LOCUS       sequence-235209-        2528 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express sgRNA. ColE1, Kan.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2528)
  AUTHORS   Cui L, Vigouroux A, Rousset F, Varet H, Khanna V, Bikard D
  TITLE     A CRISPRi screen in E. coli reveals sequence-specific toxicity of 
            dCas9.
  JOURNAL   Nat Commun. 2018 May 15;9(1):1912. doi: 10.1038/s41467-018-04209-5.
  PUBMED    29765036
REFERENCE   2  (bases 1 to 2528)
  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: "Nat 
            Commun."; date: "2018-05-15"; volume: "9(1):1912. doi"; pages: " 
            10.1038/s41467-018-04209-5"
COMMENT     Sequence Label: psgRNA
FEATURES             Location/Qualifiers
     source          1..2528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      49..135
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(520..1108)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(600..619)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1196..1290)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1321..2115)
                     /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(1432..1451)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2042..2061
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2294..2312
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2386..2420
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        2441..2516
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 tgcagcgcga tcgtaatcag gatcccatgg tacgcgtgct agaggcatca aataaaacga
       61 aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc
      121 ctgagtagga caaatccgcc gccctagacc ctccacgcac gttgtgatat gtagatgata
      181 atcattatca ctttacgggt cctttccggt gatccgacag gttacggggc ggcgacctcg
      241 cgggttttcg ctatttatga aaattttccg gtttaaggcg tttccgttct tcttcgtcat
      301 aacttaatgt ttttatttaa aataccctct gaaaagaaag gaaacgacag gtgctgaaag
      361 cgaggctttt tggggcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
      421 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
      481 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
      541 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
      601 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
      661 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
      721 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
      781 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
      841 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
      901 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
      961 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     1021 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     1081 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     1141 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgactagtg cttggattct
     1201 caccaataaa aaacgcccgg cggcaaccga gcgttctgaa caaatccaga tggagttctg
     1261 aggtcattac tggatctatc aacaggagtc caagcgagct ctcgaacccc agagtcccgc
     1321 tcagaagaac tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat
     1381 accgtaaagc acgaggaagc ggtcagccca ttcgccgcca agctcttcag caatatcacg
     1441 ggtagccaac gctatgtcct gatagcggtc cgccacaccc agccggccac agtcgatgaa
     1501 tccagaaaag cggccatttt ccaccatgat attcggcaag caggcatcgc catgggtcac
     1561 gacgagatcc tcgccgtcgg gcatgcgcgc cttgagcctg gcgaacagtt cggctggcgc
     1621 gagcccctga tgctcttcgt ccagatcatc ctgatcgaca agaccggctt ccatccgagt
     1681 acgtgctcgc tcgatgcgat gtttcgcttg gtggtcgaat gggcaggtag ccggatcaag
     1741 cgtatgcagc cgccgcattg catcagccat gatggatact ttctcggcag gagcaaggtg
     1801 agatgacagg agatcctgcc ccggcacttc gcccaatagc agccagtccc ttcccgcttc
     1861 agtgacaacg tcgagcacag ctgcgcaagg aacgcccgtc gtggccagcc acgatagccg
     1921 cgctgcctcg tcctgcagtt cattcagggc accggacagg tcggtcttga caaaaagaac
     1981 cgggcgcccc tgcgctgaca gccggaacac ggcggcatca gagcagccga ttgtctgttg
     2041 tgcccagtca tagccgaata gcctctccac ccaagcggcc ggagaacctg cgtgcaatcc
     2101 atcttgttca atcatgcgaa acgatcctca tcctgtctct tgatcagatc ttgatcccct
     2161 gcgccatcag atccttggcg gcaagaaagc catccagttt actttgcagg gcttcccaac
     2221 cttaccagag ggcgccccag ctggcaattc cgacgtctaa gaaaccatta ttatcatgac
     2281 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcgggccc attaagttct
     2341 gtgctaggag gtgactgaag tatattttag gaattctaaa gatctttgac agctagctca
     2401 gtcctaggta taatactagt tgagaccagt ctaggtctcg gttttagagc tagaaatagc
     2461 aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt
     2521 tttggtag
//
