LOCUS       sequence-486277-        6476 bp    DNA     circular SYN 24-SEP-2025
DEFINITION  PVC cargo/regulatory region - deleted toxin cargos (Pdp1/Pnf) - 
            vegfa sgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6476)
  AUTHORS   Kreitz J, Yang V, Friedrich MJ, Pham J, Macrae RK, Zhang F
  TITLE     Targeted delivery of diverse biomolecules with engineered bacterial 
            nanosyringes.
  JOURNAL   Nat Biotechnol. 2025 Aug 12. doi: 10.1038/s41587-025-02774-x.
  PUBMED    40796978
REFERENCE   2  (bases 1 to 6476)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 24, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2025 Aug 12. doi: 10.1038/s41587-025-02774-x."
COMMENT     Sequence Label: pBR322-sgRNA-pvc18-20
FEATURES             Location/Qualifiers
     source          1..6476
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             467..658
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(652..674)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    760..900
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     815..834
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(915..932)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1086..1674)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1166..1185)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1845..2636,2637..2705))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2468..2487
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2706..2810)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2855..2889
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     protein_bind    2922..2940
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    2977..2995
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     promoter        join(3110..3115,3116..3132,3133..3138)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc ggggttgcct
       61 tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact gctgctgcaa
      121 aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt cgtaaagtct
      181 ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg caggatgctg
      241 ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt gaccctgagt
      301 gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac aacgttccag
      361 taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc gtttcatcgg
      421 tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga ccaaacagga
      481 aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc ttctggagaa
      541 actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc acgaccacgc
      601 tgatgagctt taccgcagct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
      661 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
      721 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
      781 tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga
      841 gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
      901 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
      961 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     1021 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     1081 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     1141 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     1201 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     1261 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     1321 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     1381 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     1441 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     1501 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     1561 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     1621 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     1681 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     1741 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     1801 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     1861 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     1921 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     1981 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     2041 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     2101 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     2161 gcaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     2221 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     2281 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     2341 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     2401 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     2461 acacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     2521 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     2581 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     2641 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     2701 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     2761 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     2821 cgaaaagtgc cacctgacgt ctaagaaacc cctcttgaca gctagctcag tcctaggtat
