LOCUS       sequence-354058-        2305 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transiently expressing a pegRNA to introduce HEK3 CTT insertion in 
            human cells. It has a sgRNA scaffold from px330..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2305)
  AUTHORS   Li H, Busquets O, Verma Y, Syed KM, Kutnowski N, Pangilinan GR, 
            Gilbert LA, Bateup HS, Rio DC, Hockemeyer D, Soldner F
  TITLE     Highly efficient generation of isogenic pluripotent stem cell models
            using prime editing.
  JOURNAL   Elife. 2022 Sep 7;11:e79208. doi: 10.7554/eLife.79208.
  PUBMED    36069759
REFERENCE   2  (bases 1 to 2305)
  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: "Elife."; 
            date: "2022-09-7"; volume: "11:e79208. doi"; pages: " 
            10.7554/eLife.79208"
COMMENT     Sequence Label: pU6-pegRNA-HEK3-CTTins-px330-scaffold
FEATURES             Location/Qualifiers
     source          1..2305
                     /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        270..345
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     rep_origin      complement(475..1063)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(555..574)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1234..2025,2026..2094))
                     /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     1857..1876
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2095..2199)
                     /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 gcccagactg agcacgtgag ttttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgctctgc catcaaagcg
      361 tgctcagtct gtttttttaa gcttgggccg ctcgaggtac ctctctacat atgacatgtg
      421 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
      481 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
      541 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
      601 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
      661 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
      721 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
      781 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
      841 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
      901 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
      961 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     1021 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     1081 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     1141 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     1201 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     1261 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     1321 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagatcc
     1381 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     1441 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     1501 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     1561 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     1621 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     1681 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     1741 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     1801 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     1861 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     1921 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     1981 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     2041 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     2101 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     2161 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     2221 acctgacgtc gctagctgta caaaaaagca ggctttaaag gaaccaattc agtcgactgg
     2281 atccggtacc aaggtcgggc aggaa
//
