LOCUS       sequence-451878-       11131 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Group B Streptococcus inducible dCas12 CRISPRi with Modular crRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11131)
  AUTHORS   Hillebrand GH, Carlin SC, Giacobe EJ, Stephenson HA, Collins J, 
            Hooven TA
  TITLE     A Cas12a Toolbox for Rapid and Flexible Group B Streptococcus 
            Genomic Editing and CRISPRi.
  JOURNAL   Mol Microbiol. 2025 Nov;124(5):449-461. doi: 10.1111/mmi.70022. Epub
            2025 Sep 13.
  PUBMED    40944362
REFERENCE   2  (bases 1 to 11131)
  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"; doi: "10.1111/mmi.70022";
            journalName: "Mol Microbiol"; date: "2025-11"; volume: "124"; issue:
            "5"; pages: "449-461"
COMMENT     Sequence Label: pGBScrispri Sham
FEATURES             Location/Qualifiers
     source          1..11131
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(348..365)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(348..364)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(357..379)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_RNA        complement(5203..5281)
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     rep_origin      complement(5404..5992)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5484..5503)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6309..6325
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6309..6325
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(6392..7018)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     6923..6942
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    7125..7143
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
ORIGIN
        1 tatgaccatg attacgaatt cgagctcggt acccggggat cctataagtt ttaataaaac
       61 tttaaataga aaaaggcttc tctcatgaga agtctttttt atttaaaata aatataaaat
      121 aaaatagagg ctataaatag cctctatttt atgtgagaaa tccctaaata aaaagatgcc
      181 agtgtgctgg aattcgtcca ctagtcattc tacagtttat tcttgacatt gcactgtccc
      241 cctggtataa taactataat ttctactctt gtagatgaga cgatcagccg catcgtctca
      301 atttctactc ttgtagatct cgaggcctgc agacatgcaa gcttggcact ggccgtcgtt
      361 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
      421 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
      481 ttgcgcagcc tgaatggcga atggcgctag cataaaaata agaagcctgc atttgcaggc
      541 ttcttatttt tatggcgcgc cgttctgaat ccttagctaa tggttcaaca ggtaactatg
      601 acgaagatag caccctggag caaccaggaa tgaattacta tcccttttat caagaagcgc
      661 acaaaaagaa aaacgaaatg atacaccaat cagtgcaaaa aaagatataa tgggagataa
      721 gacggttcgt gttcgtgctg acttgcacca tatcataaaa atcgaaacag caaagaatgg
      781 cggaaacgta aaagaagtta tggaaataag acttagaagc aaacttaaga gtgtgttgat
      841 agtgcagtat cttaaaattt tgtataatag gaattgaagt taaattagat gctaaaaatt
      901 tgtaattaag aaggagtgat tacatgaaca aaaatataaa atattctcaa aactttttaa
      961 cgagtgaaaa agtactcaac caaataataa aacaattgaa tttaaaagaa accgataccg
     1021 tttacgaaat tggaacaggt aaagggcatt taacgacgaa actggctaaa ataagtaaac
     1081 aggtaacgtc tattgaatta gacagtcatc tattcaactt atcgtcagaa aaattaaaac