     2881 aatgctagcg gtgagtgagt gtgtgcgtgg tttgagagct agggccctga agaagggccc
     2941 tagcaagttc aaataaggct agtccgttat caacttgggc cctgaagaag ggcccaagtg
     3001 gcaccgagtc ggtgcttttt ttccgaaaaa aaaaccccgc ccctgacagg gcggggtttt
     3061 ttttaattaa cgtatcacga ggccctttcg tcttcaagaa ttctcatgtt tgacagctta
     3121 tcatcgataa gctttaatgc ggtagtttat cacagttaaa ttgctaacgc agtcaggcac
     3181 cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg caccgtcacc ctggatgctg
     3241 taggcatagg cttggttatg ccggtactgc cgggcctctt gcgggatatc gtccattccg
     3301 acagcatcgc cagtcactat ggcgtgctgc tagcgctata tgcgttgatg caatttctat
     3361 gcgcacccgt tctcggagca ctgtccgacc gctttggccg ccgcccagtc ctgctcgctt
     3421 cgctacttgg agccactatc gactacgcga tcatggcgac cacacccgtc ctgtggatcc
     3481 tatggatttt catgtcatca ggagaatatt caaaattctg tgtcacacaa aatattttac
     3541 agcgtaatca caccattaag aggggcatca aaataaatgg ttaactggga caatgttgaa
     3601 tttcattttt ataaggcatt gtccgtatta tccaccatcc agatcaacgg attcagatgg
     3661 tgtaaagaaa gcatatagct ggtaggtaaa attaagaagt gtagttttat aaagtattaa
     3721 tggagaacga tgaatttaaa gtaaatggag gaaggttagt ttcactatca aattacagga
     3781 ccaaaaatat ggagtattcc atcttctttg gtgaattaag tcttgaagac ttcaacaaat
     3841 attgcctgga actatgggcc tgtgaatggt attggagtat catttataac gactatttcg
     3901 ataggaaatt taaagtcttg ttgaaaatga ctaatcgtct aaaagtatgt ctggagaaat
     3961 aggtttaatc tctatattta acggttttat tttcatctat ttactaatgg taaatttttt
     4021 tactctttga atatattggt gagtttgtta tccccattag ttaccagggt attattaatg
     4081 agtgaaattc aattttattg atattaaagt caaatttaca aatgatgact ttattatttt
     4141 ttatttatta attttaaatt attttttgtt aatctttact ataaatgaat tttttgcttg
     4201 catggacttg atttttttat atataacatt gacaaatatt aattcatata tatgattgtt
     4261 atagtgagtt atttaattga ttcgactaac aatttcattg taatttttca agatagcagt
     4321 caattgctat aacaagcatc tttctcgtca aaggaaaatg tgtatggcct ataagcagca
     4381 acttgaaaga tgaaggatat aatgatgatt aacttaacgg agagtagata ttagaagtag
     4441 tagaaggaca atttaagact ttatacccat taaaacaaag gggtaaaata atgatgtttg
     4501 atgaaaattt agaatgtgtt aatcacgaaa ttatcaatga aaaattagat acgaaaattc
     4561 accttgctaa tccttatgcc ttttgggggg gaagggtaaa tgagaatata aatgggttaa
     4621 ttaaacaaca ttctttaaga ggaactgact ttaataaaat ttttaaccgg aagataaatt
     4681 tttttgtaaa tggaataaat aattgccccc gaaaaactcg gaatataaaa ccccctaatt
     4741 aattatttaa aggaataaga gtttatttgc tcgtagagta attatgttcc attaatatgc
     4801 taaattgctc agtaaagtta ttaacttttt ttcaacgcca gtgaataagt ctgttttatt
     4861 aaataaggag ttcaggtgtt tatttcaaaa gaattatcaa gctttattgc tgtagctaaa
     4921 aataaatcaa taaataaagc aacaacggaa ctttttatta cacagtcacc gatttgccga
     4981 agcttaaaaa aactggaatt ttcattaggt gtaaaactct ttgtaagaaa aagccaggga
     5041 ttggctttaa cagaagaagg agaatgctta tataataaag ttttgtcaat ttgtgaacgg
     5101 cttcaagaaa tagaaggtga ctacaagaat gaaaaatcaa catacaatga tctatatcaa
     5161 aaaaatatat gaggttctaa tataataagc ggttctagat aactaaagtc atctattacc
     5221 aatatctgta tactcagtga aagttttgtc ttctaggtaa gttttcataa tagtgacata
     5281 ccttattttt tcatctgggt gtctctcatc ctgaagaatg ggtgatatag ttgaaataac
     5341 tcagttcgtt ttgtggttta cgatttttgg cggatcacaa aaatagggct gagtcccttc
     5401 aagtgagttt tgaaatatac aattataccc ttcatccttt aagttgttgc tttgttggct
     5461 gttttcattc acctcagtca catagttatc tatgctcctg aggattcgtt cacttgccgt
     5521 cgtgctgcaa ctcgaaatct attgggtata tatgtaattt tcatattctg tttcatttta
     5581 acggtattat ttaaatattt atatttagaa cattaggtta ttagaataga taattttagg
     5641 taaatgtaat taattcaacc atcacggaga gtttatcagt cgaccgatgc ccttgagagc
     5701 cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg ccgcacttat
     5761 gactgtcttc tttatcatgc aactcgtagg acaggtgccg gcagcgctct gggtcatttt
     5821 cggcgaggac cgctttcgct ggagcgcgac gatgatcggc ctgtcgcttg cggtattcgg
     5881 aatcttgcac gccctcgctc aagccttcgt cactggtccc gccaccaaac gtttcggcga
     5941 gaagcaggcc attatcgccg gcatggcggc cgacgcgctg ggctacgtct tgctggcgtt
     6001 cgcgacgcga ggctggatgg ccttccccat tatgattctt ctcgcttccg gcggcatcgg
     6061 gatgcccgcg ttgcaggcca tgctgtccag gcaggtagat gacgaccatc agggacagct
     6121 tcaaggatcg ctcgcggctc ttaccagcct aacttcgatc attggaccgc tgatcgtcac
     6181 ggcgatttat gccgcctcgg cgagcacatg gaacgggttg gcatggattg taggcgccgc
     6241 cctatacctt gtctgcctcc ccgcgttgcg tcgcggtgca tggagccggg ccacctcgac
     6301 ctgaatggaa gccggcggca cctcgctaac ggattcacca ctccaagaat tggagccaat
     6361 caattcttgc ggagaactgt gaatgcgcaa accaaccctt ggcagaacat atccatcgcg
     6421 tccgccatct ccagcagccg cacgcggcgc atctcgggca gcgttgggtc ctggcc
//
