LOCUS       sequence-409777-        8626 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AcrIIC1-mediated inhibition of GeoCas9 cleavage activity in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8626)
  TITLE     van der Oost E. coli CRISPR plasmids
REFERENCE   2  (bases 1 to 8626)
  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"
COMMENT     Sequence Label: pAcrGCas9_scr
FEATURES             Location/Qualifiers
     source          1..8626
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(85..187)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(579..596)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(713..1258)
                     /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(793..812)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1291..1409
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     protein_bind    5091..5115
                     /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        complement(join(5122..5127,5128..5144,5145..5150))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(5313..5332)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        5538..5615
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5616..6698
                     /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     complement(5635..5654)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        6759..6793
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     CDS             complement(join(8051..8626,1..84))
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     join(8625..8626,1..18)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat caacggtggt
       61 atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc tcgataactc
      121 aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac ctcttacgtg
      181 ccgatcaacg tctcattttc gccaaaagtt ggcccagggc ttcccggtat caacagggac
      241 accaggattt atttattctg cgaagtgatc ttccgtcaca ggtatttatt cggcgcaaag
      301 tgcgtcgggt gatgctgcca acttactgat ttagtgtatg atggtgtttt tgaggtgctc
      361 cagtggcttc tgtttctatc agctgtccct cctgttcagc tactgacggg gtggtgcgta
      421 acggcaaaag caccgccgga catcagcgct agcggagtgt atactggctt actatgttgg
      481 cactgatgag ggtgtcagtg aagtgcttca tgtggcagga gaaaaaaggc tgcaccggtg
      541 cgtcagcaga atatgtgata caggatatat tccgcttcct cgctcactga ctcgctacgc
      601 tcggtcgttc gactgcggcg agcggaaatg gcttacgaac ggggcggaga tttcctggaa
      661 gatgccagga agatacttaa cagggaagtg agagggccgc ggcaaagccg tttttccata
      721 ggctccgccc ccctgacaag catcacgaaa tctgacgctc aaatcagtgg tggcgaaacc
      781 cgacaggact ataaagatac caggcgtttc cccctggcgg ctccctcgtg cgctctcctg
      841 ttcctgcctt tcggtttacc ggtgtcattc cgctgttatg gccgcgtttg tctcattcca
      901 cgcctgacac tcagttccgg gtaggcagtt cgctccaagc tggactgtat gcacgaaccc
      961 cccgttcagt ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggaa
     1021 agacatgcaa aagcaccact ggcagcagcc actggtaatt gatttagagg agttagtctt
     1081 gaagtcatgc gccggttaag gctaaactga aaggacaagt tttggtgact gcgctcctcc
     1141 aagccagtta cctcggttca aagagttggt agctcagaga accttcgaaa aaccgccctg
     1201 caaggcggtt ttttcgtttt cagagcaaga gattacgcgc agaccaaaac gatctcaaga
     1261 agatcatctt attaatcaga taaaatattt caccacaatt cagcaaattg tgaacatcat
     1321 cacgttcatc tttccctggt tgccaatggc ccattttcct gtcagtaacg agaaggtcgc
     1381 gaattcaggc gctttttaga ctggtcgtag ggagaccaca acggtttccc tctagaaata
     1441 attttgttta actataagaa ggagatatac atatgaataa aacatataaa atcggaaaaa
     1501 atgcgggata tgacggctgc gggttatgtt tagccgcgat ctcagaaaat gaagctatta
     1561 aagttaagta tctgcgcgat atctgtccag actatgacgg cgacgacaaa gctgaggact
     1621 ggctgagatg gggaacggat agccgcgtga aagctgcggc tttagaaatg gagcaatatg
     1681 cttatacgtc ggttgggatg gcgtcatgtt gggagtttgt tgaactatga gtcgacagga
     1741 ttaaacaaaa atggcccgct tcataagcag gccattttgt ctgatttagt cacgagtaga
     1801 ttgcagtgga cgaatggttt tgccaggttt gctgtgtgcg gaggacgcca gaccgacacg
     1861 tttctcgcca cgcactttat agatgttacc cagaacatca acttgatatt tctcgaaacg
     1921 tttcagggtg cgagagccaa cgccacgcag agagaaacgg tggtcgtgag aaatcagttc
