LOCUS       sequence-201411-        5351 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas1, Cas2 under PLTetO-1 promoter, and TetR and LacI 
            constitutively..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5351)
  AUTHORS   Sheth RU, Yim SS, Wu FL, Wang HH
  TITLE     Multiplex recording of cellular events over time on CRISPR 
            biological tape.
  JOURNAL   Science. 2017 Dec 15;358(6369):1457-1461. doi: 
            10.1126/science.aao0958. Epub 2017 Nov 23.
  PUBMED    29170279
REFERENCE   2  (bases 1 to 5351)
  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: 
            "10.1126/science.aao0958"; journalName: "Science"; date: 
            "2017-12-15- 15"; volume: "358"; issue: "6369"; pages: "1457-1461"
COMMENT     Sequence Label: pRec
FEATURES             Location/Qualifiers
     source          1..5351
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     143..161
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(182..200,201..206,207..223,224..229,230..255)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    182..200
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    207..225
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      1515..1586
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1602..1629
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        1681..1715
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     CDS             1750..2376
                     /codon_start=1
                     /gene="tetR"
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(1826..1845)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      2382..2468
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        2504..2581
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2582..3664
                     /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(2601..2620)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      complement(3875..4463)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3955..3974)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4551..4645)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4666..5325)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5240..5259
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 tcgataactc aaaaaatacg accggtagtg atcttatttc attatggtga aagttggaac
       61 ctcttacggg ccggccaacg tctcattttc gccagatatc gacgtctaag aaaccattat
      121 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc ttcacctcga
      181 gtccctatca gtgatagaga ttgacatccc tatcagtgat agagatactg agcacatcag
      241 caggacgcac tgaccgttta ataaatatat acatatcagg acaacgccta atgacctggc
      301 ttccccttaa tcccattcca ctcaaagatc gcgtctccat gatctttctg caatatgggc
      361 agatcgatgt aatagatggc gcgtttgtac ttatcgacaa gacagggatc cgcactcata
      421 ttcctgttgg ctcggttgcc tgcatcatgc tggaacctgg tacacgggtt tcgcatgcag
      481 ctgtacgcct ggctgcgcaa gttggaacat tgttggtatg ggtgggggaa gcgggcgttc
      541 gtgtttatgc ttctggtcag cctggaggtg cgcgttcaga taagctgctc tatcaggcaa
      601 aacttgctct ggatgaagat ttgcgtctga aggtcgtacg taaaatgttt gaacttcggt
      661 ttggagaacc tgcgcctgcc cggcgctccg tagagcaact cagaggtata gaaggcagtc
      721 gcgtgcgggc aacctacgca cttctggcga agcaatacgg cgtgacatgg aatggacgtc
      781 gctacgatcc gaaagactgg gaaaagggcg atacgatcaa ccaatgcatt agcgctgcaa
      841 cttcctgttt atacggcgta actgaagcgg cgatacttgc agctggttat gcaccagcta
      901 ttgggtttgt gcatacagga aagcctcttt cctttgttta cgatattgca gacatcatta
      961 aatttgacac tgttgtaccg aaagcttttg agatagcgcg tcgtaaccct ggtgagccgg
     1021 accgggaagt ccgtttggcg tgcagggata tttttcgcag tagtaaaaca ttagccaaat
     1081 tgattccgct tatagaggac gtgcttgccg ctggagaaat acaaccgccg gccccacctg
     1141 aagatgcaca gcctgttgcc attccgcttc ctgtttcact gggagatgca ggccatcgga
     1201 gtagctgaaa tgagtatgtt ggtcgtggtc actgaaaatg tacctccgcg cttacgaggc
     1261 agattagcca tctggttgtt ggaggtacgt gcaggggtat atgtaggtga tgtatccgca
     1321 aaaattcgtg aaatgatctg ggaacaaata gctggactgg cggaagaagg caatgtagtg
     1381 atggcatggg caacgaatac ggaaacggga tttgagttcc agacatttgg gttaaacagg
     1441 cgtaccccgg tagatttgga tggtttaagg ttggtgtctt ttttacctgt ttgataataa
     1501 tctagaccag gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta
     1561 tctgttgttt gtcggtgaac gctctctact agagtaacac tggctcacct tcgggtgggc
     1621 ctttctgcgt ttataggtac ccgtggatcc tctagagtcg acctgcaggc atgcaagctt
     1681 tttacggcta gctcagtcct aggtatagtg ctagcccagc cagagaaaca ctctttaaca
     1741 gggggtagta tgatgtctcg tttagataaa agtaaagtga ttaacagcgc attagagctg
     1801 cttaatgagg tcggaatcga aggtttaaca acccgtaaac tcgcccagaa gctaggtgta
     1861 gagcagccta cattgtattg gcatgtaaaa aataagcggg ctttgctcga cgccttagcc
     1921 attgagatgt tagataggca ccatactcac ttttgccctt tagaagggga aagctggcaa
     1981 gattttttac gtaataacgc taaaagtttt agatgtgctt tactaagtca tcgcgatgga
     2041 gcaaaagtac atttaggtac acggcctaca gaaaaacagt atgaaactct cgaaaatcaa
     2101 ttagcctttt tatgccaaca aggtttttca ctagagaatg cattatatgc actcagcgca
     2161 gtggggcatt ttactttagg ttgcgtattg gaagatcaag agcatcaagt cgctaaagaa
     2221 gaaagggaaa cacctactac tgatagtatg ccgccattat tacgacaagc tatcgaatta
     2281 tttgatcacc aaggtgcaga gccagccttc ttattcggcc ttgaattgat catatgcgga
     2341 ttagaaaaac aacttaaatg tgaaagtggg tcttaaggca tcaaataaaa cgaaaggctc
     2401 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
     2461 ggacaaatcc gccgccctag acgaagcggc atgcatttac gttgacacca tcgaatggcg
     2521 caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa
     2581 tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
     2641 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
     2701 ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca
     2761 gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt
     2821 cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga
     2881 acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag
     2941 tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg
     3001 cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat
     3061 tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca
     3121 gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg
     3181 ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg
     3241 gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac
     3301 tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc
     3361 cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc
     3421 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
     3481 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
     3541 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
     3601 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     3661 gtgagctttc taactaaaaa ggcctcccaa atcggggggc cttttttatt gataacaaaa
     3721 cgctcctagg cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3781 tatccacaga atcaggggat aacgcatgca agaacatgtg agcaaaaggc cagcaaaagg
     3841 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3901 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     3961 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4021 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgat
     4081 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4141 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4201 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4261 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     4321 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4381 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4441 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4501 agtggaacga aaactcacgt taagggattt tggtcatgac tagtgcttgg attctcacca
     4561 ataaaaaacg cccggcggca accgagcgtt ctgaacaaat ccagatggag ttctgaggtc
     4621 attactggat ctatcaacag gagtccaagc gagctcgata tcaaattacg ccccgccctg
     4681 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
     4741 aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
     4801 tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
     4861 actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa taaacccttt
     4921 agggaaatag gccaggtttt caccataaca cgccacatct tgcgaatata tgtgtagaaa
     4981 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
     5041 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
     5101 catacgaaat tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
     5161 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
     5221 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
     5281 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
     5341 tcctgaaaat c
//
