LOCUS       sequence-273628-        2469 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Subcloning and expression of non-targeting sgRNA for use in yeast 
            dual reporter system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2469)
  AUTHORS   Standage-Beier K, Brookhouser N, Balachandran P, Zhang Q, Brafman 
            DA, Wang X
  TITLE     RNA-Guided Recombinase-Cas9 Fusion Targets Genomic DNA Deletion and 
            Integration.
  JOURNAL   CRISPR J. 2019 Aug;2:209-222. doi: 10.1089/crispr.2019.0013.
  PUBMED    31436506
REFERENCE   2  (bases 1 to 2469)
  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: "CRISPR J.";
            date: "2019-08"; volume: "2:209-222. doi"; pages: " 
            10.1089/crispr.2019.0013"
COMMENT     Sequence Label: pSB1C3-ysg(NT)
FEATURES             Location/Qualifiers
     source          1..2469
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     40..58
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(62..105)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     promoter        114..382
                     /label=SNR52 promoter
                     /note="promoter for the S. cerevisiae small nucleolar RNA 
                     gene SNR52"
     misc_RNA        401..476
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      550..607
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(631..648)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(802..1390)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(882..901)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1573..1667)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1688..2347)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2262..2281
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggcag
       61 aatttcagat aaaaaaaatc cttagctttc gctaaggatg atttctggaa ttctctttga
      121 aaagataatg tatgattatg ctttcactca tatttataca gaaacttgat gttttctttc
      181 gagtatatac aaggtgatta catgtacgtt tgaagtacaa ctctagattt tgtagtgccc
      241 tcttgggcta gcggtaaagg tgcgcatttt ttcacaccct acaatgttct gttcaaaaga
      301 ttttggtcaa acgctgtaga agtgaaagtt ggtgcgcatg tttcggcgtt cgaaacttct
      361 ccgcagtgaa agataaatga tcagaagagc gagctcttct gttttagagc tagaaatagc
      421 aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt
      481 tttactagat aattggagac cgagctggtc tctctaggat gtgcactagt agagctgcgg
      541 ccgctgcagt ccggcaaaaa agggcaaggt gtcaccaccc tgcccttttt ctttaaaacc
      601 gaaaagatta cttcgcgtta tgcaggcttc ctcgctcact gactcgctgc gctcggtcgt
      661 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
      721 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
      781 aaaggccgcg ttgctggcgt ttttccacag gctccgcccc cctgacgagc atcacaaaaa
      841 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
      901 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
      961 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     1021 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     1081 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     1141 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     1201 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     1261 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     1321 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     1381 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     1441 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     1501 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     1561 ctcgaggctt ggattctcac caataaaaaa cgcccggcgg caaccgagcg ttctgaacaa
     1621 atccagatgg agttctgagg tcattactgg atctatcaac aggagtccaa gcgagctcga
     1681 tatcaaatta cgccccgccc tgccactcat cgcagtactg ttgtaattca ttaagcattc
     1741 tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc ggcatcagca
     1801 ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag aagttgtcca
     1861 tattggccac gtttaaatca aaactggtga aactcaccca gggattggct gagacgaaaa
     1921 acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa cacgccacat
     1981 cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc cagagcgatg
     2041 aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta tcccatatca
     2101 ccagctcacc gtctttcatt gccatacgaa attccggatg agcattcatc aggcgggcaa
     2161 gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc tttaaaaagg
     2221 ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac tgaaatgcct
     2281 caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca gtgatttttt
     2341 tctccatttt agcttcctta gctcctgaaa atctcgataa ctcaaaaaat acgcccggta
     2401 gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgcccgatc aactcgagtg
     2461 ccacctgac
//
