LOCUS       sequence-318553-       10175 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  flaB gRNA expression in B. burgdorferi.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10175)
  AUTHORS   Takacs CN, Scott M, Chang Y, Kloos ZA, Irnov I, Rosa PA, Liu J, 
            Jacobs-Wagner C
  TITLE     A CRISPR interference platform for selective downregulation of gene 
            expression in Borrelia burgdorferi.
  JOURNAL   Appl Environ Microbiol. 2020 Nov 30. pii: AEM.02519-20. doi: 
            10.1128/AEM.02519-20.
  PUBMED    33257311
REFERENCE   2  (bases 1 to 10175)
  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: "Appl 
            Environ Microbiol. 2020 Nov 30. pii: AEM.02519-20. doi: 
            10.1128/AEM.02519-20."
COMMENT     Sequence Label: pBSV2H_Psyn-sgRNAflaB
FEATURES             Location/Qualifiers
     source          1..10175
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1484..1505
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1520..1525,1526..1543,1544..1550)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1558..1574
                     /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)."
     primer_bind     1563..1585
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1582..1598
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1582..1598
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        1634..1668
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        1690..1765
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(1808..1825)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1808..1824)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1817..1839)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             4369..5388
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     CDS             6472..7491
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     rep_origin      7594..8182
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8083..8102
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 actttcaata ataaaatgag gcttaaattt gtaattttcg tatatttttt ggaaacttgt
       61 ttctaattgt ttagaattat attcattttt ttcaaattct ttttgagtgt tgcatagaat
      121 ttcctttaat ttttcttgtt tttctttcat acaagattta gtaaaatgta tatttttatt
      181 ttttagtaat ttaatttcta ttatttttat gattttaagt attttgatta tattatcttt
      241 atcaatattt aaattgaata aaattgtaag aaaaattgaa agaatttttt tacataaaaa
      301 gttacattta ttgaagtaat ttcttacttg atacttttct atttcattaa tctttctttc
      361 ttcttttata ttatttttat tatttaaaca ctcctttaaa tctacactcc catttttagt
      421 aaatctatct ttaaggcccc ttgaaactct attttgaaat cttaaatctt ttttttcttg
      481 aaagtattta tagatttttt ggtaacattc tttttttgaa aaattaagtt gatagtatat
      541 ttcggtcccc aaatttaccc ctaagtgttt gtgataatta gttgtgactt taaattcttt
      601 ttctaatttg taaaggtaat tttgcagtgt ttttagttta acgggtgttt gactatttct
      661 tttaaggttt tcattaaagt aatagagtat gtttttttgt gtgtatttct tatatgtttt
      721 atttatgtac gctagtgttg agataaggac tactaattta tactgtaatt tattgtagca
      781 agttgtggtt ttactggttt tgtttatatt agataagttt tccatttata gctccttgaa
      841 taccggattt tttatctata aagtatatat atcacaattt tgatttaaaa gtaaattctt
      901 tttttaaaaa tacaaaaatt ttttataaat tatatagacc atgatattgt attaatacta
      961 attgatttta atttaaggtt gaactaaaat aaacatagtt ttgtagaaaa taatttttca
     1021 ttattttgct acttgatatt ggatcgtaaa aatattaggg ctttacaaag ttcttttaaa
     1081 agagaattta gtaaagccct aagtctttta ataaaaaatt ttttgtaaaa aagttggcaa
     1141 aaatagtttt tgctatatat ttatatataa gaaaaaattc aatactcaag gagataaaaa
     1201 atggaaaact taaacaataa tccacaaaaa attaatcaaa taaataagca aactaatttt
     1261 attggttttg aaaattttat agaacttgat aatcaagtta atcaaactaa agaaaaaagt
