LOCUS       sequence-461753-        3036 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  M13 phagemid constitutively expressing sgRNA-A0. sgRNA-A0 is a dummy
            20-nt sequence, containing a Golden Gate cloning site (BsaI)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3036)
  AUTHORS   Pujar A, Pathania A, Hopper C, Pandi A, Calderon CR, Fugger M, Nowak
            T, Kushwaha M
  TITLE     Phage-mediated intercellular CRISPRi for biocomputation in bacterial
            consortia.
  JOURNAL   Nucleic Acids Res. 2024 Dec 27:gkae1256. doi: 10.1093/nar/gkae1256.
  PUBMED    39727169
REFERENCE   2  (bases 1 to 3036)
  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: "Nucleic 
            Acids Res. 2024 Dec 27:gkae1256. doi: 10.1093/nar/gkae1256."
COMMENT     Sequence Label: pSEVA26_sgRNA-A0_M13ps
FEATURES             Location/Qualifiers
     source          1..3036
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(14..100)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     115..137
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     promoter        168..202
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     misc_RNA        223..298
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      363..457
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(578..597)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             589..1404
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     oriT            1571..1679
                     /note="incP origin of transfer"
     rep_origin      1726..2271
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     2172..2191
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2388..2405
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2438..2947)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2633..2654)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2841..2860
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tctagggcgg cggatttgtc ctactcagga gagcgttcac cgacaaacaa cagataaaac
       61 gaaaggccca gtctttcgac tgagcctttc gttttatttg atgcctttaa ttaaagcgga
      121 taacaatttc acacaggagg ccgcctaggc cgcggccgcg cgaattcttg acagctagct
      181 cagtcctagg tataatgcta gcagagacca ctttcggtct ctgttttaga gctagaaata
      241 gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt
      301 ttctgcaggc atgcaagctt gcggccgcgt cgtgactggg aaaaccctgg cgactagtct
      361 tggactcctg ttgatagatc cagtaatgac ctcagaactc catctggatt tgttcagaac
      421 gctcggttgc cgccgggcgt tttttattgg tgagaatcca ggggtcccca ataattacga
      481 tttaaatttg tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca
      541 tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt
      601 cagcgtgaaa cgagctgtag ccgtccgcgt ctgaacagca acatggatgc ggatctgtat
      661 ggctataaat gggcgcgtga taacgtgggt cagagcggcg cgaccattta tcgtctgtat
      721 ggcaaaccgg atgcgccgga actgtttctg aaacatggca aaggcagcgt ggcgaacgat
      781 gtgaccgatg aaatggtgcg tctgaactgg ctgaccgaat ttatgccgct gccgaccatt
      841 aaacatttta ttcgcacccc ggatgatgcg tggctgctga ccaccgcgat tccgggcaaa
      901 accgcgtttc aggtgctgga agaatatccg gatagcggcg aaaacattgt ggatgcgctg
      961 gccgtgtttc tgcgtcgtct gcatagcatt ccggtgtgca actgcccgtt taacagcgat
     1021 cgtgtgtttc gtctggccca ggcgcagagc cgtatgaaca acggcctggt ggatgcgagc
     1081 gattttgatg atgaacgtaa cggctggccg gtggaacagg tgtggaaaga aatgcataaa
     1141 ctgctgccgt ttagcccgga tagcgtggtg acccacggcg attttagcct ggataacctg
     1201 attttcgatg aaggcaaact gattggctgc attgatgtgg gccgtgtggg cattgcggat
     1261 cgttatcagg atctggccat tctgtggaac tgcctgggcg aatttagccc gagcctgcaa
     1321 aaacgtctgt ttcagaaata tggcattgat aatccggata tgaacaaact gcaatttcat
     1381 ctgatgctgg atgaattttt ctaataatta attggaccgc ggtccgcgcg ttgtcctttt
     1441 ccgctgcata accctgcttc ggggtcatta tagcgatttt ttcggtatat ccatcctttt
     1501 tcgcacgata tacaggattt tgccaaaggg ttcgtgtaga ctttccttgg tgtatccaac
     1561 ggcgtcagcc gggcaggata ggtgaagtag gcccacccgc gagcgggtgt tccttcttca
     1621 ctgtccctta ttcgcacctg gcggtgctca acgggaatcc tgctctgcga ggctggccgt
     1681 aggccggccc tagaaatatt ttatctgatt aataagatga tcttcttgag atcgttttgg
     1741 tctgcgcgta atctcttgct ctgaaaacga aaaaaccgcc ttgcagggcg gtttttcgaa
     1801 ggttctctga gctaccaact ctttgaaccg aggtaactgg cttggaggag cgcagtcacc
     1861 aaaacttgtc ctttcagttt agccttaacc ggcgcatgac ttcaagacta actcctctaa
     1921 atcaattacc agtggctgct gccagtggtg cttttgcatg tctttccggg ttggactcaa
     1981 gacgatagtt accggataag gcgcagcggt cggactgaac ggggggttcg tgcatacagt
     2041 ccagcttgga gcgaactgcc tacccggaac tgagtgtcag gcgtggaatg agacaaacgc
     2101 ggccataaca gcggaatgac accggtaaac cgaaaggcag gaacaggaga gcgcacgagg
     2161 gagccgccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccaccactga
     2221 tttgagcgtc agatttcgtg atgcttgtca ggggggcgga gcctatggaa aaacggcttt
     2281 gccgcggccc tctcacttcc ctgttaagta tcttcctggc atcttccagg aaatctccgc
     2341 cccgttcgta agccatttcc gctcgccgca gtcgaacgac cgagcgtagc gagtcagtga
     2401 gcgaggaagc ggaatatatc cggcgcgaga ttgatttacc ccggttgata atcagaaaag
     2461 ccccaaaaac aggaagattg tataagcaaa tatttaaatt gtaaacgtta atattttgtt
     2521 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     2581 caaaatccct tataaatcaa aagaatagcc cgagataggg ttgagtgttg ttccagtttg
     2641 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     2701 tcagggcgat ggcccactac gtgaaccatc acccaaatca agttttttgg ggtcgaggtg
     2761 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     2821 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     2881 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     2941 ggcgcgtaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     3001 acgtgagttt tcgttccact gaggcgcgcc cagctg
//
