LOCUS       sequence-137246-         981 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  for expression of gRNA in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 981)
  AUTHORS   Li Y, Lin Z, Huang C, Zhang Y, Wang Z, Tang YJ, Chen T, Zhao X
  TITLE     Metabolic engineering of Escherichia coli using CRISPR-Cas9 
            meditated genome editing.
  JOURNAL   Metab Eng. 2015 Sep;31:13-21. doi: 10.1016/j.ymben.2015.06.006. Epub
            2015 Jun 30.
  PUBMED    26141150
REFERENCE   2  (bases 1 to 981)
  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: "Metab 
            Eng."; date: "2015-09"; volume: "31:13-21. doi"; pages: " 
            10.1016/j.ymben.2015.06.006. Epub 2015 Jun 30"
COMMENT     Sequence Label: pGRB
FEATURES             Location/Qualifiers
     source          1..981
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     87..105
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        106..140
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     primer_bind     complement(164..186)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(281..598)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MITNSSSVPGDPLESTCRHASLALAVVLQRRDWENPGVTQLNRLA
                     AHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICSDA
                     A"
     primer_bind     286..305
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     499..521
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     513..530
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     514..530
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    534..590
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(599..615)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(599..615)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(612..634)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    623..639
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(647..653,654..671,672..677))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    692..713
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(829..846)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
       61 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcttgac agctagctca
      121 gtcctaggta taatactagt tttttcggtg atgacggtga aaacctctga cacatgcagc
      181 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
      241 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga
      301 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
      361 accgcatcag gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc
      421 gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt
      481 gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt gccaagcttg
      541 catgcctgca ggtcgactct agaggatccc cgggtaccga gctcgaattc gtaatcatgt
      601 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
      661 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
      721 tgcgctcact gcccgctttc cagtcgggaa acctgtcagt tttagagcta gaaatagcaa
      781 gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg cacccttcct cgctcactga
      841 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
      901 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
      961 aaaggccagg aaccgtaaaa a
//
