LOCUS       sequence-204413-        3040 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MIDAS Level-3 destination vector. Used for initiating Level-3 
            multigene assembly..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3040)
  AUTHORS   van Dolleweerd CJ, Kessans SA, Van de Bittner KC, Bustamante LY, 
            Bundela R, Scott B, Nicholson MJ, Parker EJ
  TITLE     MIDAS: A Modular DNA Assembly System for Synthetic Biology.
  JOURNAL   ACS Synth Biol. 2018 Apr 20;7(4):1018-1029. doi: 
            10.1021/acssynbio.7b00363. Epub 2018 Apr 12.
  PUBMED    29620866
REFERENCE   2  (bases 1 to 3040)
  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"; doi: 
            "10.1021/acssynbio.7b00363"; journalName: "ACS Synth Biol"; date: 
            "2018-04-20- 20"; volume: "7"; issue: "4"; pages: "1018-1029"
COMMENT     Sequence Label: pML3
FEATURES             Location/Qualifiers
     source          1..3040
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(41..214)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGE"
     primer_bind     163..185
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     177..194
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     178..194
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(210..226)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(210..226)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(223..245)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    234..250
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(258..264,265..282,283..288))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    303..324
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(460..477)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(631..1219)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(711..730)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             1691..2482
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWHRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      complement(2574..3029)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2711..2732)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2923..2942
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gtgtcattgc ttgcaggtgt tattaccagg caaagcgcca ttcgccattc aggctgcgca
       61 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      121 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      181 aaacgacggc cagtgaatcc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
      241 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
      301 taatgagtga gctaactcac attaattgcg ttgcgccacc tgcccgccgt aggcatcact
      361 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
      421 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
      481 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
      541 cacagaatca ggggataact caggaaagaa catgtgagca aaaggccagc aaaaggccag
      601 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
      661 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
      721 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
      781 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
      841 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
      901 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
      961 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     1021 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     1081 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     1141 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     1201 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     1261 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     1321 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     1381 gtctgacaga ccggacgtcg atatccggat gaaggcacga acccagtgga cataagcctg
     1441 ttcggttcgt aagctgtaat gcaagtagcg tatgcgctca cgcaactggt ccagaacctt
     1501 gaccgaacgc agcggtggta acggcgcagt ggcggttttc atggcttgtt atgactgttt
     1561 ttttggggta cagtctatgc ctcgtgcatc caagcagcaa gcgcgttacg ccgtgggtcg
     1621 atgtttgatg ttatggagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa
     1681 gttaaacatc atgagggaag cggtgatcgc cgaagtatct actcaactat cagaggtagt
     1741 tggcgtcatc gagcgccatc tcgaaccgac gttgctggcc gtacatttgt acggctccgc
     1801 agtggatggc ggcctgaagc cacacagtga tattgatttg ctggttacgg tgaccgtaag
     1861 gcttgatgaa acaacgcggc gagctttgat caacgacctt ttggaaactt cggcttcccc
     1921 tggagagagc gagattctcc gcgctgtaga agtcaccatt gttgtgcacg acgacatcat
     1981 tccgtggcgt tatccagcta agcgcgaact gcaatttgga gaatggcatc gcaatgacat
     2041 tcttgcaggt atcttcgagc cagccacgat cgacattgat ctggctatct tgctgacaaa
     2101 agcaagagaa catagcgttg ccttggtagg tccagcggcg gaggaactct ttgatccggt
     2161 tcctgaacag gatctatttg aggcgctaaa tgaaacctta acgctatgga actcgccgcc
     2221 cgactgggct ggcgatgagc gaaatgtagt gcttacgttg tcccgcattt ggtacagcgc
     2281 agtaaccggc aaaatcgcgc cgaaggatgt cgctgccgac tgggcaatgg agcgcctgcc
     2341 ggcccagtat cagcccgtca tacttgaagc tagacaggct tatcttggac aagaagaaga
     2401 tcgcttggcc tcgcgcgcag atcagttgga agaatttgtc cactacgtga aaggcgagat
     2461 caccaaggta gtcggcaaat aagaattaat tcatgagcgg atacatattt gaatgtattt
     2521 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt
     2581 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     2641 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     2701 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaatgtcaa
     2761 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     2821 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     2881 tagatcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     2941 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     3001 cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca
//
