LOCUS       sequence-416250-        9165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MWEE01000013 Cas5-HNH operon (E. coli codon optimized) + CRISPR 
            array in pACYCDuet-1 Lac promoters .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9165)
  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 9165)
  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: pHS1232
FEATURES             Location/Qualifiers
     source          1..9165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(14..1096)
                     /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     1058..1077
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1097..1174)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      complement(1188..1259)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        complement(1400..1434)
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     protein_bind    1517..1541
                     /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(6839..6857)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6853..6900
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(7121..7780)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     7695..7714
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(7781..7883)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(8275..8292)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8409..8954)
                     /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(8489..8508)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
       61 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc
      121 ttttcaccag tgacacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt
      181 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta
      241 acggcgggat ataacatgag ctgtcctcgg tatcgtcgta tcccactacc gagatgtccg
      301 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt
      361 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa
      421 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag
      481 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg
      541 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac
      601 cgtcctcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata
      661 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat
      721 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg
      781 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc
      841 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa
      901 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat
      961 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct
     1021 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata
     1081 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca
     1141 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacggag agcgttcacc
     1201 gacaaacaac agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttgg
     1261 gcgcgccaaa aaaatcgacc tgcaggcgcg ccgagctcga attcggatcc ggttcacccc
     1321 cacatgcgtg gggaatactt tttaacagtg gccttattaa atgacttctc cggttcaccc
     1381 ccacatgcgt ggggaatacg ctagtattat acctaggact gagctagctg tcaacattaa
     1441 ttcctaattt ttctctccct tatgcgactc ctgcattaga aaattttaca ctttatgctt
