LOCUS       sequence-374004-        5620 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of anti-CRISPR protein AcrVIB1 in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5620)
  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 5620)
  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: pAcrVIB1
FEATURES             Location/Qualifiers
     source          1..5620
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     13..32
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        13..31
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    32..56
                     /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)."
     RBS             99..121
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             497..514
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(567..585)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      581..628
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      665..1120
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(752..771)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     962..983
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1212..2027)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1212..2027)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1935..1954
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2019..2038
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2149..2737
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2638..2657
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2891..2908
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2923..3062
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2988..3007)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3148..3170
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3164..3355)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4164..5246)
                     /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     5208..5227
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5247..5324)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5530..5549
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac aattcccctc
       61 taggatccga attcgagctc aggagggcta gcaataattt tgtttaactt taagaaggag
      121 atataccatg aaggacctgg acctgtccaa gcttaagggt gaagagattg cccagtggct
      181 tttgaacaac aagaaagcca ctgcaatcca actctcatcc gaacgtacgg atacggacga
      241 tgggttcatg cacattctgg tacacaagga tgagtacgtg gaaatcattt acagctactt
      301 gaagattgat gaggacgatg taatgcaaaa cttcacgatc tactcgaaac gctggggtaa
      361 catcgataat agctactttg agttgcaaac ctttgaaggg gaaatcttca cgggtgaaag
      421 cgacaagatc ctttgtggag tgttgtcatt aggcgactta actacactca agtaaaagct
      481 tgcggccgca ctcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg
      541 aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
      601 ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa
      661 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
      721 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
      781 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
      841 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
      901 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
      961 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     1021 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     1081 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa
     1141 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     1201 tgaattaatt cttagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca
     1261 ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg
     1321 aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca
     1381 tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca
     1441 tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt
     1501 tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc
     1561 attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa
     1621 acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct
     1681 gaatcaggat attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt
     1741 aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc
     1801 gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca
     1861 tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct
     1921 gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa
     1981 tttaatcgcg gcctagagca agacgtttcc cgttgaatat ggctcataac accccttgta
     2041 ttactgttta tgtaagcaga cagttttatt gttcatgacc aaaatccctt aacgtgagtt
     2101 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     2161 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     2221 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     2281 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     2341 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     2401 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     2461 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     2521 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     2581 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     2641 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     2701 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     2761 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     2821 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     2881 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct
     2941 ccttacgcat ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct
     3001 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg
     3061 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     3121 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     3181 catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt
     3241 cacagatgtc tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg
     3301 tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg
     3361 cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga
     3421 tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta
     3481 aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc
     3541 gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga
     3601 tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga
     3661 aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc
     3721 acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta
     3781 gccgggtcct caacgacagg agcacgatca tgcgcacccg tggggccgcc atgccggcga
     3841 taatggcctg cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga
     3901 gggcgtgcaa gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa
     3961 agcggtcctc gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga
     4021 taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc
     4081 tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct
     4141 aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     4201 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg
     4261 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
     4321 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
     4381 atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc
     4441 gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
     4501 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
     4561 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
     4621 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
     4681 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
     4741 agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca
     4801 tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca
     4861 tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc
     4921 gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt
     4981 tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg
     5041 gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg
     5101 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
     5161 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
     5221 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
     5281 tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc
     5341 tcccttatgc gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac
     5401 cgccgccgca aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg
     5461 gcctgccacc atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc
     5521 ttccccatcg gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat
     5581 gccggccacg atgcgtccgg cgtagaggat cgagatctcg
//
