LOCUS       sequence-126622-        8662 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TetR/PhlF/SrpR/LambdaCI/LacI BCD7 5-node v41-2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8662)
  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 8662)
  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: 5n1 (560)
FEATURES             Location/Qualifiers
     source          1..8662
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1213..1923
                     /codon_start=1
                     /gene="cI"
                     /product="phage lambda repressor"
                     /label=lambda repressor
                     /translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ
                     SGVGALFNGINALNAYNAALLAKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY
                     PVFSHVQAGMFSPELRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD
                     GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV
                     VGKVIASQWPEETFG"
     CDS             3079..4158
                     /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="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3098..3117)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4314..4330
                     /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             4435..5055
                     /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(4508..4527)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      5250..5321
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5337..5364
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(5895..6845)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     terminator      complement(7607..7701)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(7732..8526)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(7843..7862)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     8453..8472
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 gacgtcggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
       61 ttccagtcgg gaaacctggt ctcaaagccc acgcattgct gttcgattcg ttaccaattg
      121 acatgatacg aaacgtaccg tatcgttaag gttactagag aagcgggccc aagttcactt
      181 aaaaaggaga tcaacaatga aagcaatttt cgtactgaaa catcttaatc atgctgggga
      241 gggtttctaa tggcacgtaa aaccgcagca gaagcagaag aaacccgtca gcgtattatt
      301 gatgcagcac tggaagtttt tgttgcacag ggtgttagtg atgcaaccct ggatcagatt
      361 gcacgtaaag ccggtgttac ccgtggtgca gtttattggc attttaatgg taaactggaa
      421 gttctgcagg cagttctggc aagccgtcag catccgctgg aactggattt tacaccggat
      481 ctgggtattg aacgtagctg ggaagcagtt gttgttgcaa tgctggatgc agttcatagt
      541 ccgcagagca aacagtttag cgaaattctg atttatcagg gtctggatga aagcggtctg
      601 attcataatc gtatggttca ggcaagcgat cgttttctgc agtatattca tcaggttctg
      661 cgtcatgcag ttacccaggg tgaactgccg attaatctgg atctgcagac cagcattggt
      721 gtttttaaag gtctgattac cggtctgctg tatgaaggtc tgcgtagcaa agatcagcag
      781 gcacagatta tcaaagttgc actgggtagc ttttgggcac tgctgcgtga accgcctcgt
      841 tttctgctgt gtgaagaagc acagattaaa caggtgaaat ccttcgaagc agcaaacgac
      901 gaaaactacg ctttagcagc ttaatgaact ttcagccaaa aaacttaaga ccgccggtct
      961 tgtccactac cttgcagtaa tgcggtggac aggatcggcg gttttctttt ctcttctcaa
     1021 ccttgagagt cgggcagcat tgctgttcga ttcgttacca attgacagct agctcagtcc
     1081 taggtatata catacatgct tgtttgtttg taaactacta gagaagcggg cccaagttca
     1141 cttaaaaagg agatcaacaa tgaaagcaat tttcgtactg aaacatctta atcatgctgg
     1201 ggagggtttc taatgagcac aaaaaagaaa ccattaacac aagagcagct tgaggacgca
     1261 cgtcgcctta aagcaattta tgaaaaaaag aaaaatgaac ttggcttatc ccaggaatct
     1321 gtcgcagaca agatggggat ggggcagtca ggcgttggtg ctttatttaa tggcatcaat
     1381 gcattaaatg cttataacgc cgcattgctt gcaaaaattc tcaaagttag cgttgaagaa
     1441 tttagccctt caatcgccag agaaatctac gagatgtatg aagcggttag tatgcagccg
     1501 tcacttagaa gtgagtatga gtaccctgtt ttttctcatg ttcaggcagg gatgttctca
     1561 cctgagctta gaacctttac caaaggtgat gcggagagat gggtaagcac aaccaaaaaa
     1621 gccagtgatt ctgcattctg gcttgaggtt gaaggtaatt ccatgaccgc accaacaggc
     1681 tccaagccaa gctttcctga cggaatgtta attctcgttg accctgagca ggctgttgag
     1741 ccaggtgatt tctgcatagc cagacttggg ggtgatgagt ttaccttcaa gaaactgatc
