LOCUS       sequence-308791-        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      1..58
                     /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(82..99)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(253..841)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(333..352)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1024..1118)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(1130..1921,1922..1990))
                     /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     1753..1772
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1991..2095)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2137..2155
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(2159..2202)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     protein_bind    complement(2222..2269)
                     /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        2290..2345
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    2296..2314
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    2326..2344
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             2353..3558
                     /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      3591..3618
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    complement(3621..3654)
                     /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 tccggcaaaa aagggcaagg tgtcaccacc ctgccctttt tctttaaaac cgaaaagatt
       61 acttcgcgtt atgcaggctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
      121 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
      181 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
      241 gttgctggcg tttttccaca ggctccgccc ccctgacgag catcacaaaa atcgacgctc
      301 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
      361 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
      421 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
      481 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
      541 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
      601 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
      661 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
      721 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
      781 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
      841 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
      901 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
      961 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gctcgaggct
     1021 tggattctca ccaataaaaa acgcccggcg gcaaccgagc gttctgaaca aatccagatg
     1081 gagttctgag gtcattactg gatctatcaa caggagtcca agcgagctct taccaatgct
     1141 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     1201 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     1261 tgataccgcg agatccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     1321 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     1381 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     1441 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     1501 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     1561 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     1621 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     1681 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     1741 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     1801 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     1861 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     1921 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     1981 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2041 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgacgtc
     2101 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcagaat
     2161 ttcagataaa aaaaatcctt agctttcgct aaggatgatt tctggaattc ggtctctgga
     2221 ggaagttcct atactttcta gagaatagga acttcggaat aggaacttct agaacttatc
     2281 ctaatttttg ttgaaactct atcattgata gagttatttt accactccct atcagtgata
     2341 gagaaaagtg aaatgaatag ttcgacaaag atcgcattgg taattacgtt actcgatgcc
     2401 atggggattg gccttatcat gccagtcttg ccaacgttat tacgtgaatt tattgcttcg
     2461 gaagatatcg ctaaccactt tggcgtattg cttgcacttt atgcgttaat gcaggttatc
     2521 tttgctcctt ggcttggaaa aatgtctgac cgatttggtc ggcgcccagt gctgttgttg
     2581 tcattaatag gcgcatcgct ggattactta ttgctggctt tttcaagtgc gctttggatg
     2641 ctgtatttag gccgtttgct ttcagggatc acaggagcta ctggggctgt cgcggcatcg
     2701 gtcattgccg ataccacctc agcttctcaa cgcgtgaagt ggttcggttg gttaggggca
     2761 agttttgggc ttggtttaat agcggggcct attattggtg gttttgcagg agagatttca
     2821 ccgcatagtc ccttttttat cgctgcgttg ctaaatattg tcactttcct tgtggttatg
     2881 ttttggttcc gtgaaaccaa aaatacacgt gataatacag ataccgaagt aggggttgag
     2941 acgcaatcga attcggtata catcacttta tttaaaacga tgcccatttt gttgattatt
     3001 tatttttcag cgcaattgat aggccaaatt cccgcaacgg tgtgggtgct atttaccgaa
     3061 aatcgttttg gatggaatag catgatggtt ggcttttcat tagcgggtct tggtctttta
     3121 cactcagtat tccaagcctt tgtggcagga agaatagcca ctaaatgggg cgaaaaaacg
     3181 gcagtactgc tcggatttat tgcagatagt agtgcatttg cctttttagc gtttatatct
     3241 gaaggttggt tagttttccc tgttttaatt ttattggctg gtggtgggat cgctttacct
     3301 gcattacagg gagtgatgtc tatccaaaca aagagtcatc agcaaggtgc tttacaggga
     3361 ttattggtga gccttaccaa tgcaaccggt gttattggcc cattactgtt tgctgttatt
     3421 tataatcatt cactaccaat ttgggatggc tggatttgga ttattggttt agcgttttac
     3481 tgtattatta tcctgctatc gatgaccttc atgttaaccc ctcaagctca ggggagtaaa
     3541 caggagacaa gtgcttagta agcgggactc tggggttcga aatgaccgac ggctcacctt
     3601 cgggtgggcc tttctgcgag gaagttccta tactttctag agaataggaa cttccgcttg
     3661 agaccctgca g
//