     1981 caggccgccg tttgcggagt cgatcgtttt atagtaaacg aacacgtctt tcacgttaat
     2041 ttcttcgcct gcagcggttt taacggtttt ttcacgcggc agttcgatgc ggatcaggtc
     2101 gttcggatac aggctgaaac gaaacgtgta atcttcggtc atttctttcc attcagagta
     2161 cggcttgttc ggttcaatcg ctttgttcgg cagaatgcct ttcatgatgt ccatggtata
     2221 caccggtaca cagtagtatt taccatcttt ttcgaacacg tccacgcgaa cgatgttgga
     2281 attgtaggcc acggttttac catcgttcag agggatcacc tggtttttcg tgtcgataat
     2341 ttttacagta cggattaccg ggcccggttc gccgtttttt ttcggtttgt acagaggctc
     2401 ctggaatgcc tttttcgggt cgttgttatg ctccagcaga cgctggcgga tggcttcata
     2461 agtgcgtgga tcggattctt taccgtacat tgggaagtgg ccggatgcat ccagtttaat
     2521 ctcggacagt ttggttttta ctacggtctg gattttaccg gagcgttcgt cgatgcccac
     2581 gtaacggcgc agcgtttcct ggtgcgcagc gccagtaaca gaacgtttcg gcatacggct
     2641 gacaaacacc ggctgcagag attccagttt ttggtcatcg tagttaccca ggttcagagc
     2701 tttgatgctt tctttaggat gtttggacag acgagcacgc agttcgtccg cgaagtgcgg
     2761 ccacggctgc ggaaaatgcg gttcggtttt tttggccagt tccttgttct gctcacgacg
     2821 ctggtagaat gcggtcacct tagcgatatc gcttggggtg gtgcacgcca caataacggc
     2881 gtcaacagcg tggtgcagat cgctttcctc acggttcttg ttgaactccc agcgagaacg
     2941 caggtgggcg gtaacacggc cgtttacggt ataaactttc tgtttatcgt cggattccgc
     3001 aaatttcagg tgttcacgaa tgaagtttgc gaagaagcgg ctgatgtagc gagtatcgtt
     3061 caggttacgg tttttgaatt cagtctcttc gttttcatcg tagtgcagac gcagcagacg
     3121 gtcacgtttc tttttagaga actgtttatt ggtcaggacg aaggtttcga actgttgcca
     3181 acgctcagtg ccaacaccca gatattcagc tgggatgcgg ttgcctttct cacggttttc
     3241 gcgagtcaga accagaactt tgttggtata agaatcatcc agagagcggg agtacgggat
     3301 cacatgatct acttcaacgt aacctggctc cagcagacgt tcgatctcga tcggctgcag
     3361 agagtaagca cagcgaccgt tctgttcaga ccacagcttg aacttcacaa tatcgtgacc
     3421 agtagggttc agagtcagac cgtattccat cagctggcgg atcgcggttt cgtttttttt
     3481 acggttttca tcctgttcct ttttagtttt acgacgctcg tcaaaagtct ggctcaggtc
     3541 gcgcgccagt tcgatatgga tggaaaccgg gctgccgtac tttttgatga tcgcgttgac
     3601 tactttgcgc gcctgggtca gtgcacgcat tactacaggg ttcgcaatcg gcgggatgtt
     3661 cggcagcagc atagtttttt gcttcttttt cggaccggtg aaggtgtagc ctgcacgttc
     3721 acaagcggag gagtagactt caccctgttc catatacggc aggatggaac gcagagcttt
     3781 cagagacagg tgaccgaatt tagtgaagga caggttcagc agttcttcaa tcagctcgtt
     3841 atcgtaaact ttgttagcca ggttaggcat acgtttgccg ttctgttcgt actcgttgcg
     3901 caggtaagag tggatatccg catcgtcttt aaacagggtc agcgcgtaac cgaaggtatc
     3961 aaagtcgatc ggcaggaagc tgctggattt acccttgccg taaactttat ctacagcttt
     4021 acggatctgg tggtatgcat ccagttccag gaaacgaatg ttttcgttct gcttacgaga
     4081 ttcaccgcga tcgtaaacga tgcccttgaa gtaggtgtcg tccggcagat gcagcagggt
     4141 acgaatatcg tggtaggtaa ttttgttttt ctggaatgct tgttcataca gcagacgacg
     4201 ttcttcgtcg gtcaggccgc gggcgccaga cggagaaatc agacgcagtt tgttgatatg
     4261 ttcccacgcg ataaaagact ggaaggtata ggtggctttc ggcgcacgtt tttccttcgg
     4321 ttcgaaagta caaaagccta cttttttttc gatatcatct ttagacgcca ccggacgctg
     4381 agatgcccaa atggtaatat attcgttttc gaattcttcg gtgcaagaca tgttgccgaa
     4441 ttcgcgctgt ttggagaaaa tcagacggat ttcacgttcc aggtcatcgc gtgcaatggt
     4501 gttggtgtag ttttcgcctt tgttacgctt atgcagtgcg aatttcgggt ctttaacaat
     4561 catttcgccc acggtacggt agctagacag aattgcacgg ttctcctcaa tatgtttcag
     4621 catagtggag ttctctttat tggagcgttc agatttacgg ttagatttga agccacgacg
     4681 ctttgccaga tgcagcagga cacgcgccag ttcgtcgttg ttcagcttac ggtccagggc
     4741 ttctacacgc agctgccata cgtcgatctc gtgtttttct tcgaacagtt tatccagttc
     4801 ttctttcgtc aggatgcctt cacgaataac caggcgacga atacgctcca gacggtgttt
     4861 acgacgacgc aggcggcggc gtgcgctacg tgccagacga cgcggcagag ccagagattc
     4921 accggtctgc gggttctccg cacggtcaaa aatgcgcacg cccagatctt cgatacgagg
     4981 gatatccagg ttcatgactg cccaaccaac agaggtaata ccgatgtcca ggccaatctt
     5041 ataacgcata cacggtgcct gactgcgtta gcaatttaac tgtgataaac ggaattgtta
     5101 tccgctcaca attcctaccg cattaaagct tatcgatgat aagctgtcaa acatgagaat
     5161 tacaacttat atcgtatggg gctgacttca ggtgctacat ttgaagagat aaattgcact
