LOCUS       sequence-318346-        8837 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  rodA gRNA expression in B. burgdorferi.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8837)
  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 8837)
  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-sgRNArodA
FEATURES             Location/Qualifiers
     source          1..8837
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      71..659
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     560..579
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    4136..4157
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4172..4177,4178..4195,4196..4202)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4210..4226
                     /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     4215..4237
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4234..4250
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4234..4250
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        4286..4320
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        4342..4417
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(4460..4477)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4460..4476)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4469..4491)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             5683..6702
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     CDS             7786..8805
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
ORIGIN
        1 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
       61 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
      121 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
      181 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
      241 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
      301 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
      361 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
      421 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
      481 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
      541 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
      601 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
      661 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
      721 tctttcctgc actagatccc cggcaagttc atccttaggg ttttcatatc tcttaaaaag
      781 tcaatttctg cttgattaat ttcttgtgga ttattgtttt tgcggttttt cattttttat
      841 ctccttgagt attgaatttt ttcttatata taaaatatag caaaaactat ttttgccaac
      901 ttttttacaa aaaatttttt attaaaacac ttagggcttt gctaaattct cttttaaaag
      961 aacttagtaa agccctatgg atttaagaac tgattatact tacgtagtaa aaatactaca
     1021 gatattaata gtatagacca tatcaaaatc ataatcaatt tatatttatc taaaattgtg
     1081 gtccatacaa aatacaaaaa aattaggttt ttctaaattc tattttaaaa gaacttagta
     1141 acacctaatc tctaaattac ttacaaaatt ctaaaattag atttttagtt cttcatattt
     1201 ctttaaaagt tttttaagaa aatctttttg attttcaaaa atttcatgca tcataaaact
     1261 agtaaatttg gatttttttt tgtaaaaatc ataactttct tgggttttaa gttgaaatct
     1321 taatggtttt attggatttt gcctagattt ttttaacgcc ggactttctt tatcttttat
     1381 aaaattataa gaatttttaa taccattttc tattaaataa tcttctttta taagcccctc
     1441 ttgcagtgca gttgctaatc ttaaatactt atacgcttga gtttttgcta catcaaagtc
     1501 ttttataaaa tcaagaaaag ttttataacc atcaaattta taatattgat tatcttttat
     1561 ttcttttaaa attttaatat tttccaccct gtggaatatt tcttttttga aattgtcatt
     1621 taacttttcc tttagtgtct tgtaataaga aattttactt tcattaatca ttaaattttg
     1681 ttcaacataa gtatctatct tttttgttaa aactttaaca acacgttttt tttccataaa
     1741 aattttcctt tatttttgca acattttcct taattgatat aattgttact attttagatt
     1801 ttttaatatc cacttaagaa ttaatccctc actagaatat tttttattat agaattcata
     1861 tacatgtttt tctaaatcta taaatttatc aacaattgct ttattatagg gtgtatctat
     1921 cttttctttt cttaataatc gtctcatatt ctttaaataa caacaaacac tgccggtttt
     1981 aaacctaaat tctatgtaat atatttttga aaaagtatat acattttcaa ccccgtttat
     2041 ttcatacttt acaaaaacat tatttattgg ttttctataa ccatagtaaa tgcctaaaaa
     2101 tttgtccccg tctttcatag gatatagata aagtttttga atctctgatt tatctaataa
     2161 tttttcaaaa aaaattatac accgcttttg tttgttacta actactctat gcaatgcctt
