LOCUS       sequence-333685-        4258 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual control based multi-level gene expression controller (MLC).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4258)
  AUTHORS   Greco FV, Pandi A, Erb TJ, Grierson CS, Gorochowski TE
  TITLE     Harnessing the central dogma for stringent multi-level control of 
            gene expression.
  JOURNAL   Nat Commun. 2021 Mar 19;12(1):1738. doi: 10.1038/s41467-021-21995-7.
  PUBMED    33741937
REFERENCE   2  (bases 1 to 4258)
  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: "Nat 
            Commun."; date: "2021-03-19"; volume: "12(1):1738. doi"; pages: " 
            10.1038/s41467-021-21995-7"
COMMENT     Sequence Label: pDC
FEATURES             Location/Qualifiers
     source          1..4258
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(57..1139)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYMTVSRVVNQASHVSAKTREKVEAAMVEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIYSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1101..1120
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1278..1283,1284..1299,1300..1306)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1312..1331
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     terminator      1378..1464
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1556..1583
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     protein_bind    2491..2510
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     promoter        complement(join(2516..2522,2523..2538,2539..2544))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     rep_origin      complement(2663..3251)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2743..2762)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(3362..3381)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3373..4188
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             3373..4188
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3446..3465)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 agtgaaaaga aaaaaggccg cagagcggcc tttttagtta gatcggaccc cacatctcac
       61 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
      121 cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagact
      181 ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg
      241 ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat
      301 gagctatctt cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg
      361 gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca
      421 gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc
      481 cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag
      541 ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc
      601 tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcctc atgggagtaa
      661 ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg
      721 caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca
      781 ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt
      841 tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg
      901 acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac
      961 tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc caccatcgcc
     1021 gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa
     1081 acggtcatat aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcata
     1141 ttcacctcca tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttt
     1201 caccattcga tggcgcgccg cggctgggaa tctgagacat gagtcaagat atttgctcgg
     1261 taacgtatgc tggagtgttg acaattaatc atcggctcgt ataatgtgtg gaattgtgag
     1321 cgctcacaat taggtaacaa aataaagcaa taaggaatcg ctcacccaaa ggatctgctt
     1381 ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg
     1441 aacgctctcc tgagtaggac aaatccgccg ggagcggatt tgaacgttgc gaagcaacgg
     1501 cccggagggt ggcgggcagg acgcccgcca taaactgcca ggcatcaaat taagcagaag
     1561 gccatcctga cggatggcct tttggccccc gacccttaga ctctgtactc agggaccaaa
     1621 acgaaaaaag gccccccttt cgggaggcct cttttctgga atttggtacc gagcgctagt
     1681 tgtatcgtac gtcggtctag gaatcgttta gccgcctcac ttatacaact catccatgcc
     1741 taaagtaatc cccgccgcgg ttacgaactc caaaaggacc atgtgatcac gtttctcatt
     1801 tgggtccttc gataatttgg attgtgtact caggtagtga ttgtcaggaa gtaaaaccgg
     1861 gccgtcgcca atgggtgtgt tttgttgata atggtcagcc aattgcacgc tgccatcctc
     1921 aatgttgtga cgaatcttga aatttacctt aatcccgttc ttctgcttgt cagccatgat
     1981 atacacgttg tgcgaattat agttatactc cagtttatgt cccaaaatgt tgccatcttc
     2041 tttaaagtca atgcctttca attcaatgcg attcactaac gtatccccct caaacttgac
     2101 ctcggcgcgt gtcttatagt tcccgtcatc cttgaagaag attgtgcgct cttgcacata
     2161 gccttctggc attgcagatt taaagaagtc gtgctgcttc atgtgatctg ggtaacgcga
     2221 gaaacattgg accccgtatg taagtgttgt aaccagagtt ggccagggga ctggaagttt
     2281 tcctgtagta caaatgaact taagcgtcaa tttgccgtaa gtagcgtcgc cttcgccctc
     2341 gccgctaacc gaaaacttat gaccattaac gtccccatcc aactcaacca agatgggtac
     2401 aacgcctgta aacagctcct ctcctttact caccatcaaa aaatcgactc cttaatctca
     2461 atttcgttta aggaatcgct cacccatact aattgtgagc gctcacaatt ccacacatta
     2521 tacgagccga tgattaattg tcaacatgcc caaaggcggt aatacggtta tccacagaat
     2581 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2641 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2701 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2761 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2821 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2881 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2941 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3001 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3061 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     3121 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     3181 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3241 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     3301 actcacgtta agggattttg gtcatgaaca ataaaactgt ctgcttacat aaacagtaat
     3361 acaaggggtg ttatgagcca tattcaacgg gaaacgtctt gctctaggcc gcgattaaat
     3421 tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt cgggcaatca
     3481 ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc cagagttgtt tctgaaacat
     3541 ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa ctggctgacg
     3601 gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga tgcatggtta
     3661 ctcaccactg cgatccccgg gaaaacagca ttccaggtat tagaagaata tcctgattca
     3721 ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt
     3781 tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg
     3841 aataacggtt tggttgatgc gagtgatttt gatgacgagc gtaatggctg gcctgttgaa
     3901 caagtctgga aagaaatgca taaacttttg ccattctcac cggattcagt cgtcactcat
     3961 ggtgatttct cacttgataa ccttattttt gacgagggga aattaatagg ttgtattgat
     4021 gttggacgag tcggaatcgc agaccgatac caggatcttg ccatcctatg gaactgcctc
     4081 ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa aatatggtat tgataatcct
     4141 gatatgaata aattgcagtt tcatttgatg ctcgatgagt ttttctaaga attaattcat
     4201 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcggtga
//
