LOCUS       sequence-271135-        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(5..52)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(178..196)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        205..239
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     misc_RNA        261..336
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(431..450)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      complement(528..1116)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(608..627)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1227..2021)
                     /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(1338..1357)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1948..1967
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ttctcaaaaa acccctcaag acccgtttag aggccccaag gggttatgct aggggaatcg
       61 aatacgagag aagtgcgaac acttcctggg cacgaagcca tatcttctgt gcccagctcg
      121 tattcgagga ctccccagtg actgtgggag tcctcccctt ttatccaatg tagactgcct
      181 atagtgagtc gtattaatct ccaatttaca gctagctcag tcctaggtat tatgctagca
      241 tccgctcaca attccacatg gttttagagc tagaaatagc aagttaaaat aaggctagtc
      301 cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt tttacgcgcc ctgtagcggc
      361 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
      421 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgtttat gtgagcaaaa
      481 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
      541 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
      601 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
      661 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
      721 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
      781 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
      841 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
      901 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
      961 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     1021 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     1081 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     1141 gggagtgtaa ctcgatgagc tacccggtaa tgggagacca gaaacaaaaa aaggggagcg
     1201 gtttcccgct ccccttcaat aattggtcag aagaactcgt caagaaggcg atagaaggcg
     1261 atgcgctgcg aatcgggagc ggcgataccg taaagcacga ggaagcggtc agcccattcg
     1321 ccgccaagct cttcagcaat atcacgggta gccaacgcta tgtcctgata gcggtccgcc
     1381 acacccagcc ggccacagtc gatgaatcca gaaaagcggc cattttccac catgatattc
     1441 ggcaagcagg catcgccatg ggtcacgacg agatcctcgc cgtcgggcat gcgcgccttg
     1501 agcctggcga acagttcggc tggcgcgagc ccctgatgct cttcgtccag atcatcctga
     1561 tcgacaagac cggcttccat ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg
     1621 tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc gcattgcatc agccatgatg
     1681 gatactttct cggcaggagc aaggtgagat gacaggagat cctgccccgg cacttcgccc
     1741 aatagcagcc agtcccttcc cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg
     1801 cccgtcgtgg ccagccacga tagccgcgct gcctcgtcct gcagttcatt cagggcaccg
     1861 gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg ctgacagccg gaacacggcg
     1921 gcatcagagc agccgattgt ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa
     1981 gcggccggag aacctgcgtg caatccatct tgttcaatca tgcgaaacga tcctcatcct
     2041 gtctcttgat cagatcttga tcccctgcgc catcagatcc ttggcggcaa gaaagccatc
     2101 cagttttaac catcagcgaa
//
