LOCUS       sequence-291998-        3671 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mUAVbla containing a tetracycline resistance (tetA) cassette for 
            building Gene Doctoring donor plasmids from pDOC-GG, using BsaI 
            Golden Gate assembly with GGAG and CGCT overhangs.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3671)
  AUTHORS   Thomson NM, Zhang C, Trampari E, Pallen MJ
  TITLE     Creation of Golden Gate constructs for gene doctoring.
  JOURNAL   BMC Biotechnol. 2020 Oct 7;20(1):54. doi: 
            10.1186/s12896-020-00648-5.
  PUBMED    33028286
REFERENCE   2  (bases 1 to 3671)
  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: "BMC 
            Biotechnol."; date: "2020-10-7"; volume: "20(1):54. doi"; pages: " 
            10.1186/s12896-020-00648-5"
COMMENT     Sequence Label: mUAVbla:tetR 5′
FEATURES             Location/Qualifiers
     source          1..3671
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      14..71
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(95..112)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(266..854)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(346..365)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1037..1131)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(1143..1934,1935..2003))
                     /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     1766..1785
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2004..2108)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2150..2168
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(2172..2215)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     protein_bind    complement(2235..2282)
                     /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)."
     promoter        2303..2358
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    2309..2327
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    2339..2357
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             2366..3571
                     /codon_start=1
                     /gene="tetA from transposon Tn10"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MNSSTKIALVITLLDAMGIGLIMPVLPTLLREFIASEDIANHFGV
                     LLALYALMQVIFAPWLGKMSDRFGRRPVLLLSLIGASLDYLLLAFSSALWMLYLGRLLS
                     GITGATGAVAASVIADTTSASQRVKWFGWLGASFGLGLIAGPIIGGFAGEISPHSPFFI
                     AALLNIVTFLVVMFWFRETKNTRDNTDTEVGVETQSNSVYITLFKTMPILLIIYFSAQL
                     IGQIPATVWVLFTENRFGWNSMMVGFSLAGLGLLHSVFQAFVAGRIATKWGEKTAVLLG
                     FIADSSAFAFLAFISEGWLVFPVLILLAGGGIALPALQGVMSIQTKSHQQGALQGLLVS
                     LTNATGVIGPLLFAVIYNHSLPIWDGWIWIIGLAFYCIIILLSMTFMLTPQAQGSKQET
                     SA"
     terminator      3604..3631
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    complement(3634..3667)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
ORIGIN
        1 tgagaccctg cagtccggca aaaaagggca aggtgtcacc accctgccct ttttctttaa
       61 aaccgaaaag attacttcgc gttatgcagg cttcctcgct cactgactcg ctgcgctcgg
      121 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
      181 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
      241 gtaaaaaggc cgcgttgctg gcgtttttcc acaggctccg cccccctgac gagcatcaca
      301 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
      361 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
      421 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
      481 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
      541 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
      601 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
      661 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
      721 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
      781 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
      841 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
      901 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
      961 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     1021 acagctcgag gcttggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga
     1081 acaaatccag atggagttct gaggtcatta ctggatctat caacaggagt ccaagcgagc
     1141 tcttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     1201 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     1261 cccagtgctg caatgatacc gcgagatcca cgctcaccgg ctccagattt atcagcaata
     1321 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     1381 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     1441 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     1501 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     1561 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     1621 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     1681 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     1741 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     1801 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     1861 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     1921 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     1981 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     2041 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     2101 gttccgcgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     2161 cacgaggcag aatttcagat aaaaaaaatc cttagctttc gctaaggatg atttctggaa
     2221 ttcggtctct ggaggaagtt cctatacttt ctagagaata ggaacttcgg aataggaact
     2281 tctagaactt atcctaattt ttgttgacac tctatcattg atagagttat tttaccactc
     2341 cctatcagtg atagagaaaa gtgaaatgaa tagttcgaca aagatcgcat tggtaattac
     2401 gttactcgat gccatgggga ttggccttat catgccagtc ttgccaacgt tattacgtga
     2461 atttattgct tcggaagata tcgctaacca ctttggcgta ttgcttgcac tttatgcgtt
     2521 aatgcaggtt atctttgctc cttggcttgg aaaaatgtct gaccgatttg gtcggcgccc
     2581 agtgctgttg ttgtcattaa taggcgcatc gctggattac ttattgctgg ctttttcaag
     2641 tgcgctttgg atgctgtatt taggccgttt gctttcaggg atcacaggag ctactggggc
     2701 tgtcgcggca tcggtcattg ccgataccac ctcagcttct caacgcgtga agtggttcgg
     2761 ttggttaggg gcaagttttg ggcttggttt aatagcgggg cctattattg gtggttttgc
     2821 aggagagatt tcaccgcata gtcccttttt tatcgctgcg ttgctaaata ttgtcacttt
     2881 ccttgtggtt atgttttggt tccgtgaaac caaaaataca cgtgataata cagataccga
     2941 agtaggggtt gagacgcaat cgaattcggt atacatcact ttatttaaaa cgatgcccat
     3001 tttgttgatt atttattttt cagcgcaatt gataggccaa attcccgcaa cggtgtgggt
     3061 gctatttacc gaaaatcgtt ttggatggaa tagcatgatg gttggctttt cattagcggg
     3121 tcttggtctt ttacactcag tattccaagc ctttgtggca ggaagaatag ccactaaatg
     3181 gggcgaaaaa acggcagtac tgctcggatt tattgcagat agtagtgcat ttgccttttt
     3241 agcgtttata tctgaaggtt ggttagtttt ccctgtttta attttattgg ctggtggtgg
     3301 gatcgcttta cctgcattac agggagtgat gtctatccaa acaaagagtc atcagcaagg
     3361 tgctttacag ggattattgg tgagccttac caatgcaacc ggtgttattg gcccattact
     3421 gtttgctgtt atttataatc attcactacc aatttgggat ggctggattt ggattattgg
     3481 tttagcgttt tactgtatta ttatcctgct atcgatgacc ttcatgttaa cccctcaagc
     3541 tcaggggagt aaacaggaga caagtgctta gtaagcggga ctctggggtt cgaaatgacc
     3601 gacggctcac cttcgggtgg gcctttctgc gaggaagttc ctatactttc tagagaatag
     3661 gaacttccgc t
//