     5221 gaaatctaga tttcgcggga tcgagatctc gatcctctac gccggacgca tcgtggccgg
     5281 catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga
     5341 agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg
     5401 ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc
     5461 ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg
     5521 agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat
     5581 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
     5641 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
     5701 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
     5761 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
     5821 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
     5881 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
     5941 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
     6001 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
     6061 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
     6121 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
     6181 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc
     6241 aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa
     6301 ccatgcaaat gctgaatgag ggtatcgttc ccactgcgat gctggttgcc aacgatcaga
     6361 tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct
     6421 cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca
     6481 tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc
     6541 agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca
     6601 ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     6661 tggcacgaca ggtttcccga ctggaaagcg ggcagtgatc agaatcgact gatggaccaa
     6721 aaaaccccgc cgaagcgggg actttgaact agcatacatt gacagctagc tcagtcctag
     6781 gtataatgct agcctagatc cgcagtaacc ccatgggtca tagttcccct gagattatcg
     6841 ctgtggtata atgaaagtta taccacagca atgatctcag ggttactatg ataagggctt
     6901 tctgcctata ggcagactga cccgtggcgt tggggatcgc ctatcgcccg ctttcttcgg
     6961 gcattcccca ctcttaggcg aacgaagacc ttggcggaaa acgtcaaggt cttttttatg
     7021 gaagccggcg gcacctcgct aacggattca ccactccaag aattggagcc aatcaattct
     7081 tgcggagaac tgtgaatgcg caaaccaacc cttggcagaa catatccatc gcgtccgcca
     7141 tctccagcag ccgcacgcgg cgcatctcgg gcagcgttgg gtcctggcca cgggtgcgca
     7201 tgatcgtgct cctgtcgttg aggacccggc taggctggcg gggttgcctt actggttagc
     7261 agaatgaatc accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga
     7321 cctgagcaac aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga
     7381 agtcccctac gtgctgctga agttgcccgc aacagagagt ggaaccaacc ggtgatacca
     7441 cgatactatg actgagagtc aacgccatga gcggcctcat ttcttattct gagttacaac
     7501 agtccgcacc gctgccggta gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc
     7561 acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag cgcccaacag
     7621 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgccct gcaccattat
     7681 gttccggatc tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt
     7741 aacgaagcgc taaccgtttt tatcaggctc tgggaggcag aataaatgat catatcgtca
     7801 attattacct ccacggggag agcctgagca aactggcctc aggcatttga gaagcacacg
     7861 gtcacactgc ttccggtagt caataaaccg gtaaaccagc aatagacata agcggctatt
     7921 taacgaccct gccctgaacc gacgaccggg tcgaatttgc tttcgaattt ctgccattca
     7981 tccgcttatt atcacttatt caggcgtagc aaccaggcgt ttaagggcac caataactgc
     8041 cttaaaaaaa ttacgccccg ccctgccact catcgcagta ctgttgtaat tcattaagca
     8101 ttctgccgac atggaagcca tcacaaacgg catgatgaac ctgaatcgcc agcggcatca
     8161 gcaccttgtc gccttgcgta taatatttgc ccatggtgaa aacgggggcg aagaagttgt
     8221 ccatattggc cacgtttaaa tcaaaactgg tgaaactcac ccagggattg gctgagacga
     8281 aaaacatatt ctcaataaac cctttaggga aataggccag gttttcaccg taacacgcca
     8341 catcttgcga atatatgtgt agaaactgcc ggaaatcgtc gtggtattca ctccagagcg
     8401 atgaaaacgt ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca ctatcccata
     8461 tcaccagctc accgtctttc attgccatac ggaattccgg atgagcattc atcaggcggg
     8521 caagaatgtg aataaaggcc ggataaaact tgtgcttatt tttctttacg gtctttaaaa
     8581 aggccgtaat atccagctga acggtctggt tataggtaca ttgagc
//
