LOCUS       sequence-222318-        3959 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 3959)
  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 3959)
  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: pPRISM-Fuse-cmlc2-eGFP
FEATURES             Location/Qualifiers
     source          1..3959
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             31..705
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKMR
                     LKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHST
                     GA"
     CDS             31..705
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKMR
                     LKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHST
                     GA"
     primer_bind     complement(158..176)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     426..445
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     620..643
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             2468..2488
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(2492..2494,2495..2497,2498..3211)
                     /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(2492..2494,2495..2497,2498..3208)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2537..2558)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2798..2817)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3145..3166
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3216..3440
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     polyA_signal    3510..3591
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    3715..3796
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 cttctaggag gaggtagcgg aggtagcgga gcctcctccg aggacgtcat caaggagttc
       61 atgcgcttca aggtgcgcat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      121 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggc
      181 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tccagtacgg ctccaaggcc
      241 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      301 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      361 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
      421 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctccaccga gcggatgtac
      481 cccgaggacg gcgccctgaa gggcgagatc aagatgaggc tgaagctgaa ggacggcggc
      541 cactacgacg ccgaggtcaa gaccacctac atggccaaga agcccgtgca gctgcccggc
      601 gcctacaaga ccgacatcaa gctggacatc acctcccaca acgaggacta caccatcgtg
      661 gaacagtacg agcgcgccga gggccgccac tccaccggag cctaagacta gtaattaagt
      721 ctcagccacc gttaactgaa catgtcaaaa cctgtggaga ctgttgagat ttgatgttct
      781 gaaaagataa agcctataaa taaaatgttg cccaaatttc ctgcctgatg tttttctttg
      841 tctttgctac atggctttgc tgctcggatc ggctcactct gtgtatgcca cgttcacttt
      901 gtactctcct tctcacggta ggtttattat ttttagatgt gcagttagtt tctgtgaaat
      961 aacacaccac acactgatat tgtctgtgca ttgacttggt gagtgcacat tgtttttgat
     1021 cttgacatat ttatatttga ttgatcaggt gaactgtgtg aatctaaagt gctccataca
     1081 gatgttctgc attgaaaata ttctcatttt attagtggaa gtgagtgtat gccacatcca
     1141 atcaatttca gcaaacaccc cagtatgatt taatgcaaaa aaatgaaggt atcaaacacg
     1201 cattactact ttgcagttaa atatttaaca tttattccaa cacgaaaaaa agcagtaaat
     1261 aacactttga caaacacgtc aggacatctt atttttgtca ccctcacagg caatttagta
     1321 taatatatta tatatatata tatatcatat aataatattc agtataatat atatatatat
     1381 atcatattat aatattcagt ataatataaa acacaaacac atatatgtat aatataatat
     1441 aacattttta tttattgaga tgcctctatg gaccgtgtta taagaagtaa agatcaggag
     1501 aagtaaacat gaagtgtaat tatgaatact gatgttaaat taagctatga tgagttttca
     1561 ctgttaattt accatctcaa ttaaatgttg atgcctccat gaccaagtta agcagatgag
     1621 actgagacaa ctgtagaaga caagatgttc actttgctga atatagctgg cttgacagtt
     1681 atctatgact ctataaatat atatatattt ttttttttat aaaatgattt atttataact
     1741 atatatccat ttctcagaca ggtgcttcat atccctcact cccgtagctg tccatgctgg
     1801 atctgtcccc gttgttttta aaaagctaaa taagttatta acatgactgc atccagcgag
     1861 ccaaacctgt ctggtgtaca gctaccagag aagcttgaga tcctagtcac cggcggcatt
     1921 catccatcct tttcatccct caaatctctc attcacgtcc ccctccccat ctgcacactt
     1981 tatctcattt tccaccctgc tggaatctga gcacttgtgc agttatcagg gctcctgtat
     2041 ttaggaggct ctgggtgtcc atgtagggga cgaacagaaa cactgcagac ctttatagaa
     2101 gaacaaatga taagagtcct catacataaa gactccatta gaaacgtcag tgacccagga
     2161 gcccagacca acagcaaagc agacagtgaa ggatccacag cctgactaac tcgacaaacc
     2221 cccccaaacc taaggtgagt tgatctttaa gctttttaca ttttcagctc gcatatatca
     2281 attcgcgcgc attgtcacac tagcacccac tagcggtcag actgcagatt gcagcacgaa
     2341 acaggaagct gactccacat ggtcacatgc tcactgaagt gttgacttcc ctgacagctg
     2401 tgcactttct aaaccggttt tctcattcat ttacagttca gccggaattc acctcgagac
     2461 catggctcca aagaagaagc gtaaggtaac catggtgagc aagggcgagg agctgttcac
     2521 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     2581 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
     2641 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
     2701 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     2761 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     2821 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     2881 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     2941 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     3001 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     3061 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa
     3121 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     3181 cactctcggc atggacgagc tgtacaagta aagatctgtg ccttctagtt gccagccatc
     3241 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     3301 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     3361 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     3421 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc aaagggcgaa ttaattcatt
     3481 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
     3541 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
     3601 aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag
     3661 taaaacctct acaaatgtgg tcattgatga gtttggacaa accacaacta gaatgcagtg
     3721 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     3781 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     3841 ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtcatt cagatcaatt
     3901 aaccctcacc cgggctagcg atgcatggga gctctgacgt acgttgtcta gcaaggaag
//