     1141 tgaatactcg tgtcacttta attcaccaag atattctaca gtttcaattc cctaacaaac
     1201 agaggtataa aattgttggg agtattcctt accatttaag cacacaaatt attaaaaaag
     1261 tggtttttga aagccatgcg tctgacatct atctgattgt tgaagaagga ttctacaagc
     1321 gtaccttgga tattcaccga acactagggt tgctcttgca cactcaagtc tcgattcagc
     1381 aattgcttaa gctgccagcg gaatgctttc atcctaaacc aaaagtaaac agtgtcttaa
     1441 taaaacttac ccgccatacc acagatgttc cagataaata ttggaagcta tatacgtact
     1501 ttgtttcaaa atgggtcaat cgagaatatc gtcaactgtt tactaaaaat cagtttcatc
     1561 aagcaatgaa acacgccaaa gtaaacaatt taagtaccgt tacttatgag caagtattgt
     1621 ctatttttaa tagttatcta ttatttaacg ggaggaaata attctatgag tcgcttttgt
     1681 aaatttggaa agttacacgt tactaaaggg aatgtagata aattattagg tatactactg
     1741 acagcttcca aggagctaaa gaggtcccta gcgcttagaa tcgctttagg aaacacgatc
     1801 cagtccaata atcgtcgata aaaacttttg aaaaaggttg gtgaaattac ctacttttgg
     1861 aataatcaca aatcacaagt gattaatcac aaatcacaag tgattaatca cttgtttatt
     1921 aagatattaa aagctataat ttaaataaag cgtgaatttt attacacaaa aagagggggg
     1981 agaaacttgg aactagcatt tagagaaagc ttaaaaaaga tgagaggtac caaatcaaaa
     2041 gaaaaattct cccaagaatt agaaatgagt agatcaaatt attcacgaat agaatcagga
     2101 aaatcagatc caaccataaa aacactagaa caaattgcaa agttaactaa ctcaacgcta
     2161 gtagtggatt taatcccaaa tgagccaaca gaaccagaac cagaaacaga atcagaacaa
     2221 gtaacattgg atttagaaat ggaagaagaa aaaagcaatg acttcgtgtg aataatgcgc
     2281 gaaatcgttg cttatttttt ttaaaagcgg tatactagat ataacgaaac aacgaactga
     2341 atagaaacga aaaaagagcc atgacacatt tataaaatgt ttgacgacat tttataaatg
     2401 catagcccga taagattgcc aaaccaacgc ttatcagtta gtcagatgaa ctcttccctc
     2461 gtaagaagtt atttaattaa ctttgtttga agacggtata taaccgtact atcattatat
     2521 agggaaatca gagagttttc aagtatctaa gctactgaat ttaagaattg ttaagcaatc
     2581 aatcggaaat cgtttgattg ctttttttgt attcatttat agaaggtgga gtttgtatga
     2641 atcatgatga atgtaaaact tatataaaaa atagtttatt ggagataaga aaattagcaa
     2701 atatctatac actagaaacg tttaagaaag agttagaaaa gagaaatatc tacttagaaa
     2761 caaaatcaga taagtatttt tcttcggagg gggaagatta tatatataag ttaatagaaa
     2821 ataacaaaat aatttattcg attagtggaa aaaaattgac ttataaagga aaaaaatctt
     2881 tttcaaaaca tgcaatattg aaacagttga atgaaaaagc aaaccaagtt aattaaacaa
     2941 cctattttat aggatttata ggaaaggaga acagctgaat gaatatccct tttgttgtag
     3001 aaactgtgct tcatgacggc ttgttaaagt acaaatttaa aaatagtaaa attcgctcaa
     3061 tcactaccaa gccaggtaaa agcaaagggg ctatttttgc gtatcgctca aaatcaagca
     3121 tgattggcgg tcgtggtgtt gttctgactt ccgaggaagc gattcaagaa aatcaagata
     3181 catttacaca ttggacaccc aacgtttatc gttatggaac gtatgcagac gaaaaccgtt
     3241 catacacgaa aggacattct gaaaacaatt taagacaaat caataccttc tttattgatt
     3301 ttgatattca cacggcaaaa gaaactattt cagcaagcga tattttaaca accgctattg
     3361 atttaggttt tatgcctact atgattatca aatctgataa aggttatcaa gcatattttg
     3421 ttttagaaac gccagtctat gtgacttcaa aatcagaatt taaatctgtc aaagcagcca
     3481 aaataatttc gcaaaatatc cgagaatatt ttggaaagtc tttgccagtt gatctaacgt
     3541 gtaatcattt tggtattgct cgcataccaa gaacggacaa tgtagaattt tttgatccta