     1321 aaattaagta agataggtaa gaaattaccc ggaataagca gtcaagtcta gtgcggccgc
     1381 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     1441 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     1501 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     1561 tgagcggata acaatttcac acaggaaaca gctatgacat gattacgaat tcgagctcga
     1621 attctaaaga tctttgacag ctagctcagt cctaggtata atactagtga aaatttaaat
     1681 ttctgacttg ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact
     1741 tgaaaaagtg gcaccgagtc ggtgcttttt ttgaagcttg ggcccgaaca aaaactcaaa
     1801 gcttggcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     1861 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     1921 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcggc cgccctaggt
     1981 taatcttcaa taacatgtaa accacgacgt tttaaatctt ctaatgaatc taacattcct
     2041 cttattaatt caacaggatg tccttcccaa tcttcaacaa caccaacaat tcttaaaggt
     2101 tcacaagttc tataagattg tgttggatta ccaggaaatt ttttatttgt aagatttgga
     2161 tcatcttcaa atggtcctgt tggttcaact atatatatat aaccacgacc ttcaagacca
     2221 gatagtgaca tagcactgat gaatccccta atgattttta tcaaaatcat taagttaagg
     2281 tagatacaca tcttgtcata tgatcaaatg gtttcgccaa aaatcaataa tcagacaaca
     2341 aaatgtgcga actcgatatt ttacacgact ctctttacca attctgcccc gaattacact
     2401 taaaacgact caacagctta acgttggctt gccacgcctt acttgactgt aaaactctca
     2461 ctcttaccga acttggccgt aacctgccaa ccaaagcgag aacaaaacat aacatcaaac
     2521 gaatcgaccg attgttaggt aatcgtcacc tccacaaaga gcgactcgct gtataccgtt
     2581 ggcatgctag ctttatctgt tcgggcaata cgatgcccat tgtacttgtt gactggtctg
     2641 atatccgtga gcaaaaacgg cttatggtat tgcgagcttc agtcgcacta cacggtcgtt
     2701 ctgttactct ttatgagaaa gcgttcccgc tttcagagca atgttcaaag aaagctcatg
     2761 accaatttct agccgacctt gcgagcattc taccgagtaa caccacaccg ctcattgtca
     2821 gtgatgctgg ctttaaagtg ccatggtata aatccgttga gaagctgggt tggtactggt
     2881 taagtcgagt aagaggaaaa gtacaatatg cagacctagg agcggaaaac tggaaaccta
     2941 tcagcaactt acatgatatg tcatctagtc actcaaagac tttaggctat aagaggctga
     3001 ctaaaagcaa cccaatctca tgccaaattc tattgtataa atctcgctct aaaggccgaa
     3061 aaaatcagcg ctcgacacgg actcattgtc accacccgtc acctaaaatc tactcagcgt
     3121 cggcaaagga gccatggatt ctagcaacta acttacctgt tgaaattcga acacccaaac
     3181 aacttgttaa tatctattcg aagcgaatgc agattgaaga aaccttccga gacttgaaaa
     3241 gtcctgccta cggactaggc ctacgccata gccgaacgag cagctcagag cgttttgata
     3301 tcatgctgct aatcgccctg atgcttcaac taacatgttg gcttgcgggc gttcatgctc
     3361 agaaacaagg ttgggacaag cacttccagg ctaacacagt cagaaatcga aacgtactct
     3421 caacagttcg cttaggcatg gaagttttgc ggcattctgg ctacacaata acaagggaag
     3481 actcactcgt ggctgcaacc ctgcttactc aaaatctatt cacacatggt tacgttttgg
     3541 ggaaattatg aggggatctc tcaggacata gcaagttcag ctccccaaac tgctggttcc
     3601 attaaagctg aaaaataaat atgtttaaga attcgaccat cttcaaaatg agaaataaat
     3661 cctgtagtta gtaaatcacc aattgctaaa ttagctttag ttccatgata aaatggtcct
     3721 tgtacttgtt tataattatc atgagaaata ggaatccaat cttttaccat gacgttacct
     3781 cctcatgatg tttaactttg ttttagggcg actgccctgc tgcgtaacat cgttgctgct
     3841 ccataacatc aaacatcgac ccacggcgta acgcgcttgc tgcttggatg cccgaggcat
     3901 agactgtacc ccaaaaaaac agtcataaca agccatgcgg ccgtacccga gcttcaagga
     3961 agatttccta ttaaggttga acttaagagc ttaagcatag acgatttgaa aaaaatttta
     4021 aaacaaacaa aaaattcttt aataaaacaa tatgttgcga tgtttaaggt ttatgattta
     4081 gatttaaagt ttagtgagga agctatagat agaattgcag agcttacttt taatatgaat
     4141 cttgagagtg aaaatcttgg tgccagaaga cttcacggtg ttatggaaat agtgcttgca
     4201 gatctttttt ttgaagtgcc tggcagtaag ttgaaaaaat ttgaaataaa cttggactat