     1801 agggatagcg gtcaggtgtt tttacaacca ctaaacccac agtacccaat gatcccatgc
     1861 aatgagagtt gttccgttgt ggggaaagtt atcgctagtc agtggcctga agagacgttt
     1921 ggcgcagcaa acgacgaaaa ctacgcttta gcagcttgat gaactaacgc atgagaaagc
     1981 ccccggaaga tcaccttccg ggggcttttt tattgcgccc ttgagagtcg atgtgcattg
     2041 ctgttctcag tgatagagat tgacatccct atcagtgata gatataatga gcactactag
     2101 agaagcgggc ccaagttcac ttaaaaagga gatcaacaat gaaagcaatt ttcgtactga
     2161 aacatcttaa tcatgctggg gagggtttct aatggcacgt accccgagcc gtagcagcat
     2221 tggtagcctg cgtagtccgc atacccataa agcaattctg accagcacca ttgaaatcct
     2281 gaaagaatgt ggttatagcg gtctgagcat tgaaagcgtt gcacgtcgtg ccggtgcaag
     2341 caaaccgacc atttatcgtt ggtggaccaa taaagcagca ctgattgccg aagtgtatga
     2401 aaatgaaagc gaacaggtgc gtaaatttcc ggatctgggt agctttaaag ccgatctgga
     2461 ttttctgctg cgtaatctgt ggaaagtttg gcgtgaaacc atttgtggtg aagcatttcg
     2521 ttgtgttatt gcagaagcac agctggaccc tgcaaccctg acccagctga aagatcagtt
     2581 tatggaacgt cgtcgtgaga tgccgaaaaa actggttgaa aatgccatta gcaatggtga
     2641 actgccgaaa gataccaatc gtgaactgct gctggatatg atttttggtt tttgttggta
     2701 tcgcctgctg accgaacagc tgaccgttga acaggatatt gaagaattta ccttcctgct
     2761 gattaatggt gtttgtccgg gtacacagcg tgcagcaaac gacgaaaact acgctttagc
     2821 agcttaatga acttccggca attaaaaaag cggctaacca cgccgctttt tttacgtctg
     2881 caccttgaga gtcgctcggc attgctgttc ctgcaggtcg aggataaata tctaacaccg
     2941 tgcgtgttga ctattttacc tctggcggtg ataatggttg catgtactaa agcgggccca
     3001 agttcactta aaaaggagat caacaatgaa agcaattttc gtactgaaac atcttaatca
     3061 tgctggggag ggtttctaat gaaaccagta acgttatacg atgtcgcaga gtatgccggt
     3121 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
     3181 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
     3241 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
     3301 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
     3361 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
     3421 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
     3481 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca
     3541 cccatcaaca gtattatttt ctcccatgag gacggtacgc gactgggcgt ggagcatctg
     3601 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
     3661 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
     3721 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
     3781 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
     3841 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
     3901 gataccgagg acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc
     3961 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     4021 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     4081 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     4141 cgactggaaa gcgggcaggc agcaaacgac gaaaactacg ctttagcagc ttgatgaact
     4201 gttatgagtc aggaaaaaag gcgacagagt aatctgtcgc cttttttctt tgcttgcttt
     4261 ccttgagagt cgaaccgcat tgctttcata aatgtgagcg gataacattg acattgtgag
     4321 cggataacaa gatactgagc acagcaagcg ggcccaagtt cacttaaaaa ggagatcaac
     4381 aatgaaagca attttcgtac tgaaacatct taatcatgct ggggagggtt tctaatgtct
     4441 agattagata aaagtaaagt gattaacagc gcattagagc tgcttaatga ggtcggaatc
     4501 gaaggtttaa caacccgtaa actcgcccag aagctaggtg tagagcagcc tacattgtat
     4561 tggcatgtaa aaaataagcg ggctttgctc gacgccttag ccattgagat gttagatagg
     4621 caccatactc acttttgccc tttagaaggg gaaagctggc aagatttttt acgtaataac
     4681 gctaaaagtt ttagatgtgc tttactaagt catcgcgatg gagcaaaagt acatttaggt
     4741 acacggccta cagaaaaaca gtatgaaact ctcgaaaatc aattagcctt tttatgccaa
     4801 caaggttttt cactagagaa tgcattatat gcactcagcg ctgtggggca ttttacttta
     4861 ggttgcgtat tggaagatca agagcatcaa gtcgctaaag aagaaaggga aacacctact
     4921 actgatagta tgccgccatt attacgacaa gctatcgaat tatttgatca ccaaggtgca
     4981 gagccagcct tcttattcgg ccttgaattg atcatatgcg gattagaaaa acaacttaaa
     5041 tgtgaaagtg ggtctgcagc aaacgacgaa aactacgctt tagcagctta atgaactttc
     5101 agccaaaaaa cttaagaccg ccggtcttgt ccactacctt gcagtaatgc ggtggacagg
     5161 atcggcggtt ttcttttctc ttctcaacct tgagagtcgg cttgtcgtga gacctaagga
     5221 tccaaactcg agtaaggatc tccaggcatc aaataaaacg aaaggctcag tcgaaagact
     5281 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct ctactagagt cacactggct
