LOCUS       sequence-311487-        6542 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 6542)
  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 6542)
  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: pBSV2_Psyn-sgRNArodA
FEATURES             Location/Qualifiers
     source          1..6542
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        22..97
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(140..157)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(140..156)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(149..171)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             713..1522
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(780..799)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     regulatory      1773..1782
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1779..2153
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      2291..2879
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2780..2799
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6357..6378
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6393..6398,6399..6416,6417..6423)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6431..6447
                     /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     6436..6458
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6455..6471
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6455..6471
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6508..6542
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
ORIGIN
        1 gttaatctac ctaatatacc agttttagag ctagaaatag caagttaaaa taaggctagt
       61 ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt ttttgaagct tgggcccgaa
      121 caaaaactca aagcttggca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
      181 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
      241 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcg
      301 gccgctaccc gagcttcaag gaagatttcc tattaaggtt gaacttaaga gcttaagcat
      361 agacgatttg aaaaaaattt taaaacaaac aaaaaattct ttaataaaac aatatgttgc
      421 gatgtttaag gtttatgatt tagatttaaa gtttagtgag gaagctatag atagaattgc
      481 agagcttact tttaatatga atcttgagag tgaaaatctt ggtgccagaa gacttcacgg
      541 tgttatggaa atagtgcttg cagatctttt ttttgaagtg cctggcagta agttgaaaaa
      601 atttgaaata aacttggact atgttaataa aaaaatacaa attaacgaac aaaaagattt
      661 gaattattat ataatttagt taaaagcaat tttaaatgag ggaggtttcc atatgagcca
      721 tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg
      781 gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg
      841 gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt
      901 tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa
      961 gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac
     1021 agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc
     1081 agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg
     1141 cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga
     1201 ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact
     1261 tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat
     1321 ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg
     1381 ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa
     1441 acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt
     1501 gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat
     1561 tacgctgact tgacgggacg gcgcttacaa tttcctgatg cggtattttc tccttacgca
     1621 tctgtgcggt atttcacacc gcatcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     1681 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     1741 gataaatgct tcaataatag cacgtgagga gggccaccat ggccaagttg accagtgccg
     1801 ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga gttctggacc gaccggctcg
     1861 ggttctcccg ggacttcgtg gaggacgact tcgccggtgt ggtccgggac gacgtgaccc
     1921 tgttcatcag cgcggtccag gaccaggtgg tgccggacaa caccctggcc tgggtgtggg
     1981 tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt cgtgtccacg aacttccggg
     2041 acgcctccgg gccggccatg accgagatcg gcgagcagcc gtgggggcgg gagttcgccc
     2101 tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc
     2161 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     2221 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     2281 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2341 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2401 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     2461 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2521 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2581 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2641 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2701 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2761 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2821 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2881 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     2941 tctttcctgc actagatccc cggcaagttc atccttaggg ttttcatatc tcttaaaaag