     3601 attaccgtta ttctttcaaa gaatggcaag attggtcttt caaacaaaca gataataagg
     3661 gctttactcg ttcaagtcta acggttttaa gcggtacaga aggcaaaaaa caagtagatg
     3721 aaccctggtt taatctctta ttgcacgaaa cgaaattttc aggagaaaag ggtttaatag
     3781 ggcgtaataa cgtcatgttt accctctctt tagcctactt tagttcaggc tattcaatcg
     3841 aaacgtgcga atataatatg tttgagttta ataatcgatt agatcaaccc ttagaagaaa
     3901 aagaagtaat caaaattgtt agaagtgcct attcagaaaa ctatcaaggg gctaataggg
     3961 aatacattac cattctttgc aaagcttggg tatcaagtga tttaaccagt aaagatttat
     4021 ttgtccgtca agggtggttt aaattcaaga aaaaaagaag cgaacgtcaa cgtgttcatt
     4081 tgtcagaatg gaaagaagat ttaatggctt atattagcga aaaatcagat gtatacaagc
     4141 cttatttagt gacgaccaaa aaagagatta gagaagtgct aggcattcct gaacggacat
     4201 tagataaatt gctgaaggta ctgaaggcga atcaggaaat tttctttaag attaaaccag
     4261 gaagaaatgg tggcattcaa cttgctagtg ttaaatcatt gttgctatcg atcattaaag
     4321 taaaaaaaga agaaaaagaa agctatataa aggcgctgac aaattctttt gacttagagc
     4381 atacattcat tcaagagact ttaaacaagc tagcagaacg ccctaaaacg gacacacaac
     4441 tcgatttgtt tagctatgat acaggctgaa aataaaaccc gcactatgcc attacattta
     4501 tatctatgat acgtgtttgt tttttctttg ctgtttagcg aatgattagc agaaatatac
     4561 agagtaagat tttaattaat tattaggggg agaaggagag agtagcccga aaacttttag
     4621 ttggcttgga ctgaacgaag tgagggaaag gctactaaaa cgtcgagggg cagtgagagc
     4681 gaagcgaaca cttgattttt taattttcta tcttttatag gtcattagag tatacttatt
     4741 tgtcctataa actatttagc agcataatag atttattgaa taggtcattt aagttgagca
     4801 tattagagga ggaaaatctt ggagaaatat ttgaagaacc cgattacatg gattggatta
     4861 gttcttgtgg ttacgtggtt tttaactaaa agtagtgaat ttttgatttt tggtgtgtgt
     4921 gtcttgttgt cagtatttgc tagtcaaagt gattaaatag aattctcatg tttgacagct
     4981 tatcatcgga gctccgatga taagctgtca aacatgagaa ttcccgggga tccttacgaa
     5041 atcatcctgt ggagcttagt aggtttagca agatggcagc gcctaaatgt agaatgataa
     5101 aaggattaag agattaattt ccctaaaaat gataaaacaa gcgttttgaa agcgcttgtt
     5161 tttttggttt gcagtcagag tagaatagaa gtatcaaaaa aagcaccgac tcggtgccac
     5221 tttttcaagt tgataacgga ctagccttat tttaacttgc tatgctgtta agaatggttc
     5281 caacgctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5341 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5401 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5461 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5521 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5581 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5641 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5701 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5761 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5821 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     5881 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5941 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     6001 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     6061 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     6121 taaatcaatc taaagtatat atgagtaaag atgtgatccg tagcggtatg cgctccatca
     6181 agaagagcga cttcgcggag ctggtgaagt acatcaccga cgagcaaggc aagaccgatc
