LOCUS       sequence-339521-        2706 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Control vector for coselection for prime editing in human cells. 
            Tandem expression of ATP1A1 Q118R-G4 and RUNX1 +4 T to G pegRNAs 
            from two independent U6 promoters..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2706)
  AUTHORS   Levesque S, Mayorga D, Fiset JP, Goupil C, Duringer A, Loiselle A, 
            Bouchard E, Agudelo D, Doyon Y
  TITLE     Marker-free co-selection for successive rounds of prime editing in 
            human cells.
  JOURNAL   Nat Commun. 2022 Oct 7;13(1):5909. doi: 10.1038/s41467-022-33669-z.
  PUBMED    36207338
REFERENCE   2  (bases 1 to 2706)
  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: "2022-10-7"; volume: "13(1):5909. doi"; pages: " 
            10.1038/s41467-022-33669-z"
COMMENT     Sequence Label: ATP1A1_G4_Q118R_RUNX1_+4_T_to_G_Dual_pegRNA
FEATURES             Location/Qualifiers
     source          1..2706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        270..345
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     rep_origin      complement(479..1067)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(559..578)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1238..2029,2030..2098))
                     /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     1861..1880
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2099..2203)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2266..2514
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        2535..2610
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg cattttcagg aggaagcgag ttttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgctgtct gaagccctcg
      361 cttcctcctg aaaatttttt ttaagcttgg gccgctcgag gtacctctct acatatgaca
      421 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
      481 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
      541 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
      601 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
      661 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
      721 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
      781 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
      841 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
      901 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
      961 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     1021 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     1081 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     1141 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     1201 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     1261 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     1321 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     1381 atccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     1441 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     1501 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     1561 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     1621 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     1681 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     1741 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     1801 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     1861 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     1921 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     1981 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     2041 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     2101 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     2161 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     2221 tgccacctga cgtcgctagc tgtacatacc aaggtcgggc aggaagaggg cctatttccc
     2281 atgattcctt catatttgca tatacgatac aaggctgtta gagagataat tagaattaat
     2341 ttgactgtaa acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt
     2401 gggtagtttg cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact
     2461 tgaaagtatt tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccgttcct
     2521 cttctgtagc agctgtttta gagctagaaa tagcaagtta aaataaggct agtccgttat
     2581 caacttgaaa aagtggcacc gagtcggtgc tagcatcaga gctgctacag aagagttttt
     2641 ttaagcttgg gccgctcgag gtacctctct acgtcgactg gatccggtac caaggtcggg
     2701 caggaa
//
