LOCUS       sequence-446607-        3487 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mini-Tn7 cloning vector - empty backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3487)
  AUTHORS   Roghair Stroud MN, Vang DX, Halverson LJ
  TITLE     Optimized CRISPR Interference System for Investigating Pseudomonas 
            alloputida Genes Involved in Rhizosphere Microbiome Assembly.
  JOURNAL   ACS Synth Biol. 2024 Aug 20. doi: 10.1021/acssynbio.4c00312.
  PUBMED    39163848
REFERENCE   2  (bases 1 to 3487)
  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: "ACS Synth 
            Biol. 2024 Aug 20. doi: 10.1021/acssynbio.4c00312."
COMMENT     Sequence Label: pMRS-empty
FEATURES             Location/Qualifiers
     source          1..3487
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     51..73
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     RBS             154..162
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     terminator      256..350
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(652..671)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             663..1478
                     /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"
     oriT            1645..1753
                     /note="incP origin of transfer"
     rep_origin      1775..2163
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     mobile_element  2172..2337
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     terminator      complement(2351..2437)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    2472..2519
                     /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(2649..3182)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3371..3376,3377..3393,3394..3399))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    3440..3487
                     /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)."
ORIGIN
        1 agagcgcttt tgaagctaat tcgatcatgc atgagcgacg tcttaattaa agcggataac
       61 aatttcacac aggaggccgc ctagggccca agttcactta aaaaggagat caacaatgaa
      121 agcaattttc gtactgaaac atcttaatca tgctaaggag gtgagctcgg tacccgggga
      181 tcctctagag tcgacctgca ggcatgcaag cttgcggccg cgtcgtgact gggaaaaccc
      241 tggcgactag tcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg
      301 atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat ccagccacgc
      361 ccctctttaa tacgacgggc aatttgcact tcagaaaatg aagagtttgc tttagccata
      421 acaaaagtcc agtatgcttt ttcacagcat aactggactg atttcagttt acaactattc
      481 tgtctagttt aagactttat tgtcatagtt tagatctatt ttgttcagtt taagacttta
      541 ttgtccgccc acaatttaaa tttgtgtctc aaaatctctg atgttacatt gcacaagata
      601 aaaatatatc atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg
      661 ttatgagcca tattcagcgt gaaacgagct gtagccgtcc gcgtctgaac agcaacatgg
      721 atgcggatct gtatggctat aaatgggcgc gtgataacgt gggtcagagc ggcgcgacca
      781 tttatcgtct gtatggcaaa ccggatgcgc cggaactgtt tctgaaacat ggcaaaggca
      841 gcgtggcgaa cgatgtgacc gatgaaatgg tgcgtctgaa ctggctgacc gaatttatgc
      901 cgctgccgac cattaaacat tttattcgca ccccggatga tgcgtggctg ctgaccaccg
      961 cgattccggg caaaaccgcg tttcaggtgc tggaagaata tccggatagc ggcgaaaaca
