LOCUS       sequence-374009-        7430 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of PbuCas13b nuclease in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7430)
  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 7430)
  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: pPbuCas13b
FEATURES             Location/Qualifiers
     source          1..7430
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(398..415)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(532..1077)
                     /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(612..631)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1153..1172
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     RBS             1282..1290
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(4720..4737)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4890..4976
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5068..5095
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        5114..5205
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5663..6118
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5750..5769)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5960..5981
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(6674..7333)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     7248..7267
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(7334..7430,1..6))
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 cgatcaacgt ctcattttcg ccaaaagttg gcccagggct tcccggtatc aacagggaca
       61 ccaggattta tttattctgc gaagtgatct tccgtcacag gtatttattc ggcgcaaagt
      121 gcgtcgggtg atgctgccaa cttactgatt tagtgtatga tggtgttttt gaggtgctcc
      181 agtggcttct gtttctatca gctgtccctc ctgttcagct actgacgggg tggtgcgtaa
      241 cggcaaaagc accgccggac atcagcgcta gcggagtgta tactggctta ctatgttggc
      301 actgatgagg gtgtcagtga agtgcttcat gtggcaggag aaaaaaggct gcaccggtgc
      361 gtcagcagaa tatgtgatac aggatatatt ccgcttcctc gctcactgac tcgctacgct
      421 cggtcgttcg actgcggcga gcggaaatgg cttacgaacg gggcggagat ttcctggaag
      481 atgccaggaa gatacttaac agggaagtga gagggccgcg gcaaagccgt ttttccatag
      541 gctccgcccc cctgacaagc atcacgaaat ctgacgctca aatcagtggt ggcgaaaccc
      601 gacaggacta taaagatacc aggcgtttcc ccctggcggc tccctcgtgc gctctcctgt
      661 tcctgccttt cggtttaccg gtgtcattcc gctgttatgg ccgcgtttgt ctcattccac
      721 gcctgacact cagttccggg taggcagttc gctccaagct ggactgtatg cacgaacccc
      781 ccgttcagtc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggaaa
      841 gacatgcaaa agcaccactg gcagcagcca ctggtaattg atttagagga gttagtcttg
      901 aagtcatgcg ccggttaagg ctaaactgaa aggacaagtt ttggtgactg cgctcctcca
      961 agccagttac ctcggttcaa agagttggta gctcagagaa ccttcgaaaa accgccctgc
     1021 aaggcggttt tttcgttttc agagcaagag attacgcgca gaccaaaacg atctcaagaa
     1081 gatcatctta ttaatcagat aaaatatttc taggctcatg agcccgaagt ggcgagcccg
     1141 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     1201 gatgccggcc acgatgcgtc cggcgtagag gatctgctca tgtttgacag cttatcatcg
     1261 atgcattaat acgactcact ataaggaggt ctttatcatg caaaaacaag ataaattatt
     1321 tgtagataga aagaaaaatg ctatatttgc ctttcctaaa tatataacaa taatggaaaa
     1381 caaagagaaa cctgaaccaa tttattatga attgacagat aagcatttct gggctgcctt
     1441 tcttaacttg gcgcgccaca atgtctatac caccattaat cacatcaaca gacgtttgga
     1501 aatagcagag ctaaaagacg atggatatat gatgggcatt aaaggcagct ggaacgagca
     1561 agcgaaaaaa ctcgacaaaa aagttcggct gagagacttg atcatgaaac actttccttt
     1621 tctggaagct gcagcttatg aaatgacaaa ctctaaaagt cccaacaata aggagcaacg