     1501 ccggctcgat gttgggggaa ttgtgagcgg ataacaattc ccctgtagaa ataattttgt
     1561 ttaactttaa taaggagata taccatgtat tgcgcggcgg tttgctttcc gcagacaaaa
     1621 tatcagcaaa ggggaactgc tctaaagaaa ccgctcgtcg gtctgcgtaa gatgggtgtg
     1681 gaagcggctg catggaacac cctgaaggtt actagagatc gccctaagtt gacctttccg
     1741 gatctgatca ctccgcaaag caaattcctg gataacgact tatggctgaa gtacaaagtg
     1801 ccaattgagc agaaagaaca cgtcatgtat aacctgttgt gcgataactg ggttaatgtt
     1861 gtttacttga gcggtaaacc ggaccgtatt tccttagtcc aaaccctgaa ggacgcccat
     1921 tgtctgcaat tggcgtattc taacccgatg gatcgtttca ccgtgttccg cttcctgttg
     1981 gcgttgggtt actggtgttt cgcgaacacg aacgtcgagc cggagcccga caaaccgctg
     2041 ccggtgagct ggattccgtg gctggaggaa aataaagagt acttcgagct gttcggtgat
     2101 ggtaaacgtt tcttccaagc tgacccgagc agccgtattc gtgcgattac tgacctgatc
     2161 catgaaattc cgaccgctca caacctgtgc cattttaagc acgtgaccga ctatatcgac
     2221 ggcctgtgtg aagcctgctg cataaaaggt ctgttgcgtc tgccggtatt taccaccgtt
     2281 ggcggtcgtg gcattggcgc aggtatcaat aacacgccac cgttttatct gctgtggcat
     2341 gcgaacgact tagcgggcat gttggcgcag aattggcagc cttgggacaa catgggcatc
     2401 ccggcgtggc tgggctcttt tcagaaagag tcgcgggaag ttggcctgct tgcgggtatg
     2461 acctggcttc cgcgtaaagt ttacctgcac gatccggttc cgggtcaggc tgcgtgctgc
     2521 agctgtggtt tgccgagtga agctctggtt tactcatgct ccattgaggt ggaaccagtg
     2581 ccaaagggct tggagtggaa ggacccgcac ggcgtttaca ccgaccaggg caaatccctg
     2641 cagtccaaaa tcaagctgat gagcaatgac cgctatacct tcgcggaccg tgactggtat
     2701 tctccgttgt tctcgtatct tcacgccgaa ggtaatagcc gccagggtaa gctgtggctg
     2761 gtgggttttg cgagcgataa agcaaaatct atcgacatct gggataagat catcgagctg
     2821 gagggcaccg acaccaatga cgaactgttg gcgcaactcg ccaatcgtgc aaccgcgctg
     2881 aatgcaatgc gtaaaaagcc gctacgcggt gattttaaaa agagcgttgg caccccgcag
     2941 attgcggata tcatcccgca tgcggaaaac cgcatcgcca ttaacgcagg caaaatgacg
     3001 gaaaaccgtg gttactcttg gcaagatgcc gatacggagt acggagaact gctgaccaag
     3061 gtggcatact cgctggagcc ggcgcaaacg gtggatgctc gtctgaagcg cggtaacttt
     3121 atcagccgta agccgtggcc gattatcccc gagagcaaaa caaaacccgc agagggggac
     3181 cagaatgaat agaggaaccg ttgacttcat tgcaagcttg gagaatttaa aagaaggaga
     3241 tctagggata ctcagaaagt tacgtggtgc gcgtctggac gaaaaacttc cgggttttga
     3301 tctgttcagc gcgctgtggt ggccattgcg tcaaaaaaac cagcgcgcac cgaagcgcga
     3361 ggtggcttgg ctgattgcca aattgtttgc agaattccgc tttgagcaac gtgagggcgc
     3421 gacgctgccg attctgatgg gcggtatctg ccgtaagttg gaaccgaaga aagaactgcc
     3481 gcgtgtgttg gctcgtttcg accagctggc gagcctggac atcatgcaga tggaagagcc
     3541 gctgtccgtg attatgggca tcctgcgcaa gcaccagcag gtctgtctgg actgggttgg
     3601 tctgaccgat gttttgtcgt tctgggaaca agagccggtt aaacgtgagt ggtctgatag
     3661 ctttatcaaa gcgtacaaga tcaacaagga ggattcagat gttgattgaa attcatatga
     3721 ttcagaatca ctcacccgct aatctaaaca gggacgattt aggcgctcca aaaacgtgct
     3781 atttcggtgg cgtgctgcgt agccgcatct cctctcagtg tattaagcgc agcatccgta
     3841 cgagcaatga cttcaaagca ttgctcggcg gtgtacgtac cagacgtctg gcggatctga
     3901 ttcagcaaga ggcgggcgag acagagtgct ggaaaaaggc tcaagagatc ctgaataagt
     3961 gcggcttcaa aaacaaggac gacaatacca aaatgttggt ctttatgagc aaagacaaga
     4021 tcaaagacct tgcgcgtatt gttttggaca actcgctggg cctgaccgaa gccgcccaac
