LOCUS       sequence-266084-        5515 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for constitutive expression of a single guide RNA from Streptococcus
            thermophilus #3 with a seed region that targets cpaA gene's promoter
            from Caulobacter crescentus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5515)
  AUTHORS   Guzzo M, Castro LK, Reisch CR, Guo MS, Laub MT
  TITLE     A CRISPR Interference System for Efficient and Rapid Gene Knockdown 
            in Caulobacter crescentus.
  JOURNAL   mBio. 2020 Jan 14;11(1):e02415-19. doi: 10.1128/mBio.02415-19.
  PUBMED    31937638
REFERENCE   2  (bases 1 to 5515)
  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: "10.1128/mBio"; 
            journalName: "mBio"; date: "2020-01-14- 14"; volume: "11"; issue: 
            "1"; pages: "e02415-19"
COMMENT     Sequence Label: pBXMCS-2 Pconstitutive-sgRNA(Sth3)_cpaA
FEATURES             Location/Qualifiers
     source          1..5515
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      41..84
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(227..243)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(227..243)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(275..294)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(276..294)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(304..321)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(304..320)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(313..335)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(1029..1691)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      1692..2463
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             3943..4737
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCATVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3997..4016)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4607..4626
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      5085..5131
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 cttgaaaagg tggcaccgat tcggtgtttt tttttttttg ataaaacgaa aggcccagtc
       61 tttcgactga gcctttcgtt ttataaacgg gtcttgaggg gttttttaag cttctgcagc
      121 tggatccatg gatatctaga tttccgcaaa ataaaaaagg ggacctctag ggtccccaat
      181 taattagcta aagccgctag cccatgggaa ttcctgcagc ccgggggatc cactagttct
      241 agagcggccg ccaccgcggt ggagctccaa ttcgccctat agtgagtcgt attacgcgcg
      301 ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
      361 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
      421 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg aaattgtaag cgttaatatt
      481 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgta
      541 ctgcgatgag tggcagggcg gggcgtaatt tttttaaggc agttattggt gcccttaaac
      601 gcctggttgc tacgcctgaa taagtgataa taagcggatg aatggcagaa attcgaaagc
      661 aaattcgacc cggtcgtcgg ttcagggcag ggtcgttaaa tagccgctta tgtctattgc
      721 tggtttaccg gtttattgac taccggaagc agtgtgaccg tgtgcttctc aaatgcctga
      781 ggccagtttg ctcaggctct ccccgtggag gtaataattg acgatatgat catttattct
      841 gcctcccaga gcctgataaa aacggtgaat ccgttagcga ggtgccgccg gcttccattc
      901 aggtcgaggt ggcccggctc catgcaccgc gacgcaacgc ggggaggcag acaaggtata
      961 gggcggcgag gcggctacag ccgatagtct ggaacagcgc acttacgggt tgctgcgcaa
     1021 cccaagtgct accggcgcgg cagcgtgacc cgtgtcggcg gctccaacgg ctcgccatcg
     1081 tccagaaaac acggctcatc gggcatcggc aggcgctgct gcccgcgccg ttcccattcc
     1141 tccgtttcgg tcaaggctgg caggtctggt tccatgcccg gaatgccggg ctggctgggc
     1201 ggctcctcgc cggggccggt cggtagttgc tgctcgcccg gatacagggt cgggatgcgg
     1261 cgcaggtcgc catgccccaa cagcgattcg tcctggtcgt cgtgatcaac caccacggcg
     1321 gcactgaaca ccgacaggcg caactggtcg cggggctggc cccacgccac gcggtcattg
     1381 accacgtagg ccgacacggt gccggggccg ttgagcttca cgacggagat ccagcgctcg
     1441 gccaccaagt ccttgactgc gtattggacc gtccgcaaag aacgtccgat gagcttggaa
     1501 agtgtcttct ggctgaccac cacggcgttc tggtggccca tctgcgccac gaggtgatgc
     1561 agcagcattg ccgccgtggg tttcctcgca ataagcccgg cccacgcctc atgcgctttg
     1621 cgttccgttt gcacccagtg accgggcttg ttcttggctt gaatgccgat ttctctggac
     1681 tgcgtggcca tgcttatctc catgcggtag gggtgccgca cggttgcggc accatgcgca
     1741 atcagctgca acttttcggc agcgcgacaa caattatgcg ttgcgtaaaa gtggcagtca
     1801 attacagatt ttctttaacc tacgcaatga gctattgcgg ggggtgccgc aatgagctgt
     1861 tgcgtacccc ccttttttaa gttgttgatt tttaagtctt tcgcatttcg ccctatatct