     4261 gttaataaaa aaatacaaat taacgaacaa aaagatttga attattatat aatttagtta
     4321 aaagcaattt taaatgaggg aggtttccat catatggata gaagtggtaa acctgaatta
     4381 actgctacta gtgttgaaaa atttttaatt gaaaaatttg atagtgttag tgatttaatg
     4441 caattaagtg aaggtgaaga aagtagagct tttagttttg atgttggtgg tagaggttat
     4501 gttttaagag ttaatagttg tgctgatggt ttttataaag atagatatgt ttatagacat
     4561 tttgctagtg ctgctttacc tattcctgaa gttttagata ttggtgaatt tagtgaaagt
     4621 ttaacttatt gtattagtag aagagcacaa ggtgttactt tacaagattt acctgaaact
     4681 gaattacctg ctgttttaca acctgttgct gaagctatgg atgctattgc ggcggctgat
     4741 ctcagtcaaa cttcgggttt tggtcctttt ggtcctcaag gtattggtca atatactact
     4801 tggagagatt ttatttgtgc tattgctgat cctcatgttt atcattggca aactgttatg
     4861 gatgatactg ttagtgctag tgttgctcaa gcattagatg aattaatgtt atgggctgaa
     4921 gattgtcctg aagttagaca tttagttcat gctgattttg gtagtaataa tgttttaact
     4981 gataatggta gaattactgc tgttattgat tggagtgaag ctatgtttgg tgatagtcaa
     5041 tatgaagttg ctaatatttt tttttggaga ccttggttag cttgtatgga acaacaaact
     5101 agatattttg aaagaagaca tcctgaatta gctggtagtc ctagattaag agcttatatg
     5161 ttaagaattg gtttagatca attatatcaa agtttagttg atggtaattt tgatgatgct
     5221 gcttgggctc aaggtagatg tgatgctatt gttagaagtg gtgctggtac tgttggtaga
     5281 actcaaattg ctagaagaag tgctgctgtt tggactgatg gttgtgttga agttttagct
     5341 gatagtggta atagaagacc tagtactaga cctagagcta aagaataaca attcgttcaa
     5401 gccgagatcg gcttactagg ttaatcttca ataacatgta aaccacgacg ttttaaatct
     5461 tctaatgaat ctaacattcc tcttattaat tcaacaggat gtccttccca atcttcaaca
     5521 acaccaacaa ttcttaaagg ttcacaagtt ctataagatt gtgttggatt accaggaaat
     5581 tttttatttg taagatttgg atcatcttca aatggtcctg ttggttcaac tatatatata
     5641 taaccacgac cttcaagacc agatagtgac atagcaagtt cagctcccca aactgctggt
     5701 tccattaaag ctgaaaaata aatatgttta agaattcgac catcttcaaa atgagaaata
     5761 aatcctgtag ttagtaaatc accaattgct aaattagctt tagttccatg ataaaatggt
     5821 ccttgtactt gtttataatt atcatgagaa ataggaatcc aatcttttac catgacgtta
     5881 cctcctcatg atgtttaact ttgttttagg gcgactgccc tgctgcgtaa catcgttgct
     5941 gctccataac atcaaacatc gacccacggc gtaacgcgct tgctgcttgg atgcccgagg
     6001 catagactgt accccaaaaa aacagtcata acaagccatg cggccgtacc cgagcttcaa
     6061 ggaagatttc ctattaaggt tgaacttaag agcttaagca tagacgattt gaaaaaaatt
     6121 ttaaaacaaa caaaaaattc tttaataaaa caatatgttg cgatgtttaa ggtttatgat
     6181 ttagatttaa agtttagtga ggaagctata gatagaattg cagagcttac ttttaatatg
     6241 aatcttgaga gtgaaaatct tggtgccaga agacttcacg gtgttatgga aatagtgctt
     6301 gcagatcttt tttttgaagt gcctggcagt aagttgaaaa aatttgaaat aaacttggac
     6361 tatgttaata aaaaaataca aattaacgaa caaaaagatt tgaattatta tataatttag
     6421 ttaaaagcaa ttttaaatga gggaggtttc catcatatgg atagaagtgg taaacctgaa
     6481 ttaactgcta ctagtgttga aaaattttta attgaaaaat ttgatagtgt tagtgattta
     6541 atgcaattaa gtgaaggtga agaaagtaga gcttttagtt ttgatgttgg tggtagaggt
     6601 tatgttttaa gagttaatag ttgtgctgat ggtttttata aagatagata tgtttataga
     6661 cattttgcta gtgctgcttt acctattcct gaagttttag atattggtga atttagtgaa
     6721 agtttaactt attgtattag tagaagagca caaggtgtta ctttacaaga tttacctgaa
     6781 actgaattac ctgctgtttt acaacctgtt gctgaagcta tggatgctat tgcggcggct
     6841 gatctcagtc aaacttcggg ttttggtcct tttggtcctc aaggtattgg tcaatatact
     6901 acttggagag attttatttg tgctattgct gatcctcatg tttatcattg gcaaactgtt
     6961 atggatgata ctgttagtgc tagtgttgct caagcattag atgaattaat gttatgggct
     7021 gaagattgtc ctgaagttag acatttagtt catgctgatt ttggtagtaa taatgtttta
     7081 actgataatg gtagaattac tgctgttatt gattggagtg aagctatgtt tggtgatagt
