LOCUS       sequence-149167-        3267 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  plasmid for building 5n1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3267)
  AUTHORS   Niederholtmeyer H, Sun ZZ, Hori Y, Yeung E, Verpoorte A, Murray RM, 
            Maerkl SJ
  TITLE     Rapid cell-free forward engineering of novel genetic ring 
            oscillators.
  JOURNAL   Elife. 2015 Oct 2;4. doi: 10.7554/eLife.09771.
  PUBMED    26430766
REFERENCE   2  (bases 1 to 3267)
  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: "Elife."; 
            date: "2015-10-2"; volume: "4. doi"; pages: " 10.7554/eLife.09771"
COMMENT     Sequence Label: pLacO1-BCD7-tetR-ssrA P42U18C45T13 v40-5a (497)
FEATURES             Location/Qualifiers
     source          1..3267
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    38..54
                     /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)."
     CDS             159..779
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(232..251)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(1040..1057)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1211..1799)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1291..1310)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1970..2761,2762..2830))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHATRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2593..2612
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2831..2935)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3003..3021
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcattgcttt cataaatgtg agcggataac attgacattg tgagcggata acaagatact
       61 gagcacagca agcgggccca agttcactta aaaaggagat caacaatgaa agcaattttc
      121 gtactgaaac atcttaatca tgctggggag ggtttctaat gtctagatta gataaaagta
      181 aagtgattaa cagcgcatta gagctgctta atgaggtcgg aatcgaaggt ttaacaaccc
      241 gtaaactcgc ccagaagcta ggtgtagagc agcctacatt gtattggcat gtaaaaaata
      301 agcgggcttt gctcgacgcc ttagccattg agatgttaga taggcaccat actcactttt
      361 gccctttaga aggggaaagc tggcaagatt ttttacgtaa taacgctaaa agttttagat
      421 gtgctttact aagtcatcgc gatggagcaa aagtacattt aggtacacgg cctacagaaa
      481 aacagtatga aactctcgaa aatcaattag cctttttatg ccaacaaggt ttttcactag
      541 agaatgcatt atatgcactc agcgctgtgg ggcattttac tttaggttgc gtattggaag
      601 atcaagagca tcaagtcgct aaagaagaaa gggaaacacc tactactgat agtatgccgc
      661 cattattacg acaagctatc gaattatttg atcaccaagg tgcagagcca gccttcttat
      721 tcggccttga attgatcata tgcggattag aaaaacaact taaatgtgaa agtgggtctg
      781 cagcaaacga cgaaaactac gctttagcag cttaatgaac tttcagccaa aaaacttaag
      841 accgccggtc ttgtccacta ccttgcagta atgcggtgga caggatcggc ggttttcttt
      901 tctcttctca accttgagag tcggcttttg tcttcgctcc ttccggtggg cgcggggcat
      961 gactatcgtc gccgcactta tgactgtgtt ctttatcatg caactcgtag gacaggtgcc
     1021 ggcagcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     1081 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     1141 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     1201 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1261 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     1321 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     1381 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     1441 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     1501 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     1561 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     1621 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     1681 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     1741 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     1801 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     1861 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     1921 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     1981 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     2041 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2101 tgataccgcg tgacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     2161 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     2221 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     2281 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     2341 cccaacgatc aaggcgagtt gcatgatccc ccatgttgtg caaaaaagcg gttagctcct
     2401 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     2461 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     2521 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     2581 cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     2641 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     2701 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     2761 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     2821 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2881 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     2941 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     3001 aaaataggcg tatcacgagg ccctttcgtg ttcaagaatt ctggcgaatc ctctgaccag
     3061 ccagaaaacg acctttctgt ggtgaaaccg gatgctgcaa ttcagagcgc cagcaagtgg
     3121 gggacagcag atgacctgac cgccgcagag tggatgtttg acatggtgat gactatcgca
     3181 ccatcagcca gaaaaccgaa ttttgctggg tgggctaacg atatccgcct gatgcgtgaa
     3241 cgtgacggac gtaacgaaga caaaacc
//
