LOCUS       sequence-421621-        8384 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NZ_CP025815 DinG HNH proteins (E. coli codon optimized) and CRISPR 
            array in pACYCDuet-1 with Lac promoters.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8384)
  AUTHORS   Altae-Tran H, Kannan S, Suberski AJ, Mears KS, Demircioglu FE, 
            Moeller L, Kocalar S, Oshiro R, Makarova KS, Macrae RK, Koonin EV, 
            Zhang F
  TITLE     Uncovering the functional diversity of rare CRISPR-Cas systems with 
            deep terascale clustering.
  JOURNAL   Science. 2023 Nov 24;382(6673):eadi1910. doi: 
            10.1126/science.adi1910. Epub 2023 Nov 23.
  PUBMED    37995242
REFERENCE   2  (bases 1 to 8384)
  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: "Science."; 
            date: "2023-11-24"; volume: "382(6673):eadi1910. doi"; pages: " 
            10.1126/science.adi1910. Epub 2023 Nov 23"
COMMENT     Sequence Label: pHS1068
FEATURES             Location/Qualifiers
     source          1..8384
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(218..252)
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     promoter        join(303..308,309..326,327..333)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    336..360
                     /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)."
     promoter        join(3075..3080,3081..3098,3099..3105)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3108..3132
                     /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)."
     primer_bind     complement(4864..4882)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4878..4925
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(5146..5805)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5720..5739
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(5806..5908)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(6300..6317)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6434..6979)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(6514..6533)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7204..8286)
                     /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     8248..8267
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(8287..8364)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      complement(join(8378..8384,1..65))
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttcaccgaca aacaacagat aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt
       61 atttgggcgc gccaaaaaaa tcgacctgca ggcgcgccga gctcgaattc ggatcccttt
      121 agtagccacc caatttccgg atggctgtat ttttaacagt ggccttatta aatgacttct
      181 ccctttagta gccacccaat ttccggatgg ctgtattgct agtattatac ctaggactga
      241 gctagctgtc aacattaatt cctaattttt ctctccctta tgcgactcct gcattaggaa
      301 attttacact ttatgcttcc ggctcgtatg ttgggggaat tgtgagcgga taacaattcc
      361 cctgtagaaa taattttgtt taactttaat aaggagatat accatggaaa atctgatttt
      421 tacggcttcg ctcaaatcac ccctaatagc tcaaggattt atgacgtttg acagcctgct
      481 tgctgccgtt ctgttcgatc gttgtggtga cgtggagact gcgcatacct ctgttccgct
      541 gaagtgctcc gaaggcctgt tccacgcgag cgcagcgatt gtgaccccgg ttgaacgtgg
      601 tagctacagc attgcggcgt ccctgcgcgc ggaccacgat ttggacgcgg acttattccc
      661 gaaaaacaaa tctggtcgga tttatcgtaa gatgggtcgt aagcgccgtc gtgatctcgg
      721 gaacgtgctg agcacctacc atacgatcag caccccgtcg gtgcgttgga tctgcaccgg
      781 cgacctggat caaatcaaag gcctgctggg cgacttgaga ttcatcggca agaagcgcgc
      841 acagggtttt ggtgaagttg ccgcttggtc ggtcgagaaa accgatctgg acccgttggt
      901 cgatgaacag ggcctgccgc gtcgccctat tccggagaac atgtttaccg gtgaccgtag
      961 cctgccactg caagatctgg cgtggaaacc ggcatattgg aatccggagc accgcgcagc
     1021 ttgttacgcg cctcagccgc tttgaaggag ggccggatga atacctttga actcatatcc
     1081 aagggctttg ctaggcacgt actaggaaat tcaacaacgg gcgacccgat tcgtgatgct
     1141 gccaaggcgc tgacgatgaa ccacctgctg aacaacgaga aaggcacttc tagccgtttg
     1201 accggttggg cgattttccg cgctagcggt agcaccgtcc aggcgaactt ggcggctaag
     1261 ggtgtgatgg cgaacgacgg ccgtacccgt tacgaggcgt tcgcggatga gctgtcggat
     1321 ttcagagaac cggtcgtgtt cctgcaattt accaagtcca ccgtgaattt aggtaatgtt
     1381 tttgcgacct ttgacccacg tgttaccaaa atctgcgcac cggatgttgc accggagatc
     1441 tttgacttca gcgttggtac gctgccggaa accgcaggcg tggccgaacg taatatccgc
