LOCUS       sequence-110943-         978 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  leu2 deletion gRNA cassette carried by pRS42H.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 978)
  AUTHORS   Zhang GC, Kong II, Kim H, Liu JJ, Cate JH, Jin YS
  TITLE     Construction of a quadruple auxotrophic mutant of an industrial 
            polyploid saccharomyces cerevisiae strain by using RNA-guided Cas9 
            nuclease.
  JOURNAL   Appl Environ Microbiol. 2014 Dec;80(24):7694-701. doi: 
            10.1128/AEM.02310-14. Epub 2014 Oct 3.
  PUBMED    25281382
REFERENCE   2  (bases 1 to 978)
  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: 
            "10.1128/AEM.02310-14"; journalName: "Appl Environ Microbiol"; date:
            "2014-12"; volume: "80"; issue: "24"; pages: "7694-701"
COMMENT     Sequence Label: gRNA-leu-HYB
FEATURES             Location/Qualifiers
     source          1..978
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     protein_bind    95..116
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(131..136,137..154,155..161)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    169..185
                     /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)."
     primer_bind     174..196
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     193..209
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     193..209
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     228..248
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        230..248
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        267..535
                     /label=SNR52 promoter
                     /note="promoter for the S. cerevisiae small nucleolar RNA 
                     gene SNR52"
     misc_RNA        556..630
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      635..654
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     primer_bind     complement(668..687)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(669..687)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(697..714)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(697..713)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(706..728)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(942..961)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
       61 ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg
      121 caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat
      181 aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca attaaccctc
      241 actaaaggga acaaaagctg gagctctctt tgaaaagata atgtatgatt atgctttcac
      301 tcatatttat acagaaactt gatgttttct ttcgagtata tacaaggtga ttacatgtac
      361 gtttgaagta caactctaga ttttgtagtg ccctcttggg ctagcggtaa aggtgcgcat
      421 tttttcacac cctacaatgt tctgttcaaa agattttggt caaacgctgt agaagtgaaa
      481 gttggtgcgc atgtttcggc gttcgaaact tctccgcagt gaaagataaa tgatcaatca
      541 cagccgaagc cattagtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta
      601 tcaacttgaa aaagtggcac cgagtcggtg gtgctttttt tgttttttat gtctggtacc
      661 caattcgccc tatagtgagt cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg
      721 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
      781 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
      841 gaatggcgaa atggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
      901 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
      961 ttcctttctc gccacgtt
//