     7141 caatatgaag ttgctaatat ttttttttgg agaccttggt tagcttgtat ggaacaacaa
     7201 actagatatt ttgaaagaag acatcctgaa ttagctggta gtcctagatt aagagcttat
     7261 atgttaagaa ttggtttaga tcaattatat caaagtttag ttgatggtaa ttttgatgat
     7321 gctgcttggg ctcaaggtag atgtgatgct attgttagaa gtggtgctgg tactgttggt
     7381 agaactcaaa ttgctagaag aagtgctgct gtttggactg atggttgtgt tgaagtttta
     7441 gctgatagtg gtaatagaag acctagtact agacctagag ctaaagaata acaattcgtt
     7501 caagccgaga tcggcttacg cgtccaaaat cccttaacgt gagttttcgt tccactgagc
     7561 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     7621 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     7681 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     7741 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     7801 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     7861 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     7921 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     7981 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     8041 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     8101 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     8161 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     8221 ttgctggcct tttgctcaca tgttctttcc tgcactagat ccccggcaag ttcatcctta
     8281 gggttttcat atctcttaaa aagtcaattt ctgcttgatt aatttcttgt ggattattgt
     8341 ttttgcggtt tttcattttt tatctccttg agtattgaat tttttcttat atataaaata
     8401 tagcaaaaac tatttttgcc aactttttta caaaaaattt tttattaaaa cacttagggc
     8461 tttgctaaat tctcttttaa aagaacttag taaagcccta tggatttaag aactgattat
     8521 acttacgtag taaaaatact acagatatta atagtataga ccatatcaaa atcataatca
     8581 atttatattt atctaaaatt gtggtccata caaaatacaa aaaaattagg tttttctaaa
     8641 ttctatttta aaagaactta gtaacaccta atctctaaat tacttacaaa attctaaaat
     8701 tagattttta gttcttcata tttctttaaa agttttttaa gaaaatcttt ttgattttca
     8761 aaaatttcat gcatcataaa actagtaaat ttggattttt ttttgtaaaa atcataactt
     8821 tcttgggttt taagttgaaa tcttaatggt tttattggat tttgcctaga tttttttaac
     8881 gccggacttt ctttatcttt tataaaatta taagaatttt taataccatt ttctattaaa
     8941 taatcttctt ttataagccc ctcttgcagt gcagttgcta atcttaaata cttatacgct
     9001 tgagtttttg ctacatcaaa gtcttttata aaatcaagaa aagttttata accatcaaat
     9061 ttataatatt gattatcttt tatttctttt aaaattttaa tattttccac cctgtggaat
     9121 atttcttttt tgaaattgtc atttaacttt tcctttagtg tcttgtaata agaaatttta
     9181 ctttcattaa tcattaaatt ttgttcaaca taagtatcta tcttttttgt taaaacttta
     9241 acaacacgtt ttttttccat aaaaattttc ctttattttt gcaacatttt ccttaattga
     9301 tataattgtt actattttag attttttaat atccacttaa gaattaatcc ctcactagaa
     9361 tattttttat tatagaattc atatacatgt ttttctaaat ctataaattt atcaacaatt
     9421 gctttattat agggtgtatc tatcttttct tttcttaata atcgtctcat attctttaaa
     9481 taacaacaaa cactgccggt tttaaaccta aattctatgt aatatatttt tgaaaaagta
     9541 tatacatttt caaccccgtt tatttcatac tttacaaaaa cattatttat tggttttcta
     9601 taaccatagt aaatgcctaa aaatttgtcc ccgtctttca taggatatag ataaagtttt
     9661 tgaatctctg atttatctaa taatttttca aaaaaaatta tacaccgctt ttgtttgtta
     9721 ctaactactc tatgcaatgc ctttaatatt tttgtgtgat atatattttt atcacctttt
     9781 ttttctttca gcaggaagat actttctttt ggaaatttaa ttttttgttc atttattctt
     9841 gcagccttaa gtccgatttt catatttaat ccttatatag cctttctatt taattctttt
     9901 aaactcaata aacttttttg gtttttcatt agttctaata attcgcaata atatgtgcaa
     9961 aacactttat tgtattctat tttcttttgg ttattcaaat attcttttat aattggcttt
    10021 agaagttcaa tatttacttc tttttttagt tgttcaataa ggatattgaa aatgtttgtt
    10081 tttaaatctt gataattttg ttgagaattt tctttttttc tttcaattga cttctctaat
    10141 ttacgcttta tattgtttaa atcgctatat ttatg
//
