LOCUS       sequence-344973-        2337 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express forward pegRNA mediating 991-bp deletion and 44-bp 
            insertion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2337)
  AUTHORS   Jiang T, Zhang XO, Weng Z, Xue W
  TITLE     Deletion and replacement of long genomic sequences using prime 
            editing.
  JOURNAL   Nat Biotechnol. 2021 Oct 14. pii: 10.1038/s41587-021-01026-y. doi: 
            10.1038/s41587-021-01026-y.
  PUBMED    34650270
REFERENCE   2  (bases 1 to 2337)
  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 
            Biotechnol. 2021 Oct 14. pii: 10.1038/s41587-021-01026-y. doi: 
            10.1038/s41587-021-01026-y."
COMMENT     Sequence Label: HEK3_pegR_44bp
FEATURES             Location/Qualifiers
     source          1..2337
                     /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)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        271..346
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             365..388
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     rep_origin      complement(507..1095)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(587..606)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1266..2057,2058..2126))
                     /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     1889..1908
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2127..2231)
                     /gene="bla"
                     /label=AmpR promoter
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 tgatcacctg cccaaatgtg gttttagagc tagaaatagc aagttaaaat
      301 aaggctagtc cgttatcaac ttgaaaaagt gggaccgagt cggtgctagg gataacaggg
      361 taatgattac aaggatgacg acgataagtg atttgggcag gtgttttttt aagcttgggc
      421 cgctcgaggt acctctctac atatgacatg tgagcaaaag gccagcaaaa ggccaggaac
      481 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
      541 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
      601 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
      661 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
      721 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
      781 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
      841 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
      901 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
      961 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     1021 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     1081 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     1141 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     1201 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     1261 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     1321 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     1381 ggccccagtg ctgcaatgat accgcgagat ccacgctcac cggctccaga tttatcagca
     1441 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     1501 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     1561 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     1621 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     1681 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     1741 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     1801 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     1861 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     1921 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     1981 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     2041 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     2101 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     2161 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     2221 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgctagctg tacaaaaaag
     2281 caggctttaa aggaaccaat tcagtcgact ggatccggta ccaaggtcgg gcaggaa
//