     4081 aggtggcgaa cgtgatcgct caggctaccc tagctccgga catcgcgctt tgcggtcgta
     4141 tgctggaacc gaatgataag gacaaggaca agaaggtgaa gtggtcaaac actaccgttg
     4201 aggcagcgtt gcaagtggcg catgcaatta gcacccatat tgcccgtccg gaaatcgact
     4261 acttcgtggc cgcggatgat gttccgggtg aagatgcggg tgcgggccac atcggcgaat
     4321 ctatgtttgc aagcgcatgt ttttataaat acttcagcat cgattgggaa caactggtta
     4381 agaacttgaa aggtgatacc aacctggcgg ctcacaccgt cggagccttt ctgctggcgg
     4441 cggcgaaaac caatccgagc ggtaaacaga atagctttgc cgcgcacaac tacccggatg
     4501 gtattctggt cgagttcaag aactccccga tttcctatgc aaatgctttt gttcgcccag
     4561 ttagcgttgt taaagagtcg gatttggtcg agcagtccat cggtcagctg agcaactacg
     4621 tgaacgacat ccgcttgggc tactatgatg aacagagccc ggtgattggt ttctggtttt
     4681 ccccgaacaa ccgttatccg ctgggttaca aacacagcaa actggcgagc cgtaatattg
     4741 gtaacctgaa cgaactcgtg ggtgcagttc tggattacat cggcggcttc aagtgggaag
     4801 aagttcagaa aagcaaggct tatattggcg gctaacatga gtgcgccacc caacacctta
     4861 ttcttgaggt tagaaggggc tctacaatca tggggatcga acgaagcgaa gttcgccctg
     4921 cgccggaccg cagacgcgcc aactaaatcc ggcgttctgg gcctgctgtg cgcggctatg
     4981 ggcatcggtc gtgccgaggc ggcggactct tggctgccga aactggctaa tctgcgtatg
     5041 ggtgttcgta tcgatcgtcc gggcatccgc tggtgggatt ttcacaccgt aggcgcaggc
     5101 caacgtatgc gtatggcgga gctgaaggcg ccaaagaagc cgagcatggt gggcgcagcg
     5161 ctggcggaaa ccctgacccc gagcaaagtc aaaacccgtg cggaaacctt gttgtcccgc
     5221 cgtgagtacc tcgcggacgc tagcttcctg gtggcgttgc aaggtgagcc ggaacttgtc
     5281 gcaaagctgt cggcggcctt agccaaaccg gtttgggcga tttatttggg tcgtaagagc
     5341 tgtccgccat ctcgccccgt gtgcgaacat ccgccgggct tttataacac cctggaagaa
     5401 gcgctgagcg cagttccgct acaaaaacgt tggcataacg agccgctgcc gcagattctg
     5461 ccgtgtgtta tggactggat tccgggttat gatggtgagc acgctccgga tgatgcggag
     5521 atccactacg acttgccggt gagcttccag ccgcctcgtc atctgccgcg ttttgttatt
     5581 cgtcgggaac tggtcgtggg tgaggacgtt caggtttcca gagaaactgg tacgagcgtc
     5641 tggcgtccta agggtacgcg tgcggactac aacaacagcg aatacaaaaa ggtgcgcgcc
     5701 gagcgcctgg tgatggatca cgctgcctgc atggtttgca aagcaccggc aaccaccgtg
     5761 cagcacgtga attatcgtcg cgctggcggt aaagagatcc cggaggattt aagagctttg
     5821 tgccgcctgt gccatgacgc gtgtaccatg ttagaatacg gctctggtat gaccacgaat
     5881 cgtatcgacc cgtgcgatcc gatttggaga gaacgtatcc tcgctaagcg caaggagatt
     5941 gttgaatttc gtagccgcgg tcagcgtttc cgcaaaatga agccagagga ggaaaacgga
     6001 tgatctactt gtccaggcta ttgatcgata ctggaggtaa tccagataga cctaggcccg
     6061 ggcgcaaatg gctcgacaac atttataacg tgcatcgtcg tctgagcatg gcatttccga
     6121 gcggtctgcg ccgtgaacag gacccgcatt ttctgaagcc gttttctccg aatgacttcc
     6181 agaaaacgcc attcttgttt cgtgtcgaca ataatatcga cggcaacgat aaacgtgcga
     6241 ttatcatcgt gcaaagcgtc ctggagcccg actgggatta ctgctttcaa aacgccttag
     6301 acttcttggc ggcgccaccg gaaaccaaag agtacaaccc ggaatttaaa gcgggtcagt
     6361 tgctccggtt ccgcctgcgt gttaatgcat ccgtgcgtag acacatcccg gagatggttc
     6421 agcaggatgg ccagaccatt gaaactggta agatcctgca caagcgcgtg agcctgacct
     6481 gggacgctag ctcgaccccg gatcaggctt tagctgattg gctggcggcg aagagcccga
     6541 agttgggctt cacgctgcag cgttgtgaac tgctgcaact gggttgggtt tatggtagca