     1021 ttgtggatgc gctggccgtg tttctgcgtc gtctgcatag cattccggtg tgcaactgcc
     1081 cgtttaacag cgatcgtgtg tttcgtctgg cccaggcgca gagccgtatg aacaacggcc
     1141 tggtggatgc gagcgatttt gatgatgaac gtaacggctg gccggtggaa caggtgtgga
     1201 aagaaatgca taaactgctg ccgtttagcc cggatagcgt ggtgacccac ggcgatttta
     1261 gcctggataa cctgattttc gatgaaggca aactgattgg ctgcattgat gtgggccgtg
     1321 tgggcattgc ggatcgttat caggatctgg ccattctgtg gaactgcctg ggcgaattta
     1381 gcccgagcct gcaaaaacgt ctgtttcaga aatatggcat tgataatccg gatatgaaca
     1441 aactgcaatt tcatctgatg ctggatgaat ttttctaata attaattgga ccgcggtccg
     1501 cgcgttgtcc ttttccgctg cataaccctg cttcggggtc attatagcga ttttttcggt
     1561 atatccatcc tttttcgcac gatatacagg attttgccaa agggttcgtg tagactttcc
     1621 ttggtgtatc caacggcgtc agccgggcag gataggtgaa gtaggcccac ccgcgagcgg
     1681 gtgttccttc ttcactgtcc cttattcgca cctggcggtg ctcaacggga atcctgctct
     1741 gcgaggctgg ccgtaggccg gccggcgcgc cccatgtcag ccgttaagtg ttcctgtgtc
     1801 actcaaaatt gctttgagag gctctaaggg cttctcagtg cgttacatcc ctggcttgtt
     1861 gtccacaacc gttaaacctt aaaggcttta aaagccttat atattctttt ttttcttata
     1921 aaacttaaaa ccttagaggc tatttaagtt gctgatttat attaatttta ttgttcaaac
     1981 atgagagctt agtacgtgaa acatgagagc ttagtacgtt agccatgaga gcttagtacg
     2041 ttagccatga gggtttagtt cgttaaacat gagagcttag tacgttaaac atgagagctt
     2101 agtacgtgaa acatgagagc ttagtacgta ctatcaacag gttgaactgc tgatcttcag
     2161 atcggcgcgc ctgtgggcgg acaaaatagt tgggaactgg gaggggtgga aatggagttt
     2221 ttaaggatta tttagggaag agtgacaaaa tagatgggaa ctgggtgtag cgtcgtaagc
     2281 taatacgaaa attaaaaatg acaaaatagt ttggaactag atttcactta tctggtttct
     2341 agggcggcgg atttgtccta ctcaggagag cgttcaccga caaacaacag ataaaacgaa
     2401 aggcccagtc tttcgactga gcctttcgtt ttatttgatg ccgacgtccg gacgatcgaa
     2461 ttggggatct tgaagtacct attccgaagt tcctattctc tagaaagtat aggaacttca
     2521 gagcgctttt gaagctgatg tgcttaaaaa cttactcaat ggaattctag atcgaattgg
     2581 ccgcggcgtt gtgacaattt accgaacaac tccgcggccg ggaagccgat ctcggcttga
     2641 acgaattgtt aggtggcggt acttgggtcg atatcaaagt gcatcacttc ttcccgtatg
     2701 cccaactttg tatagagagc cactgcggga tcgtcaccgt aatctgcttg cacgtagatc
     2761 acataagcac caagcgcgtt ggcctcatgc ttgaggagat tgatgagcgc ggtggcaatg
     2821 ccctgcctcc ggtgctcgcc ggagactgcg agatcataga tatagatctc actacgcggc
     2881 tgctcaaact tgggcagaac gtaagccgcg agagcgccaa caaccgcttc ttggtcgaag
     2941 gcagcaagcg cgatgaatgt cttactacgg agcaagttcc cgaggtaatc ggagtccggc
     3001 tgatgttggg agtaggtggc tacgtctccg aactcacgac cgaaaagatc aagagcagcc
     3061 cgcatggatt tgacttggtc agggccgagc ctacatgtgc gaatgatgcc catacttgag
     3121 ccacctaact ttgttttagg gcgactgccc tgctgcgtaa catcgttgct gctgcgtaac
     3181 atcgttgctg ctccataaca tcaaacatcg acccacggcg taacgcgctt gctgcttgga
     3241 tgcccgaggc atagactgta caaaaaaaca gtcataacaa gccatgaaaa ccgccactgc
     3301 gccgttacca ccgctgcgtt cggtcaaggt tctggaccag ttgcgtgagc gcatacgcta
     3361 cttgcattac agtttacgaa ccgaacaggc ttatgtcaat ccgatccatt gctgttgaca
     3421 aagggaatca ggggatcttg aagttcctat tccgaagttc ctattctcta gaaagtatag
     3481 gaacttc
//