     1681 ggaaaaggaa caatccgaag ccctatcact taacaacttg aagaatgttc tattcatttt
     1741 cttggaaaaa cttcaggttc tcaggaacta ctactcccat tacaagtatt ctgaagaatc
     1801 cccaaaaccg atatttgaaa caagcctttt gaaaaacatg tataaggttt tcgatgccaa
     1861 tgtgcggtta gtaaagagag actatatgca tcatgagaat atagacatgc aaagagattt
     1921 tacacatctc aacaggaaaa aacaagtggg acgaacaaaa aacatcatag acagtcctaa
     1981 ctttcattac cattttgcag ataaagaagg caatatgacc atagcaggct tactgttctt
     2041 cgtctcttta tttcttgata aaaaagacgc tatttggatg cagaaaaagc taaagggatt
     2101 taaagatggc agaaacttgc gtgagcaaat gacaaacgaa gtattttgca gaagccgtat
     2161 atctttgcct aagctaaaat tagagaatgt acagacaaaa gactggatgc aacttgatat
     2221 gcttaacgaa ttggttcgct gtcccaaatc gctgtatgaa cggttacgag aaaaagatag
     2281 ggaaagtttt aaagtacctt tcgatatttt ctctgacgac tataatgctg aagaagaacc
     2341 atttaagaac acgcttgtaa ggcatcaaga ccgctttcca tattttgtct tgagatattt
     2401 cgatttaaat gagatattcg aacaactccg cttccaaata gatttaggta cctatcattt
     2461 ctctatctat aacaaacgta tcggtgacga agatgaagtt cgccatctca cacatcatct
     2521 atatggcttt gcgcgcattc aggactttgc accacaaaac caaccagaag aatggagaaa
     2581 attagtgaaa gacttagatc attttgagac atcgcaagag ccttacatat ctaagactgc
     2641 tccacattac catttagaga acgaaaagat aggcattaag ttttgcagtg cccacaacaa
     2701 cctctttcca tctcttcaga ccgacaaaac atgcaatgga agaagtaaat ttaatctcgg
     2761 aacacaattt acagcagaag catttttaag tgtccacgaa ttgttgccga tgatgtttta
     2821 ttatctgtta ttgaccaagg attactccag aaaagaaagc gcagacaaag tagagggaat
     2881 tataagaaaa gaaataagca acatatacgc tatctacgat gcttttgcca ataatgagat
     2941 aaatagcatt gcagacttaa cccgtcggct tcaaaataca aatatactac aaggccattt
     3001 gccaaaacaa atgatttcca tattgaaagg cagacaaaaa gatatgggaa aagaggctga
     3061 gcgcaaaatt ggggaaatga tagacgacac ccaaagaaga cttgacttgc tctgcaagca
     3121 gacaaatcag aagataagga ttggcaaaag gaatgccggt ctgctaaaat cgggaaaaat
     3181 tgcagattgg ttggtaaatg acatgatgcg cttccaacct gttcaaaaag accaaaataa
     3241 cattccaata aacaatagca aggctaacag caccgaatat cgaatgcttc agcgtgcgct
     3301 tgcccttttc ggcagtgaga acttccgcct taaggcatat tttaatcaaa tgaatcttgt
     3361 ggggaatgac aatccgcatc ctttccttgc agaaacacaa tgggaacatc aaactaacat
     3421 tctgtctttc taccgaaact atttggaagc gaggaaaaag tatcttaagg gcttaaaacc
     3481 acaaaactgg aaacaatatc agcattttct catacttaaa gtacagaaga ccaaccgcaa
     3541 cacattagtt acaggatgga aaaacagttt caatctcccc cgtggaatat ttacccaacc
     3601 aataagggaa tggtttgaga aacataataa ttcaaaaaga atatacgacc aaatactatc
     3661 ttttgaccga gtagggtttg ttgcaaaagc tattccctta tactttgcag aagaatataa
     3721 agacaatgta cagcctttct acgattatcc gttcaatatc ggcaacagac ttaaaccaaa
     3781 aaaaaggcaa ttcttagata aaaaagagcg tgtcgaattg tggcaaaaga ataaagagct
     3841 attcaaaaat tatccgtctg aaaagaaaaa gacagactta gcttatcttg attttttatc
     3901 atggaaaaag ttcgaacgcg aactgcgact tatcaagaat caggatatcg tcacatggtt
     3961 gatgttcaaa gaactattca atatggctac tgtggaagga ttgaagattg gagaaataca
     4021 tctgagggac atcgatacaa acacggcaaa tgaggaaagc aacaatattc taaatagaat
     4081 catgcccatg aagttgcctg ttaaaacata cgaaacagac aataaaggaa atatattaaa
     4141 ggaaagacct ttagcaacct tttatatcga agaaacggaa accaaagtcc tgaaacaggg
     4201 caatttcaaa gcattggtaa aagaccgaag actcaatggg ctattctcat ttgcagaaac
     4261 tacagacttg aacttagagg aacatcctat cagcaaattg agtgtagacc ttgagcttat
     4321 aaaatatcag actacaagga ttagtatctt tgagatgaca ttaggactgg aaaagaaact
     4381 catagacaaa tattccacac tacctacaga tagcttcaga aatatgcttg aaaggtggtt
