LOCUS       sequence-235255-        2350 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express sgRNA, p15A, CRM.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2350)
  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 2350)
  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: psgRNAc
FEATURES             Location/Qualifiers
     source          1..2350
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     134..153
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(220..322)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(714..731)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(848..1393)
                     /direction=LEFT
                     /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     complement(928..947)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        1510..1544
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        1565..1640
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             complement(join(1910..2350,1..219))
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
ORIGIN
        1 gaattccgga tgagcattca tcaggcgggc aagaatgtga ataaaggccg gataaaactt
       61 gtgcttattt ttctttacgg tctttaaaaa ggccgtaata tccagctgaa cggtctggtt
      121 ataggtacat tgagcaactg actgaaatgc ctcaaaatgt tctttacgat gccattggga
      181 tatatcaacg gtggtatatc cagtgatttt tttctccatt ttagcttcct tagctcctga
      241 aaatctcgat aactcaaaaa atacgcccgg tagtgatctt atttcattat ggtgaaagtt
      301 ggaacctctt acgtgccgat caacgtctca ttttcgccaa aagttggccc ggggcttccc
      361 ggtatcaaca gggacaccag gatttattta ttctgcgaag tgatcttccg tcacaggtat
      421 ttattcggcg caaagtgcgt cgggtgatgc tgccaactta ctgatttagt gtatgatggt
      481 gtttttgagg tgctccagtg gcttctgttt ctatcagctg tccctcctgt tcagctactg
      541 acggggtggt gcgtaacggc aaaagcaccg ccggacatca gcgctagcgg agtgtatact
      601 ggcttactat gttggcactg atgagggtgt cagtgaagtg cttcatgtgg caggagaaaa
      661 aaggctgcac cggtgcgtca gcagaatatg tgatacagga tatattccgc ttcctcgctc
      721 actgactcgc tacgctcggt cgttcgactg cggcgagcgg aaatggctta cgaacggggc
      781 ggagatttcc tggaagatgc caggaagata cttaacaggg aagtgagagg gccgcggcaa
      841 agccgttttt ccataggctc cgcccccctg acaagcatca cgaaatctga cgctcaaatc
      901 agtggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggcggctccc
      961 tcgtgcgctc tcctgttcct gcctttcggt ttaccggtgt cattccgctg ttatggccgc
     1021 gtttgtctca ttccacgcct gacactcagt tccgggtagg cagttcgctc caagctggac
     1081 tgtatgcacg aaccccccgt tcagtccgac cgctgcgcct tatccggtaa ctatcgtctt
     1141 gagtccaacc cggaaagaca tgcaaaagca ccactggcag cagccactgg taattgattt
     1201 agaggagtta gtcttgaagt catgcgccgg ttaaggctaa actgaaagga caagttttgg
     1261 tgactgcgct cctccaagcc agttacctcg gttcaaagag ttggtagctc agagaacctt
     1321 cgaaaaaccg ccctgcaagg cggttttttc gttttcagag caagagatta cgcgcagacc
     1381 aaaacgatct caagaagatc atcttattaa tcagataaaa tatttctaga tttcagtgca
     1441 atttatctct tcaaatgtag cacctggcta ggaggtgact gaagtatatt ttaggaattc
     1501 taaagatctt tgacagctag ctcagtccta ggtataatac tagttgagac cagtctaggt
     1561 ctcggtttta gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa
     1621 aagtggcacc gagtcggtgc tttttttggt agtgcagcgc gatcgtaatc aggggggaga
     1681 gcctgagcaa actggcctca ggcatttgag aagcacacgg tcacactgct tccggtagtc
     1741 aataaaccgg taaaccagca atagacataa gcggctattt aacgaccctg ccctgaaccg
     1801 acgaccgggt cgaatttgct ttcgaatttc tgccattcat ccgcttatta tcacttattc
     1861 aggcgtagca accaggcgtt taagggcacc aataactgcc ttaaaaaaat tacgccccgc
     1921 cctgccactc atcgcagtac tgttgtaatt cattaagcat tctgccgaca tggaagccat
     1981 cacaaacggc atgatgaacc tgaatcgcca gcggcatcag caccttgtcg ccttgcgtat
     2041 aatatttgcc catggtgaaa acgggggcga agaagttgtc catattggcc acgtttaaat
     2101 caaaactggt gaaactcacc cagggattgg ctgagacgaa aaacatattc tcaataaacc
     2161 ctttagggaa ataggccagg ttttcaccgt aacacgccac atcttgcgaa tatatgtgta
     2221 gaaactgccg gaaatcgtcg tggtattcac tccagagcga tgaaaacgtt tcagtttgct
     2281 catggaaaac ggtgtaacaa gggtgaacac tatcccatat caccagctca ccgtctttca
     2341 ttgccatacg
//
