LOCUS       sequence-147384-        3718 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 3718)
  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 3718)
  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: pCI-BCD7-lacI-ssrA P62U18C50T16 v40-4a (496)
FEATURES             Location/Qualifiers
     source          1..3718
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             181..1260
                     /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(200..219)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     primer_bind     complement(1621..1639)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1707..1811
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1812..1880,1881..2672)
                     /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     complement(2030..2049)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2843..3431
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3332..3351
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3585..3602
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcattgctgt tcctgcaggt cgaggataaa tatctaacac cgtgcgtgtt gactatttta
       61 cctctggcgg tgataatggt tgcatgtact aaagcgggcc caagttcact taaaaaggag
      121 atcaacaatg aaagcaattt tcgtactgaa acatcttaat catgctgggg agggtttcta
      181 atgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt
      241 tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg
      301 gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag
      361 tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc
      421 gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa
      481 cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctagcgca acgcgtcagt
      541 gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc
      601 actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt
      661 ttctcccatg aggacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag
      721 caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc
      781 tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg
      841 agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact
      901 gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc
      961 gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga ggacagctca
     1021 tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc
     1081 gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc
     1141 gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc
     1201 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag
     1261 gcagcaaacg acgaaaacta cgctttagca gcttgatgaa ctgttatgag tcaggaaaaa
     1321 aggcgacaga gtaatctgtc gccttttttc tttgcttgct ttccttgaga gtcgcgagtt
     1381 gtcttcgtta cgtccgtcac gttcacgcat caggcggata tcgttagccc acccagcaaa
     1441 attcggtttt ctggctgatg gtgcgatagt catcaccatg tcaaacatcc actctgcggc
     1501 ggtcaggtca tctgctgtcc cccacttgct ggcgctctga attgcagcat ccggtttcac
     1561 cacagaaagg tcgttttctg gctggtcaga ggattcgcca gaattcttga acacgaaagg
     1621 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     1681 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     1741 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1801 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1861 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1921 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1981 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     2041 cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     2101 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     2161 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     2221 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     2281 caactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     2341 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     2401 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     2461 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     2521 agcgtgggtc acgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     2581 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     2641 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     2701 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     2761 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2821 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2881 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2941 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     3001 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     3061 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3121 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3181 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3241 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3301 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3361 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3421 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3481 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3541 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3601 aggaagcgga agagcgctgc cggcacctgt cctacgagtt gcatgataaa gaacacagtc
     3661 ataagtgcgg cgacgatagt catgccccgc gcccaccgga aggagcgaag acaaggtt
//