     2221 taatattttt gtgtgatata tatttttatc accttttttt tctttcagca ggaagatact
     2281 ttcttttgga aatttaattt tttgttcatt tattcttgca gccttaagtc cgattttcat
     2341 atttaatcct tatatagcct ttctatttaa ttcttttaaa ctcaataaac ttttttggtt
     2401 tttcattagt tctaataatt cgcaataata tgtgcaaaac actttattgt attctatttt
     2461 cttttggtta ttcaaatatt cttttataat tggctttaga agttcaatat ttacttcttt
     2521 ttttagttgt tcaataagga tattgaaaat gtttgttttt aaatcttgat aattttgttg
     2581 agaattttct ttttttcttt caattgactt ctctaattta cgctttatat tgtttaaatc
     2641 gctatattta tgactttcaa taataaaatg aggcttaaat ttgtaatttt cgtatatttt
     2701 ttggaaactt gtttctaatt gtttagaatt atattcattt ttttcaaatt ctttttgagt
     2761 gttgcataga atttccttta atttttcttg tttttctttc atacaagatt tagtaaaatg
     2821 tatattttta ttttttagta atttaatttc tattattttt atgattttaa gtattttgat
     2881 tatattatct ttatcaatat ttaaattgaa taaaattgta agaaaaattg aaagaatttt
     2941 tttacataaa aagttacatt tattgaagta atttcttact tgatactttt ctatttcatt
     3001 aatctttctt tcttctttta tattattttt attatttaaa cactccttta aatctacact
     3061 cccattttta gtaaatctat ctttaaggcc ccttgaaact ctattttgaa atcttaaatc
     3121 ttttttttct tgaaagtatt tatagatttt ttggtaacat tctttttttg aaaaattaag
     3181 ttgatagtat atttcggtcc ccaaatttac ccctaagtgt ttgtgataat tagttgtgac
     3241 tttaaattct ttttctaatt tgtaaaggta attttgcagt gtttttagtt taacgggtgt
     3301 ttgactattt cttttaaggt tttcattaaa gtaatagagt atgttttttt gtgtgtattt
     3361 cttatatgtt ttatttatgt acgctagtgt tgagataagg actactaatt tatactgtaa
     3421 tttattgtag caagttgtgg ttttactggt tttgtttata ttagataagt tttccattta
     3481 tagctccttg aataccggat tttttatcta taaagtatat atatcacaat tttgatttaa
     3541 aagtaaattc tttttttaaa aatacaaaaa ttttttataa attatataga ccatgatatt
     3601 gtattaatac taattgattt taatttaagg ttgaactaaa ataaacatag ttttgtagaa
     3661 aataattttt cattattttg ctacttgata ttggatcgta aaaatattag ggctttacaa
     3721 agttctttta aaagagaatt tagtaaagcc ctaagtcttt taataaaaaa ttttttgtaa
     3781 aaaagttggc aaaaatagtt tttgctatat atttatatat aagaaaaaat tcaatactca
     3841 aggagataaa aaatggaaaa cttaaacaat aatccacaaa aaattaatca aataaataag
     3901 caaactaatt ttattggttt tgaaaatttt atagaacttg ataatcaagt taatcaaact
     3961 aaagaaaaaa gtaaattaag taagataggt aagaaattac ccggaataag cagtcaagtc
     4021 tagtgcggcc gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     4081 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     4141 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     4201 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac atgattacga
     4261 attcgagctc gaattctaaa gatctttgac agctagctca gtcctaggta taatactagt
     4321 gttaatctac ctaatatacc agttttagag ctagaaatag caagttaaaa taaggctagt
     4381 ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt ttttgaagct tgggcccgaa
     4441 caaaaactca aagcttggca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     4501 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     4561 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcg
     4621 gccgccctag gttaatcttc aataacatgt aaaccacgac gttttaaatc ttctaatgaa
     4681 tctaacattc ctcttattaa ttcaacagga tgtccttccc aatcttcaac aacaccaaca
     4741 attcttaaag gttcacaagt tctataagat tgtgttggat taccaggaaa ttttttattt
     4801 gtaagatttg gatcatcttc aaatggtcct gttggttcaa ctatatatat ataaccacga
     4861 ccttcaagac cagatagtga catagcaagt tcagctcccc aaactgctgg ttccattaaa
     4921 gctgaaaaat aaatatgttt aagaattcga ccatcttcaa aatgagaaat aaatcctgta
     4981 gttagtaaat caccaattgc taaattagct ttagttccat gataaaatgg tccttgtact
     5041 tgtttataat tatcatgaga aataggaatc caatctttta ccatgacgtt acctcctcat
     5101 gatgtttaac tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctccataa
     5161 catcaaacat cgacccacgg cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg
     5221 taccccaaaa aaacagtcat aacaagccat gcggccgtac ccgagcttca aggaagattt
     5281 cctattaagg ttgaacttaa gagcttaagc atagacgatt tgaaaaaaat tttaaaacaa
     5341 acaaaaaatt ctttaataaa acaatatgtt gcgatgttta aggtttatga tttagattta
     5401 aagtttagtg aggaagctat agatagaatt gcagagctta cttttaatat gaatcttgag
     5461 agtgaaaatc ttggtgccag aagacttcac ggtgttatgg aaatagtgct tgcagatctt