     1921 agttctttgg tgcccaaaga agggcacccc tgcggggttc ccccacgcct tcggcgcggc
     1981 tccccctccg gcaaaaagtg gcccctccgg ggcttgttga tcgactgcgc ggccttcggc
     2041 cttgcccaag gtggcgctgc ccccttggaa cccccgcact cgccgccgtg aggctcgggg
     2101 ggcaggcggg cgggcttcgc ccttcgactg cccccactcg cataggcttg ggtcgttcca
     2161 ggcgcgtcaa ggccaagccg ctgcgcggtc gctgcgcgag ccttgacccg ccttccactt
     2221 ggtgtccaac cggcaagcga agcgcgcagg ccgcaggccg gatgcttttc cccagagaaa
     2281 attaaaaaaa ttgatggggc aaggccgcag gccgcgcagt tggagccggt gggtatgtgg
     2341 tcgaaggctg ggtagccggt gggcaatccc tgtggtcaag ctcgtgggca ggcgcagcct
     2401 gtccatcagc ttgtccagca gggttgtcca cgtgccgagc gaagcgagcc agccggtggc
     2461 cgctcgcggc catcgtccac atatccacgg gctggcaagg gagcgcagcg accgcgcagg
     2521 gcgaagcccg gagagcaagc ccgtagggcg ccgcagccgc cgtaggcggt catgactttg
     2581 cgaagcaaag tctagtgagt atactcaagc attgagtggc ccgccggagg caccgccttg
     2641 cgctgccccc gtcgagccgg ttggacacca aaagggaggg gcaggcatgg cggcatacgc
     2701 gatcatgcga tgcaagaagc tggcgaaaat gtgcaacgtg gcggccagtc tcaagcacgc
     2761 ctaccgcgag cgcgagacgc ccaacgctga cgccagcagg acgccagaga acgagcactg
     2821 ggcggccagc agcaccgatg aagcgatggg ccgactgcgc gagttgctgc cagagaagcg
     2881 gcgcaaggac gctgtgttgg cggtcgagta cgtcatgacg gccagcccgg aatggtggaa
     2941 gtcggccagc caagaacagc aggcggcgtt cttcgagaag gcgcacaagt ggctggcgga
     3001 caagtacggg gcggatcgca tcgtgacggc cagcatccac cgtgacgaaa caagcccgca
     3061 catgaccgcg ttcgtggtgc cgctgacgca ggacggcagg ctgtcggcca aggagttcat
     3121 cggcaacaaa gcgcagatga cccgcgacca gaccacgttt gcggccgctg tggccgatct
     3181 agggctgcaa cggggcatcg agggcagcaa ggcacgtcac acgcgcattc aggcgttcta
     3241 cgaggccctg gagcggccac cagtgggcca cgtcaccatc agcccgcaag cggtcgagcc
     3301 acgcgcctat gcaccgcagg gattggccga aaagctggga atctcaaagc gcgttgagac
     3361 gccggaagcc gtggccgacc ggctgacaaa agcggttcgg caggggtatg agcctgccct
     3421 acaggccgcc acaggagcgc gtgagatgcg caagaaggcc gatcaagccc aagagacggc
     3481 ccgagacctt cgggagcgcc tgaagcccgt tctggacgcc ctggggccgt tgaatcggga
     3541 tatgcaggcc aaggccgccg cgatcatcaa ggccgtgggc gaaaagctgc tgacggaaca
     3601 gcgggaagtc cagcgccaga aacaggccca gcgccagcag gaacgcgggc gcgcacattt
     3661 ccccgaaaag tgccacctgg gatgaatgtc agctactggg ctatctggac aagggaaaac
     3721 gcaagcgcaa agagaaagca ggtagcttgc agtgggctta catggcgata gctagactgg
     3781 gcggttttat ggacagcaag cgaaccggaa ttgccagctg gggcgccctc tggtaaggtt
     3841 gggaagccct gcaaagtaaa ctggatggct ttcttgccgc caaggatctg atggcgcagg
     3901 ggatcaagat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     3961 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     4021 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     4081 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     4141 ctatcgtggc tggccacgac gggcgttcct tgcgcaactg tgctcgacgt tgtcactgaa
     4201 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     4261 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     4321 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     4381 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     4441 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     4501 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4561 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4621 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4681 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4741 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4801 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4861 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc catgggcaaa
     4921 tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtt
     4981 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagttttt atataagctt
     5041 tcagtcggcg cgaatgctcc acgccgcgcc cagcggcgcg gatccaaata aaacgaaagg
     5101 ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcgctgcag ccagccgtgg
     5161 tcgggcagca ggtagaaggc gccctcgtcc tgatcctcgc ccgaaacctc cagcccccgg
     5221 tcgatggctt cgacgacata gccggccgcg cggcaggtgt cggtgagcgc ggccagcagg
     5281 gcggcttcct ggtcaggggt caggtcggtc atgggcaaga ggtccaggtc gtggtttgtc
     5341 ggcggcttct agcatggacc gcccgcgccc gtgaggccga ggatttcgcg ctggtcagac
     5401 aacctacttg ccgtccccac atgttagcgc taccaagtgc cgagagcgtg acgcgagtgg
     5461 cttgttttag agctgtgaaa acagcgagtt aaaataaggc ttagtccgta ctcaa
//
