LOCUS       sequence-496424-        4440 bp    DNA     circular SYN 07-JAN-2026
DEFINITION  Expresses Non-targeting sgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4440)
  AUTHORS   Gupta D, Patinios C, Bassett HV, Kibe A, Collins SP, Kamm C, Wang Y,
            Zhao C, Vollen K, Toussaint C, Calvin I, Cullot G, Aird EJ, Polkoff 
            KM, Nguyen-Vo TP, Migur A, Schut F, Al'Abri IS, Achmedov T, Del Re 
            A, Corn JE, Saliba AE, Crook N, Stepanova AN, Alonso JM, Beisel CL
  TITLE     Targeted DNA ADP-ribosylation triggers templated repair in bacteria 
            and base mutagenesis in eukaryotes.
  JOURNAL   Nat Biotechnol. 2025 Sep 4. doi: 10.1038/s41587-025-02802-w.
  PUBMED    40908325
REFERENCE   2  (bases 1 to 4440)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 7, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2025 Sep 4. doi: 10.1038/s41587-025-02802-w."
COMMENT     Sequence Label: pCBS-5737
FEATURES             Location/Qualifiers
     source          1..4440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     448..467
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        661..929
                     /label=SNR52 promoter
                     /note="promoter for the S. cerevisiae small nucleolar RNA 
                     gene SNR52"
     misc_RNA        950..1025
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        1378..1479
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             1480..2154
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     rep_origin      2224..3206
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     CDS             complement(join(3331..4122,4123..4191))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3954..3973
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4192..4296)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(4399..4440,1..547)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
       61 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
      121 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
      181 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
      241 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
      301 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
      361 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
      421 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
      481 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
      541 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
      601 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtgcgg gctgccaatg
      661 tctttgaaaa gataatgtat gattatgctt tcactcatat ttatacagaa acttgatgtt
      721 ttctttcgag tatatacaag gttattacat gtacgtttga agtacaactc tagattttgt
      781 agtgccctct tgggctagcg gtaaaggtgc gcattttttc acaccctaca atgttctgtt
      841 caaaagattt tggtcaaacg ctgtagaagt gaaagttggt gcgcatgttt cggcgttcga
      901 aacttctccg cagtgaaaga taaatgatcg agtgatcgta gccatccagg ttttagagct
      961 agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
     1021 ggtgcttttg ttttttatgt ctggactgac agctgtggta gagccacaaa cagccggtac
     1081 aagcaacgat ctccaggacc atctgaatca tgcgcggatg acacgaactc acgacggcga
     1141 tcacagacat taacccacag tacagacact gcgacaacgt ggcaattcgt cgcaatactg
     1201 gccaacgggg tcatcacggc tcatcatgcg ccaaacaaat gtgtgcaata cacgctcgga
     1261 tgactgcatg atgaccgcac tgactgggga cagcagatcc acctaagcct gtgagagaag
     1321 cagacacccg acagatcaag gcagttaact agtgcactgc agtacagccg attaagaatt
     1381 ccggtcgaaa aaagaaaagg agagggccaa gagggagggc attggtgact attgagcacg
     1441 tgagtatacg tgattaagca cacaaaggca gcttggagta tgtctgttat taatttcaca
     1501 ggtagttctg gtccattggt gaaagtttgc ggcttgcaga gcacagaggc cgcagaatgt
     1561 gctctagatt ccgatgctga cttgctgggt attatatgtg tgcccaatag aaagagaaca
     1621 attgacccgg ttattgcaag gaaaatttca agtcttgtaa aagcatataa aaatagttca
     1681 ggcactccga aatacttggt tggcgtgttt cgtaatcaac ctaaggagga tgttttggct
     1741 ctggtcaatg attacggcat tgatatcgtc caactgcatg gagatgagtc gtggcaagaa
     1801 taccaagagt tcctcggttt gccagttatt aaaagactcg tatttccaaa agactgcaac
     1861 atactactca gtgcagcttc acagaaacct cattcgttta ttcccttgtt tgattcagaa
     1921 gcaggtggga caggtgaact tttggattgg aactcgattt ctgactgggt tggaaggcaa
     1981 gagagccccg aaagcttaca ttttatgtta gctggtggac tgacgccaga aaatgttggt
     2041 gatgcgctta gattaaatgg cgttattggt gttgatgtaa gcggaggtgt ggagacaaat
     2101 ggtgtaaaag actctaacaa aatagcaaat ttcgtcaaaa atgctaagaa ataggttatt
     2161 actgagtagt atttatttaa gtattgtttg tgcacttgcc tgcaggcctt ttgaaaagcg
     2221 agtaacgaag catctgtgct tcattttgta gaacaaaaat gcaacgcgag agcgctaatt
     2281 tttcaaacaa agaatctgag ctgcattttt acagaacaga aatgcaacgc gaaagcgcta
     2341 ttttaccaac gaagaatctg tgcttcattt ttgtaaaaca aaaatgcaac gcgagagcgc
     2401 taatttttca aacaaagaat ctgagctgca tttttacaga acagaaatgc aacgcgagag
     2461 cgctatttta ccaacaaaga atctatactt cttttttgtt ctacaaaaat gcatcccgag
     2521 agcgctattt ttctaacaaa gcatcttaga ttactttttt tctcctttgt gcgctctata
     2581 atgcagtctc ttgataactt tttgcactgt aggtccgtta aggttagaag aaggctactt
     2641 tggtgtctat tttctcttcc ataaaaaaag cctgactcca cttcccgcgt ttactgatta
     2701 ctagcgaagc tgcgggtgca ttttttcaag ataaaggcat ccccgattat attctatacc
     2761 gatgtggatt gcgcatactt tgtgaacaga aagtgatagc gttgatgatt cttcattggt
     2821 cagaaaatta tgaacggttt cttctatttt gtctctatat actacgtata ggaaatgttt
     2881 acattttcgt attgttttcg attcactcta tgaatagttc ttactacaat ttttttgtct
     2941 aaagagtaat actagagata aacataaaaa atgtagaggt cgagtttaga tgcaagttca
     3001 aggagcgaaa ggtggatggg taggttatat agggatatag cacagagata tatagcaaag
     3061 agatactttt gagcaatgtt tgtggaagcg gtattcgcaa tattttagta gctcgttaca
     3121 gtccggtgcg tttttggttt tttgaaagtg cgtcatcaga gcgcttttgg ttttcaaaag
     3181 cgctctgaag ttcctatact ttctagctag agaataggaa cttcccgagc ggccgcgatt
     3241 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     3301 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     3361 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     3421 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gggacccacg
     3481 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     3541 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     3601 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     3661 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     3721 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     3781 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     3841 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     3901 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     3961 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     4021 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     4081 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     4141 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     4201 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     4261 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     4321 tgtcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     4381 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
//
