LOCUS       sequence-410551-        5892 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor plasmid for Frame 0 with mCherry tag and triple Puromycin 
            resistant marker (3xPuroR).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5892)
  AUTHORS   Kim J, Kratz AF, Chen S, Sheng J, Kim HK, Zhang L, Singh BK, Chavez 
            A
  TITLE     High-throughput tagging of endogenous loci for rapid 
            characterization of protein function.
  JOURNAL   Sci Adv. 2024 May 3;10(18):eadg8771. doi: 10.1126/sciadv.adg8771. 
            Epub 2024 May 1.
  PUBMED    38691600
REFERENCE   2  (bases 1 to 5892)
  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: "Sci Adv."; 
            date: "2024-05-3"; volume: "10(18):eadg8771. doi"; pages: " 
            10.1126/sciadv.adg8771. Epub 2024 May 1"
COMMENT     Sequence Label: pDNR-mCherry-T2A-3xPuroR-Frame 0
FEATURES             Location/Qualifiers
     source          1..5892
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    166..300
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    420..554
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    674..808
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             907..1611
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1049..1067)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1317..1336
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1621..1674
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2272..2325
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2923..2976
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     polyA_signal    3600..3721
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3969..3986)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4140..4728)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4220..4239)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4928..5719,5720..5788))
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 cgcgatgaat aaatgaaagc ttgcagatct gcgactctag aggatctgcg actctagagg
       61 atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac
      121 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca
      181 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
      241 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc
      301 tgcgactcta gaggatcata atcagccata ccacatttgt agaggtttta cttgctttaa
      361 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
      421 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
      481 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
      541 atcatgtctg gatctgcgac tctagaggat cataatcagc cataccacat ttgtagaggt
      601 tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc
      661 aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
      721 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
      781 catcaatgta tcttatcatg tctggatccc catcaagctg atccggaact aactaagtaa
      841 gctagcgaat cgactcgaac ttcgtgtgcg agtggcgggt ctggtggcag tggaggggga
      901 tccctggtga gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc
      961 aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag
     1021 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg
     1081 cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag
     1141 caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag
     1201 cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag
     1261 gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc
     1321 gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac
     1381 ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac
     1441 gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac
     1501 gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac
     1561 gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa gtccggagga
     1621 gagggcagag gaagtctgct aacatgcggt gacgtagaag aaaacccagg gcctctgact
     1681 gaatacaagc ctactgtcag gttggctaca agagacgacg ttcctagagc cgtgagaact
     1741 ctggctgcag ccttcgccga ctaccccgcc acgagacaca ccgttgaccc agatcggcat
