LOCUS       sequence-292294-        2120 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR DuMPLING negative control plasmid with probe/barcode P1 and 
            negative control lacO1array spacer in the sgRNA. Also template for 
            library PCRs..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2120)
  AUTHORS   Camsund D, Lawson MJ, Larsson J, Jones D, Zikrin S, Fange D, Elf J
  TITLE     Time-resolved imaging-based CRISPRi screening.
  JOURNAL   Nat Methods. 2020 Jan;17(1):86-92. doi: 10.1038/s41592-019-0629-y. 
            Epub 2019 Nov 18.
  PUBMED    31740817
REFERENCE   2  (bases 1 to 2120)
  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: 
            "10.1038/s41592-019-0629-y"; journalName: "Nat Methods"; date: 
            "2020-01"; volume: "17"; issue: "1"; pages: "86-92"
COMMENT     Sequence Label: pGuide-P1-O1a (EL702)
FEATURES             Location/Qualifiers
     source          1..2120
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(1..48)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(174..192)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        201..235
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     misc_RNA        257..332
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(427..446)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      complement(524..1112)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(604..623)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1223..2017)
                     /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
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1334..1353)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1944..1963
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctaggg gaatcgaata
       61 cgagagaagt gcgaacactt cctgggcacg aagccatatc ttctgtgccc agctcgtatt
      121 cgaggactcc ccagtgactg tgggagtcct ccccttttat ccaatgtaga ctgcctatag
      181 tgagtcgtat taatctccaa tttacagcta gctcagtcct aggtattatg ctagcatccg
      241 ctcacaattc cacatggttt tagagctaga aatagcaagt taaaataagg ctagtccgtt
      301 atcaacttga aaaagtggca ccgagtcggt gcttttttta cgcgccctgt agcggcgcat
      361 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
      421 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gtttatgtga gcaaaaggcc
      481 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
      541 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
      601 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
      661 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
      721 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
      781 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
      841 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
      901 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
      961 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     1021 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     1081 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacgggga
     1141 gtgtaactcg atgagctacc cggtaatggg agaccagaaa caaaaaaagg ggagcggttt
     1201 cccgctcccc ttcaataatt ggtcagaaga actcgtcaag aaggcgatag aaggcgatgc
     1261 gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa gcggtcagcc cattcgccgc
     1321 caagctcttc agcaatatca cgggtagcca acgctatgtc ctgatagcgg tccgccacac
     1381 ccagccggcc acagtcgatg aatccagaaa agcggccatt ttccaccatg atattcggca
     1441 agcaggcatc gccatgggtc acgacgagat cctcgccgtc gggcatgcgc gccttgagcc
     1501 tggcgaacag ttcggctggc gcgagcccct gatgctcttc gtccagatca tcctgatcga
     1561 caagaccggc ttccatccga gtacgtgctc gctcgatgcg atgtttcgct tggtggtcga
     1621 atgggcaggt agccggatca agcgtatgca gccgccgcat tgcatcagcc atgatggata
     1681 ctttctcggc aggagcaagg tgagatgaca ggagatcctg ccccggcact tcgcccaata
     1741 gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa ggaacgcccg
     1801 tcgtggccag ccacgatagc cgcgctgcct cgtcctgcag ttcattcagg gcaccggaca
     1861 ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga cagccggaac acggcggcat
     1921 cagagcagcc gattgtctgt tgtgcccagt catagccgaa tagcctctcc acccaagcgg
     1981 ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg aaacgatcct catcctgtct
     2041 cttgatcaga tcttgatccc ctgcgccatc agatccttgg cggcaagaaa gccatccagt
     2101 tttaaccatc agcgaattct
//