     3001 tcaatttctg cttgattaat ttcttgtgga ttattgtttt tgcggttttt cattttttat
     3061 ctccttgagt attgaatttt ttcttatata taaatatata gcaaaaacta tttttgccaa
     3121 cttttttaca aaaaattttt tattaaaaca cttagggctt tgctaaattc tcttttaaaa
     3181 gaacttagta aagccctatg gatttaagaa ctgattatac ttacgtagta aaaatactac
     3241 agatattaat agtatagacc atatcaaaat cataatcaat ttatatttat ctaaaattgt
     3301 agtccataca aaatacaaaa aaattaggtt tttctaaatt ctattttaaa agaacttagt
     3361 aacacctaat ctctaaatta cttacaaaat tctaaaatta gatttttagt tcttcatatt
     3421 tctttaaaag ttttttaaga aaatcttttt gattttcaaa aatttcatgc atcataaaac
     3481 tagtaaattt ggattttttt ttgtaaaaat cataactttc ttgggtttta agttgaaatc
     3541 ttaatggttt tattggattt tgcctagatt tttttaacgc cggactttct ttatctttta
     3601 taaaattata agaattttta ataccatttt ctattaaata atcttctttt ataagcccct
     3661 cttgcagtgc agttgctaat cttaaatact tatacgcttg agtttttgct acatcaaagt
     3721 cttttataaa atcaagaaaa gttttataac catcaaattt ataatattga ttatctttta
     3781 tttcttttaa aattttaata ttttccaccc tgtggaatat ttcttttttg aaattgtcat
     3841 ttaacttttc ctttagtgtc ttgtaataag aaattttact ttcattaatc attaaatttt
     3901 gttcaacata agtatctatc ttttttgtta aaactttaac aacacgtttt ttttccataa
     3961 aaattttcct ttatttttgc aacattttcc ttaattgata taattgttac tattttagat
     4021 tttttaatat ccacttaaga attaatccct cactagaata ttttttatta tagaattcat
     4081 atacatgttt ttctaaatct ataaatttat caacaattgc tttattatag ggtgtatcta
     4141 tcttttcttt tcttaataat cgtctcatat tctttaaata acaacaaaca ctgccggttt
     4201 taaacctaaa ttctatgtaa tatatttttg aaaaagtata tacattttca accccgttta
     4261 tttcatactt tacaaaaaca ttatttattg gttttctata accatagtaa atgcctaaaa
     4321 atttgtcccc gtctttcata ggatatagat aaagtttttg aatctctgat ttatctaata
     4381 atttttcaaa aaaaattata caccgctttt gtttgttact aactactcta tgcaatgcct
     4441 ttaatatttt tgtgtgatat atatttttat cacctttttt ttctttcagc aggaagatac
     4501 tttcttttgg aaatttaatt ttttgttcat ttattcttgc agccttaagt ccgattttca
     4561 tatttaatcc ttatatagcc tttctattta attcttttaa actcaataaa cttttttggt
     4621 ttttcattag ttctaataat tcgcaataat atgtgcaaaa cactttattg tattctattt
     4681 tcttttggtt attcaaatat tcttttataa ttggctttag aagttcaata tttacttctt
     4741 tttttagttg ttcaataagg atattgaaaa tgtttgtttt taaatcttga taattttgtt
     4801 gagaattttc tttttttctt tcaattgact tctctaattt acgctttata ttgtttaaat
     4861 cgctatattt atgactttca ataataaaat gaggcttaaa tttgtaattt tcgtatattt
     4921 tttggaaact tgtttctaat tgtttagaat tatattcatt tttttcaaat tctttttgag
     4981 tgttgcatag aatttccttt aatttttctt gtttttcttt catacaagat ttagtaaaat
     5041 gtatattttt attttttagt aatttaattt ctattatttt tatgatttta agtattttga
     5101 ttatattatc tttatcaata tttaaattga ataaaattgt aagaaaaatt gaaagaattt
     5161 ttttacataa aaagttacat ttattgaagt aatttcttac ttgatacttt tctatttcat
     5221 taatctttct ttcttctttt atattatttt tattatttaa acactccttt aaatctacac
     5281 tcccattttt agtaaatcta tctttaaggc cccttgaaac tctattttga aatcttaaat
     5341 cttttttttc ttgaaagtat ttatagattt tttggtaaca ttcttttttt gaaaaattaa
     5401 gttgatagta tatttcggtc cccaaattta cccctaagtg tttgtgataa ttagttgtga
     5461 ctttaaattc tttttctaat ttgtaaaggt aattttgcag tgtttttagt ttaacgggtg
     5521 tttgactatt tcttttaagg ttttcattaa agtaatagag tatgtttttt tgtgtgtatt
     5581 tcttatatgt tttatttatg tacgctagtg ttgagataag gactactaat ttatactgta
     5641 atttattgta gcaagttgtg gttttactgg ttttgtttat attagataag ttttccattt
     5701 atagctcctt gaataccgga ttttttatct ataaagtata tatatcacaa ttttgattta
     5761 aaagtaaatt ctttttttaa aaatacaaaa attttttata aattatatag accatgatat
     5821 tgtattaata ctaattgatt ttaatttaag gttgaactaa aataaacata gttttgtaga
     5881 aaataatttt tcattatttt gctacttgat attggatcgt aaaaatatta gggctttaca
     5941 aagttctttt aaaagagaat ttagtaaagc cctaagtctt ttaataaaaa attttttgta
     6001 aaaaagttgg caaaaatagt ttttgctata tatttatata taagaaaaaa ttcaatactc
     6061 aaggagataa aaaatggaaa acttaaacaa taatccacaa aaaattaatc aaataaataa
     6121 gcaaactaat tttattggtt ttgaaaattt tatagaactt gataatcaag ttaatcaaac
     6181 taaagaaaaa agtaaattaa gtaagatagg taagaaatta cccggaataa gcagtcaagt
     6241 ctagtgcggc cgcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     6301 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     6361 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     6421 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
     6481 gaattcgagc tcgaattcta aagatctttg acagctagct cagtcctagg tataatacta
     6541 gt
//
