ZeoR mini-Tn7 vector, mobilizable, for GFP tagging bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5430)
  AUTHORS   Choi KH, Schweizer HP
  TITLE     mini-Tn7 insertion in bacteria with single attTn7 sites: example 
            Pseudomonas aeruginosa.
  JOURNAL   Nat Protoc. 2006;1(1):153-61.
  PUBMED    17406227
REFERENCE   2  (bases 1 to 5430)
  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: "Nat 
            Protoc."; date: "2006"; volume: "1(1)"; pages: "153-61"
COMMENT     Sequence Label: pUC18T-mini-Tn7T-Zeo-gfpmut3
FEATURES             Location/Qualifiers
     source          1..5430
                     /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    complement(966..1013)
                     /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(1096..1470)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(1489..1536)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     protein_bind    complement(1647..1694)
                     /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)."
     terminator      complement(1748..1834)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(1937..2031)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2054..2767)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(2094..2114)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     2709..2737
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     RBS             2774..2785
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     mobile_element  2947..3112
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     oriT            3271..3379
                     /note="incP origin of transfer"
     rep_origin      complement(3611..4199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3691..3710)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4370..5161,5162..5230))
                     /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     4993..5012
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5231..5335)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5403..5421
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac 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 gcgtaagcgg 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 atcgaattag cttcaaaagc
      961 gctctgaagt tcctatactt tctagagaat aggaacttcg gaataggaac ttcaagatcc
     1021 cctgattccc tttgtcaaca gcaatggatt tatctctgat gttacattgc acaagataaa
     1081 aatatatcat catgatcagt cctgctcctc ggccacgaag tgcacgcagt tgccggccgg
     1141 gtcgcgcagg gcgaactccc gcccccacgg ctgctcgccg atctcggtca tggccggccc
     1201 ggaggcgtcc cggaagttcg tggacacgac ctccgaccac tcggcgtaca gctcgtccag
     1261 gccgcgcacc cacacccagg ccagggtgtt gtccggcacc acctggtcct ggaccgcgct
     1321 gatgaacagg gtcacgtcgt cccggaccac accggcgaag tcgtcctcca cgaagtcccg
     1381 ggagaacccg agccggtcgg tccagaactc gaccgctccg gcgacgtcgc gcgcggtgag
     1441 caccggaacg gcactggtca acttggccat ggtttagttc ctcaccttgt cgtattatac
     1501 tatgccgata tactatgccg atgattaatt gtcaacacgt gctgatcatg accaaaatcc
     1561 cttaacgtga gttacgcgtc gaagccgaat atctagaatt attccattga gtaagttttt
     1621 aagcacatca gcttcaaaag cgctctgaag ttcctatact ttctagagaa taggaacttc
     1681 ggaataggta cttcaagatc cccaattcga tcatgcatga gctcgggccg caagctccta
     1741 gcggcggatt tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg
     1801 cccagtcttt cgactgagcc tttcgtttta tttgatgcct caagctagag agtcattacc
     1861 ccaggcgttt aagggcacca ataactgcct taaaaaaatt acgccccgcc ctgccactca
     1921 tcgcagtcta gcttggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga
     1981 acaaatccag atggagttct gaggtcatta ctggatctat caacaggagt ccaagctcag
     2041 ctaattaagc ttatttgtat agttcatcca tgccatgtgt aatcccagca gctgttacaa
     2101 actcaagaag gaccatgtgg tctctctttt cgttgggatc tttcgaaagg gcagattgtg
     2161 tggacaggta atggttgtct ggtaaaagga cagggccatc gccaattgga gtattttgtt
     2221 gataatggtc tgctagttga acgcttccat cttcaatgtt gtgtctaatt ttgaagttaa
     2281 ctttgattcc attcttttgt ttgtctgcca tgatgtatac attgtgtgag ttatagttgt
     2341 attccaattt gtgtccaaga atgtttccat cttctttaaa atcaatacct tttaactcga
     2401 ttctattaac aagggtatca ccttcaaact tgacttcagc acgtgtcttg tagttcccgt
     2461 catctttgaa aaatatagtt ctttcctgta cataaccttc gggcatggca ctcttgaaaa
     2521 agtcatgctg tttcatatga tctgggtatc tcgcaaagca ttgaacacca taaccgaaag
     2581 tagtgacaag tgttggccat ggaacaggta gttttccagt agtgcaaata aatttaaggg
     2641 taagttttcc gtatgttgca tcaccttcac cctctccact gacagaaaat ttgtgcccat
     2701 taacatcacc atctaattca acaagaattg ggacaactcc agtgaaaagt tcttctcctt
     2761 tacgcatgct taatttctcc tctttaattc tagagtggtg tcgcattata gggagttatt
     2821 ccggcctgac tagaggaaat ttaaaataat tttctgaccg cgcaacgagc tcactagtgg
     2881 atcccccggg ctgcaggaat tcctcgagaa gcttgggccc ggtacctcgc gaaggccttg
     2941 caggccaacc agataagtga aatctagttc caaactattt tgtcattttt aattttcgta
     3001 ttagcttacg acgctacacc cagttcccat ctattttgtc actcttccct aaataatcct
     3061 taaaaactcc atttccaccc ctcccagttc ccaactattt tgtccgccca cagcggggca
     3121 tttttcttcc tgttatgttt gggcgctgca ttaatgaatc ggccaacgcg cggggagagg
     3181 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gacccgctgc gctcggtcgt
     3241 tcggctgcgg cgagcggtat cagagcttat cggccagcct cgcagagcag gattcccgtt
     3301 gagcaccgcc aggtgcgaat aagggacagt gaagaaggaa cacccgctcg cgggtgggcc
     3361 tacttcacct atcctgcccg gctgacgccg ttggatacac caaggaaagt ctacacgaac
     3421 cctttggcaa aatcctgtat atcgtgcgaa aaaggatgga tataccgaaa aaatcgctat
     3481 aatgaccccg aagcagggtt atgcagcgga aagtatacct taaggaatcc cctgataacg
     3541 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3601 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcga
     3661 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3721 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3781 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3841 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3901 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3961 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4021 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     4081 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4141 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4201 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4261 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4321 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4381 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4441 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4501 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4561 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4621 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4681 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4741 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4801 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4861 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4921 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     4981 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     5041 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     5101 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5161 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5221 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5281 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5341 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     5401 aaaataggcg tatcacgagg ccctttcgtc
//