     6601 aaccggaacc gaagaacgtg aaagttaaag aacaaggcca aggctattgg cgtgagcaca
     6661 aatacaaccc gctgcgcttc cgtgccgcat tgcttgaggg cgttctggag gttgatgatc
     6721 cgaagctgtt cctgaaaacc ctgagcagcg gtattggtaa ggcgaagagt tttggtttcg
     6781 gcctgctgtc cgttctgcca atcaggaatg atggataaat taacctaggc tgctgccacc
     6841 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     6901 ctgaaacctc aggcatttga gaagcacacg gtcacactgc ttccggtagt caataaaccg
     6961 gtaaaccagc aatagacata agcggctatt taacgaccct gccctgaacc gacgaccggg
     7021 tcgaatttgc tttcgaattt ctgccattca tccgcttatt atcacttatt caggcgtagc
     7081 aaccaggcgt ttaagggcac caataactgc cttaaaaaaa ttacgccccg ccctgccact
     7141 catcgcagta ctgttgtaat tcattaagca ttctgccgac atggaagcca tcacaaacgg
     7201 catgatgaac ctgaatcgcc agcggcatca gcaccttgtc gccttgcgta taatatttgc
     7261 ccatagtgaa aacgggggcg aagaagttgt ccatattggc cacgtttaaa tcaaaactgg
     7321 tgaaactcac ccagggattg gctgacacga aaaacatatt ctcaataaac cctttaggga
     7381 aataggccag gttttcaccg taacacgcca catcttgcga atatatgtgt agaaactgcc
     7441 ggaaatcgtc gtggtattca ctccagagcg atgaaaacgt ttcagtttgc tcatggaaaa
     7501 cggtgtaaca agggtgaaca ctatcccata tcaccagctc accgtctttc attgccatac
     7561 ggaactccgg atgagcattc atcaggcggg caagaatgtg aataaaggcc ggataaaact
     7621 tgtgcttatt tttctttacg gtctttaaaa aggccgtaat atccagctga acggtctggt
     7681 tataggtaca ttgagcaact gactgaaatg cctcaaaatg ttctttacga tgccattggg
     7741 atatatcaac ggtggtatat ccagtgattt ttttctccat tttagcttcc ttagctcctg
     7801 aaaatctcga taactcaaaa aatacgcccg gtagtgatct tatttcatta tggtgaaagt
     7861 tggaacctct tacgtgccga tcaacgactc attttcgcca aaagttggcc cagggcttcc
     7921 cggtatcaac agggacacca ggatttattt attctgcgaa gtgatcttcc gtcacaggta
     7981 tttattcggc gcaaagtgcg tcgggtgatg ctgccaactt actgatttag tgtatgatgg
     8041 tgtttttgag gtgctccagt ggcttctgtt tctatcagct gtccctcctg ttcagctact
     8101 gacggggtgg tgcgtaacgg caaaagcacc gccggacatc agcgctagcg gagtgtatac
     8161 tggcttacta tgttggcact gatgagggtg tcagtgaagt gcttcatgtg gcaggagaaa
     8221 aaaggctgca ccggtgcgtc agcagaatat gtgatacagg atatattccg cttcctcgct
     8281 cactgactcg ctacgctcgg tcgttcgact gcggcgagcg gaaatggctt acgaacgggg
     8341 cggagatttc ctggaagatg ccaggaagat acttaacagg gaagtgagag ggccgcggca
     8401 aagccgtttt tccataggct ccgcccccct gacaagcatc acgaaatctg acgctcaaat
     8461 cagtggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggcggctcc
     8521 ctcgtgcgct ctcctgttcc tgcctttcgg tttaccggtg tcattccgct gttatggccg
     8581 cgtttgtctc attccacgcc tgacactcag ttccgggtag gcagttcgct ccaagctgga
     8641 ctgtatgcac gaaccccccg ttcagtccga ccgctgcgcc ttatccggta actatcgtct
     8701 tgagtccaac ccggaaagac atgcaaaagc accactggca gcagccactg gtaattgatt
     8761 tagaggagtt agtcttgaag tcatgcgccg gttaaggcta aactgaaagg acaagttttg
     8821 gtgactgcgc tcctccaagc cagttacctc ggttcaaaga gttggtagct cagagaacct
     8881 tcgaaaaacc gccctgcaag gcggtttttt cgttttcaga gcaagagatt acgcgcagac
     8941 caaaacgatc tcaagaagat catcttatta atcagataaa atatttctag atttcagtgc
     9001 aatttatctc ttcaaatgta gcacctgaag tcagccccat acgatataag ttgtaattct
     9061 catgttagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
     9121 catcggtcga gatcccggtg cctaatgagt gagctaactt acatt
//