     6241 gggccccctg caggataaaa aaattgtaga taaattttat aaaatagttt tatctacaat
     6301 ttttttatca ggaaacagct atgaccgcgg ccgctgtatc cacataaaaa taagaagcct
     6361 gcatttgcag gcttcttatt tttatggtac cttaagaccc actttcacat ttaagttgtt
     6421 tttctaatcc gcatatgatc aattcaaggc cgaataagaa ggctggctct gcaccttggt
     6481 gatcaaataa ttcgatagct tgtcgtaata atggcggcat actatcagta gtaggtgttt
     6541 ccctttcttc tttagcgact tgatgctctt gatcttccaa tacgcaacct aaagtaaaat
     6601 gccccacagc gctgagtgca tataatgcat tctctagtga aaaaccttgt tggcataaaa
     6661 aggctaattg attttcgaga gtttcatact gtttttctgt aggccgtgta cctaaatgta
     6721 cttttgctcc atctcggtga cttagtaaag cacatctaaa acttttagcg ttattacgta
     6781 aaaaatcttg ccagctttcc ccttctaaag ggcaaaagtg agtatggtgc ctatctaaca
     6841 tctcaatggc taaggcgtcg agcaaagccc gcttattttt tacatgccaa tacaatgtag
     6901 gctgctctac acctagcttc tgggcgagtt tacgggttgt taaaccttcg attccgacct
     6961 cattaagcag ctctaatgcg ctgttaatca ctttactttt atctaatcta gacatcatta
     7021 attcctcctt tttgttgaca ttatatcatt gatagagtta tttgtcaaac tagtttttta
     7081 tttcgatgcc ctggacttca tgaaaaacta aaaaaaatat tgacactcta tcattgatag
     7141 agtataatta aaataagctt gatcgtagcg ttaacagatc tgagctcctg cagtaaagga
     7201 ggtttttatg acacagtttg aaggcttcac caatctctac caggtcagca agacgctacg
     7261 ttttgagctt atcccgcagg gaaaaaccct gaaacacatt caggaacagg ggttcataga
     7321 ggaagataag gcgcgtaacg accattataa agaactgaag cctataatcg accgtattta
     7381 taaaacgtac gcggatcagt gcctgcagct ggttcagctg gattgggaga atctgtccgc
     7441 ggctattgat agctatcgca aagagaagac cgaggaaacc cgtaacgcac tgattgaaga
     7501 gcaggcgacc tatcggaatg cgatccatga ttacttcatc ggccgcaccg acaacctgac
     7561 cgatgcaatt aacaaacgtc acgcagagat ttacaaaggt ctgtttaaag cagagttatt
     7621 caatggcaag gttctgaaac agctgggtac ggtcaccacc accgaacacg aaaacgcact
     7681 gctgaggagc tttgataaat ttaccacata tttcagcggt ttctatgaaa atcgtaagaa
     7741 tgtatttagc gccgaagata tttccaccgc aattcctcat cgtattgtgc aggataattt
     7801 tccgaagttt aaagaaaatt gtcatatttt tacccgtctg atcaccgcgg taccgagcct
     7861 gcgagagcat tttgaaaacg ttaagaaagc cattggaatt tttgtcagta ccagcattga
     7921 agaagtgttt tcgttcccgt tctataacca actgctgacc cagacccaga ttgatctgta
     7981 caatcagctg ctggggggca taagccgcga ggcaggtacc gaaaagataa agggactcaa
     8041 tgaggtgctg aatctggcaa ttcagaagaa tgatgaaacg gctcatatca ttgctagcct
     8101 gccgcatcgt ttcattcccc tgtttaagca aatcctgagc gatcgcaata cactgagctt
     8161 tatcctcgaa gagtttaaat cggacgaaga agttatccag agcttttgca aatacaaaac
     8221 cctgctgcgg aacgaaaatg tgctggagac cgctgaagca ctgtttaatg aactgaactc
     8281 gatcgacctc acccatattt ttatatccca caaaaaactg gaaaccataa gcagcgctct
     8341 gtgtgaccat tgggataccc tgcgcaacgc cctgtatgaa cggcgtatca gcgagctgac
     8401 cgggaaaatc accaaatccg caaaggaaaa agttcagcgt agtctgaaac acgaggacat
     8461 caacctgcaa gaaattatta gcgcagcagg taaagagctg agcgaagcat tcaaacagaa
     8521 aaccagcgaa atcctgagcc atgcccatgc tgcactggat cagccgctgc cgaccaccct
     8581 gaaaaaacag gaggaaaagg agattctgaa aagccaactg gacagcctgc tgggcctgta
     8641 tcacctgctg gactggtttg cagtcgatga gagcaacgag gttgatcctg agttctccgc
