LOCUS       sequence-227880-        3334 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ORF insert for replicon MoClo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3334)
  AUTHORS   Wagner TE, Becraft JR, Bodner K, Teague B, Zhang X, Woo A, Porter E,
            Alburquerque B, Dobosh B, Andries O, Sanders NN, Beal J, Densmore D,
            Kitada T, Weiss R
  TITLE     Small-molecule-based regulation of RNA-delivered circuits in 
            mammalian cells.
  JOURNAL   Nat Chem Biol. 2018 Nov;14(11):1043-1050. doi: 
            10.1038/s41589-018-0146-9. Epub 2018 Oct 16.
  PUBMED    30327560
REFERENCE   2  (bases 1 to 3334)
  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: 
            "10.1038/s41589-018-0146-9"; journalName: "Nat Chem Biol"; date: 
            "2018-11"; volume: "14"; issue: "11"; pages: "1043-1050"
COMMENT     Sequence Label: pTW120
FEATURES             Location/Qualifiers
     source          1..3334
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(146..469)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    396..452
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(465..481)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(465..481)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(478..500)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    489..505
                     /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(513..519,520..537,538..543))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    558..579
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1344..1361)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1515..2103)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1595..1614)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2274..3065,3066..3134))
                     /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     2897..2916
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3135..3239)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3307..3325
                     /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 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acccggggat
      421 cctctagagt cgacctgcag gcatgcaagc ttggcgtaat catggtcata gctgtttcct
      481 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
      541 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
      601 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
      661 agaggcggtt tgcgtattgg gcgctcttcc gctggtctca caccatgtca agactcgaca
      721 agagcaaggt gattaacagt gcactggaac ttctcaatga agttgggatc gaggggctga
      781 ctactagaaa actcgcacag aaactggggg ttgagcagcc caccttgtac tggcacgtta
      841 aaaacaaaag ggccctgctg gatgctctgg ccatcgagat gctggatagg catcataccc
      901 acttctgccc tctggaagga gaatcctggc aggatttcct tagaaacaac gccaagtcct
      961 ttcgctgtgc ccttcttagc caccgggatg gtgctaaagt ccatctcggc acacgaccaa
     1021 ctgagaagca gtacgaaact ctcgagaacc agctggcctt tctctgtcaa cagggctttt
     1081 ctcttgaaaa cgccctgtac gcactgagtg cagttgggca ctttacactc ggatgtgttc
     1141 tggaggacca agaacatcag gtggcaaagg aagagaggga gacacctacg actgactcca
     1201 tgccccctct cttgaggcag gcaatagaat tgttcgacca tcagggcgca gaacccgcct
     1261 ttctgtttgg gctggaactg attatctgcg gtcttgagaa acagctgaag tgcgagtccg
     1321 ggagctaata atgagaccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     1381 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     1441 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     1501 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     1561 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     1621 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     1681 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     1741 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     1801 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     1861 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     1921 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     1981 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     2041 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     2101 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     2161 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     2221 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     2281 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     2341 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     2401 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     2461 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     2521 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     2581 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     2641 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     2701 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     2761 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     2821 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     2881 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     2941 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     3001 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     3061 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     3121 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     3181 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     3241 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc
     3301 tataaaaata ggcgtatcac gaggcccttt cgtc
//