     1501 aaagccgttc tgaagaagga gaaaagcaat gctgcatgat cacgtcatcc ccaagccgcc
     1561 cacactagga gcatatgcta gtcaccaatt gctggagggc ctgactgacg gcgcaagacc
     1621 actgtttgtt gataatggcg accacctctt ggttcgttcc gtggatcaat cgctgaccca
     1681 atctagcaaa ccggtcccga cggtgaacct gggtgacgtg ttaggcttcg aactgcgcgc
     1741 cagctgcggt gtgaaggtga agggtaaaca tcgttacttc ccgctgaagg actggcgtgc
     1801 tcgccatgcg tggctgaatc gtcgtgcgag cggttttgat gtcctggctg tgaccgttag
     1861 cgcacgctcc accccgatta aaaagaacat ccgtattgat cagagcgatt ttgcgggtgt
     1921 attgcgtgtt accgacatcg aagcgttcaa ccaggttctt gagcacggca tcggcggtcc
     1981 gggtaaagcc ttcggccacg gaatgctggt aatctgaagg agaacaccat gaaccaatca
     2041 actttaatta ccgaagtcgt aataaggtta gaagctcccc tatcaattgc tatgccggtg
     2101 gcagagtaca ccaagccgaa caaatgggaa aactttccgg tggctccggt gggttatgat
     2161 gaagagggtc aaccgatcca gtcaggcttc ctgccggcaa ccaccattcg tggtgcgatc
     2221 cgccgttttg cggtgctgcc agaaatggca gcggctgcgg agaagggtgc gcactaccgc
     2281 ctggagcgtg catacagcga actcattggt caggacccgg gctccgaaca gcagccggac
     2341 gagatcgatc tgactgccat cgccgaagcg cgtgaggaat ccccgattgt tgatctgttt
     2401 ggctctggcc tgggtgtcaa aagccgcttg ctggtgagca atttcgtgcc ggaccatccg
     2461 acgctgccga tggcgttcac ctgggttcgt aaggacatcg gcgacacgga aggtgttttg
     2521 gaggcgttgc cagacgagga tcgttctgcg tacctggcgc gtgagcaagc gaaccgtcac
     2581 cgtgcccaag cggaggcctt ggttgcggac ctgaagcgca aagaatcgaa agccgttaaa
     2641 accggttccg atgatgcagc tgacttgaag gtgcagctgg aagctgctga gaagctcgcg
     2701 gctaaatata aagaggacat gggtgacatg caagtttcca cccgtagcct gggagttagc
     2761 tatgcgttac cgcagggtct ggagctcaag gggaagttcg tcgtgcgtag cgcgcgcgag
     2821 agggatcagg acatcctgct gcgcggcctg gatggcttat ctcgttggcc cgtattgggc
     2881 gcacaaagcg cgagaggcgc aggtgaaatt agcggtactg cgagctttaa actggacggc
     2941 aaaattacca agattgtttc gttcggcggt ttcgaaccgc taaaaattcg cgactttgaa
     3001 gcaacttaag cggccgcata atgcttaagt cgaacagaaa gtaatcgtat tgtacacggc
     3061 cgcataatcg aaattttaca ctttatgctt ccggctcgta tgttggggga attgtgagcg
     3121 gataacaatt ccccatctta gtatattagt taagtataag aaggagatat acatatgttc
     3181 agatcgccag aacaagctga gttcaaccaa tttgtagcta atcacataag taatccggat
     3241 gcaccgctgt tgattgaggg cgcaactggc ctgggtaaga cccgtgccta cctggcggcg
     3301 gtgttccaga ccgacaaacg tgtggcgatc tgcctggcca cgaacgccct tatcgagcag
     3361 attctgaaca gctctgatct gccgtgggcg caggagctgg cgccagataa aaccgttgcg
     3421 gtgttcagaa gccgtcgtta cttcgaagat gaccgtgagg cgtacgaagc tcagcgtgag
     3481 gcggcccaaa tcgccgatat tctgatctgc accgcgtcct ccgtcatctt cgaccaacgc
     3541 ttgtccggta gctataacgg cgttaccaat cgcgacgtta tcgtgttcga cgaagccgat
     3601 cagattccgg gtctggcggc tttggcgagc gacctgagca ttgatcgtaa aacgttgcgc
     3661 gacttaaact gcgcggcctc gaccgcactt gaggttgcgg gtaagctgct gagcctgccg
     3721 cagctggata gcgaaattcg cgcaaaggct aagatcatcg ctgaaattgc ttccgaaccg
     3781 gaagtttggt ataaacgtgt gggtatgacc gaagaaggtg gcgttagcgt tatccaccgt
     3841 ctgcctggcc gcttgctgaa aaaaattagt aatcgcccga gcaccatttt tattagcgca
     3901 acgttgtcga tcaacggcca atttaatgac tttaagaggg ctatgggtat tggcgacagc
     3961 tcttcgttgt cacgtatcat agagccgaaa caccacggtc atctgagctt tagcttcttt
     4021 acggacgatc cggttgattc cgatgagtgg ctggcgaccg tcgtggagca aatcgaggct
     4081 tcggacaccc cggtgctggt cgccacaccg agccataagc tggcaacgga attaggcgat
     4141 agaatttctg gctctacggt gcgtcaacgt gatgaaacga tgaccgatgc tgttggtcgt
     4201 atgggcgaca gagacattgt gatcggcgca ggtgcatggg cgggtatgga caccccggtg
     4261 caatgggcga ccgtgatcat cccgcgtgtt ccatttaccg gtccgaatga actgttcgac
     4321 acctggaccg atgaggacgc gtttcgtatt ggcgacccga tgaccagcta tttcgatagc
     4381 aaaaacgcgg cgacccgccg actcaagcag gttttcggtc gtggtttgcg ccaccccgac
     4441 gctcgttgtg ctatcgtcat ttgcgatccg cgtatcagcc agctgggcga cgtggcccca
     4501 gtgcgttttc gcgaaggttg gttcgagggt cgtcgcgtgg aggttgttca aagcaaagcg
     4561 gagcgcaacc cggcactgcg ccgtgacgcg ctgcgtcatc atggaaccga ttgtcagacc
     4621 tgtggtcacc agccgctgat tctccgcgaa gttgaagtcc accaccttaa tccgattgct
     4681 gaggcgaagg gaatgatcgc gacttccatg gacgacgtgg cggttctgtg ccgtatctgc
     4741 catgcacgtg cgcacaaaga tggtaacaac gtaattccgc tggagcgtct gcgtgagatc
     4801 gcgaaggcaa agtctcgcac agaagcccag cagctaatta gtttctgacc taggctgctg