     8701 tcgtctgacc ggaatcaagc tggagatgga accgagtctg tcgttttaca ataaagcgcg
     8761 taattacgcg accaagaaac cgtatagcgt ggaaaaattc aaactgaact ttcagatgcc
     8821 gacccttgca agcggatggg acgttaacaa agaaaaaaac aatggggcaa ttctgtttgt
     8881 gaaaaatggc ctctattatc tgggtatcat gccgaaacag aaagggcgct acaaagccct
     8941 gtcatttgag ccgaccgaga aaacctcaga gggtttcgac aagatgtact acgattattt
     9001 cccggatgcg gcaaaaatga tacccaaatg tagcacccaa ctgaaggcag ttacagccca
     9061 ctttcagacc cataccaccc cgatcctgct gtcgaacaat tttatagagc cgctggaaat
     9121 taccaaagag atttatgatc tgaataatcc ggaaaaggag cccaagaaat ttcagacggc
     9181 gtatgcaaaa aagaccgggg atcagaaagg ttatcgtgaa gcgctgtgca aatggattga
     9241 ctttacccgt gactttctgt caaaatatac caaaacgacg agcattgatc tgagcagcct
     9301 acgtccgagc agccaatata aggatctggg cgaatattac gccgaactga atccgctgct
     9361 ctaccatatt tccttccaac gaatcgctga aaaagaaata atggacgccg ttgaaaccgg
     9421 caaactgtat ctgtttcaaa tctacaacaa agatttcgcc aaaggccatc acggtaagcc
     9481 gaacctgcat accctgtatt ggaccggtct gtttagcccg gagaatctgg ccaaaaccag
     9541 catcaagctg aacggacagg cagaactgtt ttaccgcccc aaaagccgta tgaaaaggat
     9601 ggcacaccgc ctgggcgaaa aaatgctgaa taagaaactc aaagatcaga aaacgccgat
     9661 accggatacc ctttatcagg agctgtatga ttatgttaac caccggctga gccatgacct
     9721 gagcgacgaa gcgcgtgcac tgctgccgaa cgtgattacc aaggaagtct cgcatgaaat
     9781 tattaaagat cggcgcttca ccagtgataa atttttcttc catgtaccga tcaccctgaa
     9841 ttatcaagcc gcaaatagcc cttccaaatt taatcaacgc gtgaatgcgt acctgaaaga
     9901 gcatccggag accccaatta ttggcataga ccgaggagaa cgcaatctca tttatatcac
     9961 cgtcattgat agcaccggta agatcctgga acagcgtagc ctgaatacca ttcagcagtt
    10021 tgactaccag aaaaagctgg acaacagaga aaaggaacgt gtagccgccc ggcaggcttg
    10081 gagtgtggtg ggtactatca aggatctgaa gcaggggtat ctctcccaag ttatccatga
    10141 aattgtcgat ctaatgattc actatcaagc agtagtggta ctggcaaatc tgaatttcgg
    10201 tttcaaaagc aaacgtacag ggatcgctga aaaagccgtt tatcagcagt tcgagaaaat
    10261 gctgatagac aagctgaatt gcctggttct gaaagattat ccggcagaga aggtgggcgg
    10321 tgtgctgaac ccgtaccagc tgactgatca atttacgagc tttgcaaaaa tgggaacgca
    10381 gagcggtttc ctgttctatg ttccggcgcc atataccagc aagatagacc cgctgacagg
    10441 tttcgtagat ccgtttgtct ggaaaaccat taaaaatcat gaaagtcgca aacattttct
    10501 ggagggcttt gattttctgc actatgacgt gaaaaccggc gacttcattc tgcattttaa
    10561 aatgaaccgt aatctgtcct ttcagcgcgg cctgcctggc tttatgccgg cgtgggacat
    10621 tgtttttgaa aagaatgaga cacagtttga tgccaaaggt acccccttta ttgcggggaa
    10681 acgcattgtg cccgttatag aaaatcaccg cttcaccgga cggtataggg acttgtaccc
    10741 ggcaaatgaa ttgatagcgc tgctggagga gaaaggtatt gtctttcggg atggatcaaa
    10801 catcctgccg aagctgctgg agaacgatga cagccacgca atagacacca tggtagcgct
    10861 gatccgaagc gtgctgcaga tgcgtaacag taatgcggct acgggggaag actacattaa
    10921 tagcccggtc cgtgatctga acggcgtttg tttcgatagc agatttcaaa atccggagtg
    10981 gccgatggat gccgatgcca atggagctta ccatatcgct ctcaaaggtc agctcctact
    11041 gaaccatttg aaagaatcaa aagatctgaa actgcagaac ggcatctcga atcaggactg
    11101 gctggcctac attcaagaac tgagaaacta a
//
