LOCUS       sequence-368138-        2300 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pegRNA plasmid in order to make a Y66H (TAC>CAT) conversion in the 
            GFP gene, creating a BFP gene..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2300)
  AUTHORS   Schene IF, Joore IP, Baijens JHL, Stevelink R, Kok G, Shehata S, 
            Ilcken EF, Nieuwenhuis ECM, Bolhuis DP, van Rees RCM, Spelier SA, 
            van der Doef HPJ, Beekman JM, Houwen RHJ, Nieuwenhuis EES, Fuchs SA
  TITLE     Mutation-specific reporter for optimization and enrichment of prime 
            editing.
  JOURNAL   Nat Commun. 2022 Mar 1;13(1):1028. doi: 10.1038/s41467-022-28656-3.
  PUBMED    35232966
REFERENCE   2  (bases 1 to 2300)
  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: "2022-03-1"; volume: "13(1):1028. doi"; pages: " 
            10.1038/s41467-022-28656-3"
COMMENT     Sequence Label: pegRNA-GFP>BFP_Y66H-NGG
FEATURES             Location/Qualifiers
     source          1..2300
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     430..449
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(668..772)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        879..1127
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     879..899
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1050..1069
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1148..1223
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     rep_origin      complement(1348..1936)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1428..1447)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2107..2300,1..598,599..667))
                     /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"
ORIGIN
        1 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
       61 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
      121 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
      181 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
      241 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
      301 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
      361 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
      421 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
      481 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
      541 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
      601 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
      661 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
      721 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
      781 cccgaaaagt gccacctgac gtcgctagct gtacaaaaaa gcaggcttta aaggaaccaa
      841 ttcagtcgac tggatccggt accaaggtcg ggcaggaaga gggcctattt cccatgattc
      901 cttcatattt gcatatacga tacaaggctg ttagagagat aattagaatt aatttgactg
      961 taaacacaaa gatattagta caaaatacgt gacgtagaaa gtaataattt cttgggtagt
     1021 ttgcagtttt aaaattatgt tttaaaatgg actatcatat gcttaccgta acttgaaagt
     1081 atttcgattt cttggcttta tatatcttgt ggaaaggacg aaacaccgct gaagcactgc
     1141 acgccgtgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg
     1201 aaaaagtggc accgagtcgg tgcctgaccc atggcgtgca gtgctttttt taagcttggg
     1261 ccgctcgagg tacctctcta catatgacat gtgagcaaaa ggccagcaaa aggccaggaa
     1321 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     1381 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     1441 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     1501 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     1561 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     1621 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     1681 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     1741 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     1801 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     1861 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     1921 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     1981 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     2041 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     2101 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     2161 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     2221 tggccccagt gctgcaatga taccgcgaga tccacgctca ccggctccag atttatcagc
     2281 aataaaccag ccagccggaa
//
