LOCUS       sequence-222430-        1855 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Donor for precise CRISPR directed genomic integration using the 
            GeneWeld method.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1855)
  AUTHORS   Wierson WA, Welker JM, Almeida MP, Mann CM, Webster DA, Torrie ME, 
            Weiss TJ, Kambakam S, Vollbrecht MK, Lan M, McKeighan KC, Levey J, 
            Ming Z, Wehmeier A, Mikelson CS, Haltom JA, Kwan KM, Chien CB, 
            Balciunas D, Ekker SC, Clark KJ, Webber BR, Moriarity BS, Solin SL, 
            Carlson DF, Dobbs DL, McGrail M, Essner J
  TITLE     Efficient targeted integration directed by short homology in 
            zebrafish and mammalian cells.
  JOURNAL   Elife. 2020 May 15;9. pii: 53968. doi: 10.7554/eLife.53968.
  PUBMED    32412410
REFERENCE   2  (bases 1 to 1855)
  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: "2020-05-15"; volume: "9. pii: 53968. doi"; pages: " 
            10.7554/eLife.53968"
COMMENT     Sequence Label: pGTag-NLS-eGFP-B-actin
FEATURES             Location/Qualifiers
     source          1..1855
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             10..66
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             73..93
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(94..96,97..99,100..813)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(94..96,97..99,100..810)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(139..160)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(400..419)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     747..768
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 ggatccggag ccacgaactt ctctctgtta aagcaagcag gagacgtgga agaaaacccc
       61 ggtcctatgg ctccaaagaa gaagcgtaag gtaatggtga gcaagggcga ggagctgttc
      121 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      181 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
      241 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
      301 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
      361 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
      421 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
      481 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
      541 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
      601 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
      661 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
      721 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
      781 atcactctcg gcatggacga gctgtacaag taactcgagg gcgcgccacg gactgttacc
      841 acttcacgcc gactcaaact gcgcagagaa acttcaaacg acaacattgg catggctttt
      901 gttatttttg gcgcttgact caggatctaa aaactggaac ggtgaaggtg acggcaatgt
      961 tttttggcaa ataagcatcc ccgaagttct acaatgcatc tgaggactcc atgttttgtt
     1021 ttctttagtc attccaaatg tttgttaaat gcattgttcc gaaacttatt tgcctctatg
     1081 aaggctgccc agtaattggg agcatactta acattgtagt attgtatgta aattatgtaa
     1141 caaaacaatg tctgggtttt tgtacttttc agccttaaaa tcttgggttt attttttttt
     1201 ctttgttcca aaaaaacgaa gctttaccat tcaagatgta aaggtttcca ttccccctgg
     1261 gcatattgta aaagctgtgt ggaacgtggc ggtgccagac atttggtggg gccaacctgt
     1321 acactgacta attcaattcc aataaaagtg cacatgtaag acatcctacc cctgtctcat
     1381 ttcctgtctt ggctctgagt ggcttcaata aaaatggctt ttgtgaagtt gcaccatgtt
     1441 aattacattt cagcactttc acaccaggtt tttgactaat ttgaatttat cgaagtttga
     1501 cctttttttt tttttttctt caagctaatt tgaggttatc aaaccatttc attggaggaa
     1561 acttaatgtt taaaagcata aattcgaaac atgattcatt tatttaactg cttaatggtg
     1621 aaacaaactt caagtaatac accaagtgtt ccctcatatg aaggtagcta actgcaaacg
     1681 gccttaactt tccatcttaa gttgagctac ataaaggctt taacaggaag gaactgccag
     1741 tttaaaagtc aaaccaccat gactgatata aattcttgtc catataacct tgcgtttcac
     1801 acaatatagc tatctactgc taaataaatt accatggcgt tgtctagcaa ggaag
//