     5341 caccttcggg tgggcctttc tgcgtttata cctagggtac gggttttgct gcccgcaaac
     5401 gggctgttct ggtgttgcta gtttgttatc agaatcgcag atccggcttc agccggtttg
     5461 ccggctgaaa gcgctatttc ttccagaatt gccatgattt tttccccacg ggaggcgtca
     5521 ctggctcccg tgttgtcggc agctttgatt cgataagcag catcgcctgt ttcaggctgt
     5581 ctatgtgtga ctgttgagct gtaacaagtt gtctcaggtg ttcaatttca tgttctagtt
     5641 gctttgtttt actggtttca cctgttctat taggtgttac atgctgttca tctgttacat
     5701 tgtcgatctg ttcatggtga acagctttga atgcaccaaa aactcgtaaa agctctgatg
     5761 tatctatctt ttttacaccg ttttcatctg tgcatatgga cagttttccc tttgatatgt
     5821 aacggtgaac agttgttcta cttttgtttg ttagtcttga tgcttcactg atagatacaa
     5881 gagccataag aacctcagat ccttccgtat ttagccagta tgttctctag tgtggttcgt
     5941 tgtttttgcg tgagccatga gaacgaacca ttgagatcat acttactttg catgtcactc
     6001 aaaaattttg cctcaaaact ggtgagctga atttttgcag ttaaagcatc gtgtagtgtt
     6061 tttcttagtc cgttatgtag gtaggaatct gatgtaatgg ttgttggtat tttgtcacca
     6121 ttcattttta tctggttgtt ctcaagttcg gttacgagat ccatttgtct atctagttca
     6181 acttggaaaa tcaacgtatc agtcgggcgg cctcgcttat caaccaccaa tttcatattg
     6241 ctgtaagtgt ttaaatcttt acttattggt ttcaaaaccc attggttaag ccttttaaac
     6301 tcatggtagt tattttcaag cattaacatg aacttaaatt catcaaggct aatctctata
     6361 tttgccttgt gagttttctt ttgtgttagt tcttttaata accactcata aatcctcata
     6421 gagtatttgt tttcaaaaga cttaacatgt tccagattat attttatgaa tttttttaac
     6481 tggaaaagat aaggcaatat ctcttcacta aaaactaatt ctaatttttc gcttgagaac
     6541 ttggcatagt ttgtccactg gaaaatctca aagcctttaa ccaaaggatt cctgatttcc
     6601 acagttctcg tcatcagctc tctggttgct ttagctaata caccataagc attttcccta
     6661 ctgatgttca tcatctgagc gtattggtta taagtgaacg ataccgtccg ttctttcctt
     6721 gtagggtttt caatcgtggg gttgagtagt gccacacagc ataaaattag cttggtttca
     6781 tgctccgtta agtcatagcg actaatcgct agttcatttg ctttgaaaac aactaattca
     6841 gacatacatc tcaattggtc taggtgattt taatcactat accaattgag atgggctagt
     6901 caatgataat tactagtcct tttcccgggt gatctgggta tctgtaaatt ctgctagacc
     6961 tttgctggaa aacttgtaaa ttctgctaga ccctctgtaa attccgctag acctttgtgt
     7021 gttttttttg tttatattca agtggttata atttatagaa taaagaaaga ataaaaaaag
     7081 ataaaaagaa tagatcccag ccctgtgtat aactcactac tttagtcagt tccgcagtat
     7141 tacaaaagga tgtcgcaaac gctgtttgct cctctacaaa acagacctta aaaccctaaa
     7201 ggcttaagta gcaccctcgc aagctcgggc aaatcgctga atattccttt tgtctccgac
     7261 catcaggcac ctgagtcgct gtctttttcg tgacattcag ttcgctgcgc tcacggctct
     7321 ggcagtgaat gggggtaaat ggcactacag gcgcctttta tggattcatg caaggaaact
     7381 acccataata caagaaaagc ccgtcacggg cttctcaggg cgttttatgg cgggtctgct
     7441 atgtggtgct atctgacttt ttgctgttca gcagttcctg ccctctgatt ttccagtctg
     7501 accacttcgg attatcccgt gacaggtcat tcagactggc taatgcaccc agtaaggcag
     7561 cggtatcatc aacaggctta cccgtcttac tgtccctagt gcttggattc tcaccaataa
     7621 aaaacgcccg gcggcaaccg agcgttctga acaaatccag atggagttct gaggtcatta
     7681 ctggatctat caacaggagt ccaagcgagc tctcgaaccc cagagtcccg ctcagaagaa
     7741 ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga taccgtaaag
     7801 cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac gggtagccaa
     7861 cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga atccagaaaa
     7921 gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca cgacgagatc
     7981 ctcgccgtcg ggcatgcgcg ccttgagcct ggcgaacagt tcggctggcg cgagcccctg
     8041 atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag tacgtgctcg
     8101 ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa gcgtatgcag
     8161 ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt gagatgacag
     8221 gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt cagtgacaac
     8281 gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc gcgctgcctc
     8341 gtcctgcagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa ccgggcgccc
     8401 ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt gtgcccagtc
     8461 atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc catcttgttc
     8521 aatcatgcga aacgatcctc atcctgtctc ttgatcagat catgatcccc tgcgccatca
     8581 gatccttggc ggcaagaaag ccatccagtt tactttgcag ggcttcccaa ccttaccaga
     8641 gggcgcccca gctggcaatt cc
//