     5521 ttttttgaag tgcctggcag taagttgaaa aaatttgaaa taaacttgga ctatgttaat
     5581 aaaaaaatac aaattaacga acaaaaagat ttgaattatt atataattta gttaaaagca
     5641 attttaaatg agggaggttt ccatcatatg gatagaagtg gtaaacctga attaactgct
     5701 actagtgttg aaaaattttt aattgaaaaa tttgatagtg ttagtgattt aatgcaatta
     5761 agtgaaggtg aagaaagtag agcttttagt tttgatgttg gtggtagagg ttatgtttta
     5821 agagttaata gttgtgctga tggtttttat aaagatagat atgtttatag acattttgct
     5881 agtgctgctt tacctattcc tgaagtttta gatattggtg aatttagtga aagtttaact
     5941 tattgtatta gtagaagagc acaaggtgtt actttacaag atttacctga aactgaatta
     6001 cctgctgttt tacaacctgt tgctgaagct atggatgcta ttgcggcggc tgatctcagt
     6061 caaacttcgg gttttggtcc ttttggtcct caaggtattg gtcaatatac tacttggaga
     6121 gattttattt gtgctattgc tgatcctcat gtttatcatt ggcaaactgt tatggatgat
     6181 actgttagtg ctagtgttgc tcaagcatta gatgaattaa tgttatgggc tgaagattgt
     6241 cctgaagtta gacatttagt tcatgctgat tttggtagta ataatgtttt aactgataat
     6301 ggtagaatta ctgctgttat tgattggagt gaagctatgt ttggtgatag tcaatatgaa
     6361 gttgctaata tttttttttg gagaccttgg ttagcttgta tggaacaaca aactagatat
     6421 tttgaaagaa gacatcctga attagctggt agtcctagat taagagctta tatgttaaga
     6481 attggtttag atcaattata tcaaagttta gttgatggta attttgatga tgctgcttgg
     6541 gctcaaggta gatgtgatgc tattgttaga agtggtgctg gtactgttgg tagaactcaa
     6601 attgctagaa gaagtgctgc tgtttggact gatggttgtg ttgaagtttt agctgatagt
     6661 ggtaatagaa gacctagtac tagacctaga gctaaagaat aacaattcgt tcaagccgag
     6721 atcggcttac gcggttaatc ttcaataaca tgtaaaccac gacgttttaa atcttctaat
     6781 gaatctaaca ttcctcttat taattcaaca ggatgtcctt cccaatcttc aacaacacca
     6841 acaattctta aaggttcaca agttctataa gattgtgttg gattaccagg aaatttttta
     6901 tttgtaagat ttggatcatc ttcaaatggt cctgttggtt caactatata tatataacca
     6961 cgaccttcaa gaccagatag tgacatagca agttcagctc cccaaactgc tggttccatt
     7021 aaagctgaaa aataaatatg tttaagaatt cgaccatctt caaaatgaga aataaatcct
     7081 gtagttagta aatcaccaat tgctaaatta gctttagttc catgataaaa tggtccttgt
     7141 acttgtttat aattatcatg agaaatagga atccaatctt ttaccatgac gttacctcct
     7201 catgatgttt aactttgttt tagggcgact gccctgctgc gtaacatcgt tgctgctcca
     7261 taacatcaaa catcgaccca cggcgtaacg cgcttgctgc ttggatgccc gaggcataga
     7321 ctgtacccca aaaaaacagt cataacaagc catgcggccg tacccgagct tcaaggaaga
     7381 tttcctatta aggttgaact taagagctta agcatagacg atttgaaaaa aattttaaaa
     7441 caaacaaaaa attctttaat aaaacaatat gttgcgatgt ttaaggttta tgatttagat
     7501 ttaaagttta gtgaggaagc tatagataga attgcagagc ttacttttaa tatgaatctt
     7561 gagagtgaaa atcttggtgc cagaagactt cacggtgtta tggaaatagt gcttgcagat
     7621 cttttttttg aagtgcctgg cagtaagttg aaaaaatttg aaataaactt ggactatgtt
     7681 aataaaaaaa tacaaattaa cgaacaaaaa gatttgaatt attatataat ttagttaaaa
     7741 gcaattttaa atgagggagg tttccatcat atggatagaa gtggtaaacc tgaattaact
     7801 gctactagtg ttgaaaaatt tttaattgaa aaatttgata gtgttagtga tttaatgcaa
     7861 ttaagtgaag gtgaagaaag tagagctttt agttttgatg ttggtggtag aggttatgtt
     7921 ttaagagtta atagttgtgc tgatggtttt tataaagata gatatgttta tagacatttt
     7981 gctagtgctg ctttacctat tcctgaagtt ttagatattg gtgaatttag tgaaagttta
     8041 acttattgta ttagtagaag agcacaaggt gttactttac aagatttacc tgaaactgaa
     8101 ttacctgctg ttttacaacc tgttgctgaa gctatggatg ctattgcggc ggctgatctc
     8161 agtcaaactt cgggttttgg tccttttggt cctcaaggta ttggtcaata tactacttgg
     8221 agagatttta tttgtgctat tgctgatcct catgtttatc attggcaaac tgttatggat
     8281 gatactgtta gtgctagtgt tgctcaagca ttagatgaat taatgttatg ggctgaagat
     8341 tgtcctgaag ttagacattt agttcatgct gattttggta gtaataatgt tttaactgat
     8401 aatggtagaa ttactgctgt tattgattgg agtgaagcta tgtttggtga tagtcaatat
     8461 gaagttgcta atattttttt ttggagacct tggttagctt gtatggaaca acaaactaga
     8521 tattttgaaa gaagacatcc tgaattagct ggtagtccta gattaagagc ttatatgtta
     8581 agaattggtt tagatcaatt atatcaaagt ttagttgatg gtaattttga tgatgctgct
     8641 tgggctcaag gtagatgtga tgctattgtt agaagtggtg ctggtactgt tggtagaact
     8701 caaattgcta gaagaagtgc tgctgtttgg actgatggtt gtgttgaagt tttagctgat
     8761 agtggtaata gaagacctag tactagacct agagctaaag aataacaatt cgttcaagcc
     8821 gagatcggct tacgcgt
//
