LOCUS       sequence-272944-        2888 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SGP insert for replicon MoClo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2888)
  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 2888)
  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: pTW099
FEATURES             Location/Qualifiers
     source          1..2888
                     /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(898..915)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1069..1657)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1149..1168)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1828..2619,2620..2688))
                     /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     2451..2470
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2689..2793)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2861..2879
                     /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 gctggtctcc gacttccatc atagttatgg
      721 ccatgactac tctagctagc agtgttaaat cattcagcta cctgagaggg gcccctataa
      781 ctctctacgg ctaacctgaa tggactacga catgggcgtg atccgaaagg tgacccggat
      841 ctggggcgtg atccgaaagg tgacccggat ccaccggtcg ccacctgaga ccgcttcctc
      901 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
      961 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     1021 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     1081 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     1141 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     1201 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     1261 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     1321 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     1381 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     1441 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     1501 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     1561 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     1621 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     1681 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     1741 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     1801 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     1861 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     1921 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     1981 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     2041 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     2101 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     2161 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     2221 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     2281 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     2341 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     2401 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     2461 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     2521 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     2581 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     2641 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     2701 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     2761 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     2821 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     2881 ctttcgtc
//
