LOCUS       sequence-373965-        7816 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a single-spacer CRISPR array with spacer #1 targeting 
            the MS2 phage genome and expression of PbuCas13b in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7816)
  AUTHORS   Wandera KG, Alkhnbashi OS, Bassett HVI, Mitrofanov A, Hauns S, Migur
            A, Backofen R, Beisel CL
  TITLE     Anti-CRISPR prediction using deep learning reveals an inhibitor of 
            Cas13b nucleases.
  JOURNAL   Mol Cell. 2022 May 24. pii: S1097-2765(22)00437-3. doi: 
            10.1016/j.molcel.2022.05.003.
  PUBMED    35649413
REFERENCE   2  (bases 1 to 7816)
  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: "Mol Cell. 
            2022 May 24. pii: S1097-2765(22)00437-3. doi: 
            10.1016/j.molcel.2022.05.003."
COMMENT     Sequence Label: pPlacPbuCas13b-gRNA-T-MS2
FEATURES             Location/Qualifiers
     source          1..7816
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(489..506)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(623..1168)
                     /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(703..722)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(1280..1285,1286..1302,1303..1308)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     promoter        join(1458..1463,1464..1481,1482..1487)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     promoter        4887..4921
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     primer_bind     complement(5279..5298)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(7151..7810)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     7725..7744
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(7811..7816,1..97))
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 ttccttagct cctgaaaatc tcgataactc aaaaaatacg cccggtagtg atcttatttc
       61 attatggtga aagttggaac ctcttacgtg ccgatcaacg tctcattttc gccaaaagtt
      121 ggcccagggc ttcccggtat caacagggac accaggattt atttattctg cgaagtgatc
      181 ttccgtcaca ggtatttatt cggcgcaaag tgcgtcgggt gatgctgcca acttactgat
      241 ttagtgtatg atggtgtttt tgaggtgctc cagtggcttc tgtttctatc agctgtccct
      301 cctgttcagc tactgacggg gtggtgcgta acggcaaaag caccgccgga catcagcgct
      361 agcggagtgt atactggctt actatgttgg cactgatgag ggtgtcagtg aagtgcttca
      421 tgtggcagga gaaaaaaggc tgcaccggtg cgtcagcaga atatgtgata caggatatat
      481 tccgcttcct cgctcactga ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg
      541 gcttacgaac ggggcggaga tttcctggaa gatgccagga agatacttaa cagggaagtg
      601 agagggccgc ggcaaagccg tttttccata ggctccgccc ccctgacaag catcacgaaa
      661 tctgacgctc aaatcagtgg tggcgaaacc cgacaggact ataaagatac caggcgtttc
      721 cccctggcgg ctccctcgtg cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc
      781 cgctgttatg gccgcgtttg tctcattcca cgcctgacac tcagttccgg gtaggcagtt
      841 cgctccaagc tggactgtat gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc
      901 ggtaactatc gtcttgagtc caacccggaa agacatgcaa aagcaccact ggcagcagcc
      961 actggtaatt gatttagagg agttagtctt gaagtcatgc gccggttaag gctaaactga
     1021 aaggacaagt tttggtgact gcgctcctcc aagccagtta cctcggttca aagagttggt
     1081 agctcagaga accttcgaaa aaccgccctg caaggcggtt ttttcgtttt cagagcaaga
     1141 gattacgcgc agaccaaaac gatctcaaga agatcatctt attaatcaga taaaatattt
     1201 ctagatttca gtgcaattta tctcttcaaa tgtagcacct gaagtcagcc ccatacgata
     1261 taagttgtaa ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat
     1321 cacagttaaa ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg
     1381 tcatcctcgg caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc
     1441 cgggcctctt gcgggatttt acactttatg cttccggctc gtatgttagg aggtctttat
     1501 catgcaaaaa caagataaat tatttgtaga tagaaagaaa aatgctatat ttgcctttcc
     1561 taaatatata acaataatgg aaaacaaaga gaaacctgaa ccaatttatt atgaattgac