     1801 attgagagag tgactgaact gcaggagctg tttcttacaa gagttggcct cgacataggc
     1861 aaggtgtggg tggcggacga cggcgccgcc gtggccgtct ggaccactcc cgaatcagtt
     1921 gaggctggcg ccgtattcgc tgagatcggc ccgagaatgg ctgagctcag cgggagtagg
     1981 ctcgcggcac agcagcaaat ggagggactg ctggcaccac acaggcccaa agaacccgcc
     2041 tggttcctgg caaccgtcgg tgtatctccc gatcatcagg ggaaaggtct gggctctgcc
     2101 gtagtgctcc ctggcgtgga ggcagctgag agagcaggag tacctgcctt cttggagacc
     2161 tccgctccaa ggaatcttcc cttctatgaa cggttgggct tcaccgtgac agccgacgtg
     2221 gaagtccccg aaggcccccg cacttggtgc atgacgagga agcctggagc ggaaggtaga
     2281 ggttctttgc ttacctgcgg ggacgtggaa gagaacccag gcccactgac tgaatacaag
     2341 cctactgtca ggttggctac aagagacgac gttcctagag ccgtgagaac tctggctgca
     2401 gccttcgccg actaccccgc cacgagacac accgttgacc cagatcggca tattgagaga
     2461 gtgactgaac tgcaggagct gtttcttaca agagttggcc tcgacatagg caaggtgtgg
     2521 gtggcggacg acggcgccgc cgtggccgtc tggaccactc ccgaatcagt tgaggctggc
     2581 gccgtattcg ctgagatcgg cccgagaatg gctgagctca gcgggagtag gctcgcggca
     2641 cagcagcaaa tggagggact gctggcacca cacaggccca aagaacccgc ctggttcctg
     2701 gcaaccgtcg gtgtatctcc cgatcatcag gggaaaggtc tgggctctgc cgtagtgctc
     2761 cctggcgtgg aggcagctga gagagcagga gtacctgcct tcttggagac ctccgctcca
     2821 aggaatcttc ccttctatga acggttgggc ttcaccgtga cagccgacgt ggaagtcccc
     2881 gaaggccccc gcacttggtg catgacgagg aagcctggag cggagggccg agggtcactc
     2941 cttacttgtg gtgatgtcga ggagaatcct ggcccactga ctgaatacaa gcctactgtc
     3001 aggttggcta caagagacga cgttcctaga gccgtgagaa ctctggctgc agccttcgcc
     3061 gactaccccg ccacgagaca caccgttgac ccagatcggc atattgagag agtgactgaa
     3121 ctgcaggagc tgtttcttac aagagttggc ctcgacatag gcaaggtgtg ggtggcggac
     3181 gacggcgccg ccgtggccgt ctggaccact cccgaatcag ttgaggctgg cgccgtattc
     3241 gctgagatcg gcccgagaat ggctgagctc agcgggagta ggctcgcggc acagcagcaa
     3301 atggagggac tgctggcacc acacaggccc aaagaacccg cctggttcct ggcaaccgtc
     3361 ggtgtatctc ccgatcatca ggggaaaggt ctgggctctg ccgtagtgct ccctggcgtg
     3421 gaggcagctg agagagcagg agtacctgcc ttcttggaga cctccgctcc aaggaatctt
     3481 cccttctatg aacggttggg cttcaccgtg acagccgacg tggaagtccc cgaaggcccc
     3541 cgcacttggt gcatgacgag gaagcctgga gcgtgagggc ccgcttcgag cagacatgat
     3601 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     3661 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     3721 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt
     3781 ttaaagcaag taaaacctct acaaatgtgg taaaatcgat aaagatccgt cgaccgatgc
     3841 ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg
     3901 ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg gcagcgctct
     3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagcg gccgcaaatg ctaaaccact
     4921 gcagtggtta ccagtgcttg atcagtgagg caccgatctc agcgatctgc ctatttcgtt
     4981 cgtccatagt ggcctgactc cccgtcgtgt agatcactac gattcgtgag ggcttaccat
     5041 caggccccag cgcagcaatg atgccgcgag agccgcgttc accggccccc gatttgtcag
     5101 caatgaacca gccagcaggg agggccgagc gaagaagtgg tcctgctact ttgtccgcct
     5161 ccatccagtc tatgagctgc tgtcgtgatg ctagagtaag aagttcgcca gtgagtagtt
     5221 tccgaagagt tgtggccatt gctactggca tcgtggtatc acgctcgtcg ttcggtatgg
     5281 cttcgttcaa ctctggttcc cagcggtcaa gccgggtcac atgatcaccc atattatgaa
     5341 gaaatgcagt cagctcctta gggcctccga tcgttgtcag aagtaagttg gccgcggtgt
     5401 tgtcgctcat ggtaatggca gcactacaca attctcttac cgtcatgcca tccgtaagat
     5461 gcttttccgt gaccggcgag tactcaacca agtcgttttg tgagtagtgt atacggcgac
     5521 caagctgctc ttgcccggcg tctatacggg acaacaccgc gccacatagc agtactttga
     5581 aagtgctcat catcgggaat cgttcttcgg ggcggaaaga ctcaaggatc ttgccgctat
     5641 tgagatccag ttcgatatag cccactcttg cacccagttg atcttcagca tcttttactt
     5701 tcaccagcgt ttcggggtgt gcaaaaacag gcaagcaaaa tgccgcaaag aagggaatga
     5761 gtgcgacacg aaaatgttgg atgctcatac tcgtcctttt tcaatattat tgaagcattt
     5821 atcagggtta ctagtacgtc tctcaaggat aagtaagtaa tattaaggta cgggaggtat
     5881 tggacaggcc gc
//