     4441 gcagtgcaaa gctaatagac cggaattaaa gaattacgtt aatagtttga ttgcagtgag
     4501 aaatgctttt tcccacaacc agtatcccat gtatgatgcc acactctttg ctgaagttaa
     4561 aaaatttact cttttccctt ctgtcgatac aaaaaagatt gagctaaata tagcgccaca
     4621 gttactggaa atagtaggaa aagccataaa agaaattgaa aaatcagaaa ataaaaacta
     4681 aaagcttggc tgttttggcg gatgagagaa gattttcagc ctgatacaga ttaaatcaga
     4741 acgcagaagc ggtctgataa aacagaattt gcctggcggc agtagcgcgg tggtcccacc
     4801 tgaccccatg ccgaactcag aagtgaaacg ccgtagcgcc gatggtagtg tggggtctcc
     4861 ccatgcgaga gtagggaact gccaggcatc aaataaaacg aaaggctcag tcgaaagact
     4921 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc
     4981 cgggagcgga tttgaacgtt gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc
     5041 cataaactgc caggcatcaa attaagcaga aggccatcct gacggatggc ctttttgcgt
     5101 ttctacaaac tcttttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     5161 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     5221 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     5281 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcagcaaa ctattaactg
     5341 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     5401 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     5461 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     5521 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     5581 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     5641 catatatact ttagattgat ttacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     5701 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     5761 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     5821 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     5881 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     5941 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     6001 tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     6061 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttaa
     6121 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     6181 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     6241 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     6301 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     6361 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     6421 agcaccgcct acatacctcg ctctgctaat cctgttacca gtcaggcatt tgagaagcac
     6481 acggtcacac tgcttccggt agtcaataaa ccggtaaacc agcaatagac ataagcggct
     6541 atttaacgac cctgccctga accgacgacc gggtcgaatt tgctttcgaa tttctgccat
     6601 tcatccgctt attatcactt attcaggcgt agcaaccagg cgtttaaggg caccaataac
     6661 tgccttaaaa aaattacgcc ccgccctgcc actcatcgca gtactgttgt aattcattaa
     6721 gcattctgcc gacatggaag ccatcacaaa cggcatgatg aacctgaatc gccagcggca
     6781 tcagcacctt gtcgccttgc gtataatatt tgcccatggt gaaaacgggg gcgaagaagt
     6841 tgtccatatt ggccacgttt aaatcaaaac tggtgaaact cacccaggga ttggctgaga
     6901 cgaaaaacat attctcaata aaccctttag ggaaataggc caggttttca ccgtaacacg
     6961 ccacatcttg cgaatatatg tgtagaaact gccggaaatc gtcgtggtat tcactccaga
     7021 gcgatgaaaa cgtttcagtt tgctcatgga aaacggtgta acaagggtga acactatccc
     7081 atatcaccag ctcaccgtct ttcattgcca tacggaattc cggatgagca ttcatcaggc
     7141 gggcaagaat gtgaataaag gccggataaa acttgtgctt atttttcttt acggtcttta
     7201 aaaaggccgt aatatccagc tgaacggtct ggttataggt acattgagca actgactgaa
     7261 atgcctcaaa atgttcttta cgatgccatt gggatatatc aacggtggta tatccagtga
     7321 tttttttctc cattttagct tccttagctc ctgaaaatct cgataactca aaaaatacgc
     7381 ccggtagtga tcttatttca ttatggtgaa agttggaacc tcttacgtgc
//