     1621 agataagcat ttctgggctg cctttcttaa cttggcgcgc cacaatgtct ataccaccat
     1681 taatcacatc aacagacgtt tggaaatagc agagctaaaa gacgatggat atatgatggg
     1741 cattaaaggc agctggaacg agcaagcgaa aaaactcgac aaaaaagttc ggctgagaga
     1801 cttgatcatg aaacactttc cttttctgga agctgcagct tatgaaatga caaactctaa
     1861 aagtcccaac aataaggagc aacgggaaaa ggaacaatcc gaagccctat cacttaacaa
     1921 cttgaagaat gttctattca ttttcttgga aaaacttcag gttctcagga actactactc
     1981 ccattacaag tattctgaag aatccccaaa accgatattt gaaacaagcc ttttgaaaaa
     2041 catgtataag gttttcgatg ccaatgtgcg gttagtaaag agagactata tgcatcatga
     2101 gaatatagac atgcaaagag attttacaca tctcaacagg aaaaaacaag tgggacgaac
     2161 aaaaaacatc atagacagtc ctaactttca ttaccatttt gcagataaag aaggcaatat
     2221 gaccatagca ggcttactgt tcttcgtctc tttatttctt gataaaaaag acgctatttg
     2281 gatgcagaaa aagctaaagg gatttaaaga tggcagaaac ttgcgtgagc aaatgacaaa
     2341 cgaagtattt tgcagaagcc gtatatcttt gcctaagcta aaattagaga atgtacagac
     2401 aaaagactgg atgcaacttg atatgcttaa cgaattggtt cgctgtccca aatcgctgta
     2461 tgaacggtta cgagaaaaag atagggaaag ttttaaagta cctttcgata ttttctctga
     2521 cgactataat gctgaagaag aaccatttaa gaacacgctt gtaaggcatc aagaccgctt
     2581 tccatatttt gtcttgagat atttcgattt aaatgagata ttcgaacaac tccgcttcca
     2641 aatagattta ggtacctatc atttctctat ctataacaaa cgtatcggtg acgaagatga
     2701 agttcgccat ctcacacatc atctatatgg ctttgcgcgc attcaggact ttgcaccaca
     2761 aaaccaacca gaagaatgga gaaaattagt gaaagactta gatcattttg agacatcgca
     2821 agagccttac atatctaaga ctgctccaca ttaccattta gagaacgaaa agataggcat
     2881 taagttttgc agtgcccaca acaacctctt tccatctctt cagaccgaca aaacatgcaa
     2941 tggaagaagt aaatttaatc tcggaacaca atttacagca gaagcatttt taagtgtcca
     3001 cgaattgttg ccgatgatgt tttattatct gttattgacc aaggattact ccagaaaaga
     3061 aagcgcagac aaagtagagg gaattataag aaaagaaata agcaacatat acgctatcta
     3121 cgatgctttt gccaataatg agataaatag cattgcagac ttaacccgtc ggcttcaaaa
     3181 tacaaatata ctacaaggcc atttgccaaa acaaatgatt tccatattga aaggcagaca
     3241 aaaagatatg ggaaaagagg ctgagcgcaa aattggggaa atgatagacg acacccaaag
     3301 aagacttgac ttgctctgca agcagacaaa tcagaagata aggattggca aaaggaatgc
     3361 cggtctgcta aaatcgggaa aaattgcaga ttggttggta aatgacatga tgcgcttcca
     3421 acctgttcaa aaagaccaaa ataacattcc aataaacaat agcaaggcta acagcaccga
     3481 atatcgaatg cttcagcgtg cgcttgccct tttcggcagt gagaacttcc gccttaaggc
     3541 atattttaat caaatgaatc ttgtggggaa tgacaatccg catcctttcc ttgcagaaac
     3601 acaatgggaa catcaaacta acattctgtc tttctaccga aactatttgg aagcgaggaa
     3661 aaagtatctt aagggcttaa aaccacaaaa ctggaaacaa tatcagcatt ttctcatact
     3721 taaagtacag aagaccaacc gcaacacatt agttacagga tggaaaaaca gtttcaatct
     3781 cccccgtgga atatttaccc aaccaataag ggaatggttt gagaaacata ataattcaaa
     3841 aagaatatac gaccaaatac tatcttttga ccgagtaggg tttgttgcaa aagctattcc
     3901 cttatacttt gcagaagaat ataaagacaa tgtacagcct ttctacgatt atccgttcaa
     3961 tatcggcaac agacttaaac caaaaaaaag gcaattctta gataaaaaag agcgtgtcga
     4021 attgtggcaa aagaataaag agctattcaa aaattatccg tctgaaaaga aaaagacaga
     4081 cttagcttat cttgattttt tatcatggaa aaagttcgaa cgcgaactgc gacttatcaa
     4141 gaatcaggat atcgtcacat ggttgatgtt caaagaacta ttcaatatgg ctactgtgga
     4201 aggattgaag attggagaaa tacatctgag ggacatcgat acaaacacgg caaatgagga
     4261 aagcaacaat attctaaata gaatcatgcc catgaagttg cctgttaaaa catacgaaac
     4321 agacaataaa ggaaatatat taaaggaaag acctttagca accttttata tcgaagaaac
     4381 ggaaaccaaa gtcctgaaac agggcaattt caaagcattg gtaaaagacc gaagactcaa
     4441 tgggctattc tcatttgcag aaactacaga cttgaactta gaggaacatc ctatcagcaa
     4501 attgagtgta gaccttgagc ttataaaata tcagactaca aggattagta tctttgagat
     4561 gacattagga ctggaaaaga aactcataga caaatattcc acactaccta cagatagctt
     4621 cagaaatatg cttgaaaggt ggttgcagtg caaagctaat agaccggaat taaagaatta
     4681 cgttaatagt ttgattgcag tgagaaatgc tttttcccac aaccagtatc ccatgtatga