     4861 ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt
     4921 ttttgctgaa acctcaggca tttgagaagc acacggtcac actgcttccg gtagtcaata
     4981 aaccggtaaa ccagcaatag acataagcgg ctatttaacg accctgccct gaaccgacga
     5041 ccgggtcgaa tttgctttcg aatttctgcc attcatccgc ttattatcac ttattcaggc
     5101 gtagcaacca ggcgtttaag ggcaccaata actgccttaa aaaaattacg ccccgccctg
     5161 ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga agccatcaca
     5221 aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt gcgtataata
     5281 tttgcccata gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt ttaaatcaaa
     5341 actggtgaaa ctcacccagg gattggctga cacgaaaaac atattctcaa taaacccttt
     5401 agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata tgtgtagaaa
     5461 ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag tttgctcatg
     5521 gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt ctttcattgc
     5581 catacggaac tccggatgag cattcatcag gcgggcaaga atgtgaataa aggccggata
     5641 aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca gctgaacggt
     5701 ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt tacgatgcca
     5761 ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag cttccttagc
     5821 tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt cattatggtg
     5881 aaagttggaa cctcttacgt gccgatcaac gactcatttt cgccaaaagt tggcccaggg
     5941 cttcccggta tcaacaggga caccaggatt tatttattct gcgaagtgat cttccgtcac
     6001 aggtatttat tcggcgcaaa gtgcgtcggg tgatgctgcc aacttactga tttagtgtat
     6061 gatggtgttt ttgaggtgct ccagtggctt ctgtttctat cagctgtccc tcctgttcag
     6121 ctactgacgg ggtggtgcgt aacggcaaaa gcaccgccgg acatcagcgc tagcggagtg
     6181 tatactggct tactatgttg gcactgatga gggtgtcagt gaagtgcttc atgtggcagg
     6241 agaaaaaagg ctgcaccggt gcgtcagcag aatatgtgat acaggatata ttccgcttcc
     6301 tcgctcactg actcgctacg ctcggtcgtt cgactgcggc gagcggaaat ggcttacgaa
     6361 cggggcggag atttcctgga agatgccagg aagatactta acagggaagt gagatggccg
     6421 cggcaaagcc gtttttccat aggctccgcc cccctgacaa gcatcacgaa atctgacgct
     6481 caaatcagtg gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggcg
     6541 gctccctcgt gcgctctcct gttcctgcct ttcggtttac cggtgtcatt ccgctgttat
     6601 ggccgcgttt gtctcattcc acgcctgaca ctcagttccg ggtaggcagt tcgctccaag
     6661 ctggactgta tgcacgaacc ccccgttcag tccgaccgct gcgccttatc cggtaactat
     6721 cgtcttgagt ccaacccgga aagacatgca aaagcaccac tggcagcagc cactggtaat
     6781 tgatttagag gagttagtct tgaagtcatg cgccggttaa ggctaaactg aaaggacaag
     6841 ttttggtgac tgcgctcctc caagccagtt acctcggttc aaagagttgg tagctcagag
     6901 aaccttcgaa aaaccgccct gcaaggcggt tttttcgttt tcagagcaag agattacgcg
     6961 cagaccaaaa cgatctcaag aagatcatct tattaatcag ataaaatatt tctagatttc
     7021 agtgcaattt atctcttcaa atgtagcacc tgaagtcagc cccatacgat ataagttgta
     7081 attctcatgt tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc
     7141 aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg
     7201 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     7261 caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag
     7321 tgacacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg
     7381 gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat
     7441 ataacatgag ctgtcctcgg tatcgtcgta tcccactacc gagatgtccg caccaacgcg
     7501 cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag
     7561 catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat
     7621 ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt
     7681 atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc
     7741 gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcctcatg
     7801 ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac
     7861 attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat
     7921 cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc
     7981 gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat
     8041 cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag
     8101 caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc
     8161 catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac
     8221 gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg
     8281 tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa
     8341 ggttttgcgc cattcgatgg tgtccgggat ctcgacagag agcg
//