     4741 tgccacactc tttgctgaag ttaaaaaatt tactcttttc ccttctgtcg atacaaaaaa
     4801 gattgagcta aatatagcgc cacagttact ggaaatagta ggaaaagcca taaaagaaat
     4861 tgaaaaatca gaaaataaaa actaaattga cagctagctc agtcctaggt ataatgctag
     4921 cgctgagttg catctgcctt ctttttgaaa ggtaaaaaca actacataaa gagttgaact
     4981 tctttgttgt ctgttgcatc tgccttcttt ttgaaaggta aaaacaacat cgtccattcc
     5041 gacagcatcg ccagtcacta tggcgtgctg ctagcgctat atgcgttgat gcaatttcta
     5101 tgcgcacccg ttctcggagc actgtccgac cgctttggcc gccgcccagt cctgctcgct
     5161 tcgctacttg gagccactat cgactacgcg atcatggcga ccacacccgt cctgtggatc
     5221 ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt tgctggcgcc
     5281 tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct catgagcgct
     5341 tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg cgccatctcc
     5401 ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc tcaacctact actgggctgc
     5461 ttcctaatgc aggagtcgca taagggagag cgtcgaccga tgcccttgag agccttcaac
     5521 ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact tatgactgtc
     5581 ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tctgggtcat tttcggcgag
     5641 gaccgctttc gctggagcgc gacgatgatc ggcctgtcgc ttgcggtatt cggaatcttg
     5701 cacgccctcg ctcaagcctt cgtcactggt cccgccacca aacgtttcgg cgagaagcag
     5761 gccattatcg ccggcatggc ggccgacgcg ctgggctacg tcttgctggc gttcgcgacg
     5821 cgaggctgga tggccttccc cattatgatt cttctcgctt ccggcggcat cgggatgccc
     5881 gcgttgcagg ccatgctgtc caggcaggta gatgacgacc atcagggaca gcttcaagga
     5941 tcgctcgcgg ctcttaccag cctaacttcg atcattggac cgctgatcgt cacggcgatt
     6001 tatgccgcct cggcgagcac atggaacggg ttggcatgga ttgtaggcgc cgccctatac
     6061 cttgtctgcc tccccgcgtt gcgtcgcggt gcatggagcc gggccacctc gacctgaatg
     6121 gaagccggcg gcacctcgct aacggattca ccactccaag aattggagcc aatcaattct
     6181 tgcggagaac tgtgaatgcg caaaccaacc cttggcagaa catatccatc gcgtccgcca
     6241 tctccagcag ccgcacgcgg cgcatctcgg gcagcgttgg gtcctggcca cgggtgcgca
     6301 tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt actggttagc
     6361 agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga
     6421 cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga
     6481 agtcccctac gtgctgctga agttgcccgc aacagagagt ggaaccaacc ggtgatacca
     6541 cgatactatg actgagagtc aacgccatga gcggcctcat ttcttattct gagttacaac
     6601 agtccgcacc gctgccggta gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc
     6661 acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag cgcccaacag
     6721 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgccct gcaccattat
     6781 gttccggatc tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt
     6841 aacgaagcgc taaccgtttt tatcaggctc tgggaggcag aataaatgat catatcgtca
     6901 attattacct ccacggggag agcctgagca aactggcctc aggcatttga gaagcacacg
     6961 gtcacactgc ttccggtagt caataaaccg gtaaaccagc aatagacata agcggctatt
     7021 taacgaccct gccctgaacc gacgaccggg tcgaatttgc tttcgaattt ctgccattca
     7081 tccgcttatt atcacttatt caggcgtagc aaccaggcgt ttaagggcac caataactgc
     7141 cttaaaaaaa ttacgccccg ccctgccact catcgcagta ctgttgtaat tcattaagca
     7201 ttctgccgac atggaagcca tcacaaacgg catgatgaac ctgaatcgcc agcggcatca
     7261 gcaccttgtc gccttgcgta taatatttgc ccatggtgaa aacgggggcg aagaagttgt
     7321 ccatattggc cacgtttaaa tcaaaactgg tgaaactcac ccagggattg gctgagacga
     7381 aaaacatatt ctcaataaac cctttaggga aataggccag gttttcaccg taacacgcca
     7441 catcttgcga atatatgtgt agaaactgcc ggaaatcgtc gtggtattca ctccagagcg
     7501 atgaaaacgt ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca ctatcccata
     7561 tcaccagctc accgtctttc attgccatac ggaattccgg atgagcattc atcaggcggg
     7621 caagaatgtg aataaaggcc ggataaaact tgtgcttatt tttctttacg gtctttaaaa
     7681 aggccgtaat atccagctga acggtctggt tataggtaca ttgagcaact gactgaaatg
     7741 cctcaaaatg ttctttacga tgccattggg atatatcaac ggtggtatat ccagtgattt
     7801 ttttctccat tttagc
//
