LOCUS       sequence-153105-        3262 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPaint gene tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3262)
  AUTHORS   Schmid-Burgk JL, Honing K, Ebert TS, Hornung V
  TITLE     CRISPaint allows modular base-specific gene tagging using a 
            ligase-4-dependent mechanism.
  JOURNAL   Nat Commun. 2016 Jul 28;7:12338. doi: 10.1038/ncomms12338.
  PUBMED    27465542
REFERENCE   2  (bases 1 to 3262)
  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 
            Commun."; date: "2016-07-28"; volume: "7:12338. doi"; pages: " 
            10.1038/ncomms12338"
COMMENT     Sequence Label: pCRISPaint-3xflag-PuroR
FEATURES             Location/Qualifiers
     source          1..3262
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..49
                     /note="synthetic polyadenylation signal"
     misc_feature    63..154
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     103..122
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     CDS             218..283
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             293..346
                     /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    970..1091
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(981..1000)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1035..1054
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1339..1356)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1510..2098)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1590..1609)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2298..3089,3090..3158))
                     /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 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctcttaatga tagctagcgg gccagtaccc
      181 aaaaagcggg gggtctggtg gcagtggagg gggatccgat tacaaggacc acgacggcga
      241 ttataaggac catgatattg attataagga cgatgacgat aagtccggag gagagggcag
      301 aggaagtctg ctaacatgcg gtgacgtcga ggagaatcct ggcccaatga ctgaatacaa
      361 gcctactgtc aggttggcta caagagacga cgttcctaga gccgtgagaa ctctggctgc
      421 agccttcgcc gactaccccg ccacgagaca caccgttgac ccagatcggc atattgagag
      481 agtgactgaa ctgcaggagc tgtttcttac aagagttggc ctcgacatag gcaaggtgtg
      541 ggtggcggac gacggcgccg ccgtggccgt ctggaccact cccgaatcag ttgaggctgg
      601 cgccgtattc gctgagatcg gcccgagaat ggctgagctc agcgggagta ggctcgcggc
      661 acagcagcaa atggagggac tgctggcacc acacaggccc aaagaacccg cctggttcct
      721 ggcaaccgtc ggtgtatctc ccgatcatca ggggaaaggt ctgggctctg ccgtagtgct
      781 ccctggcgtg gaggcagctg agagagcagg agtacctgcc ttcttggaga cctccgctcc
      841 aaggaatctt cccttctatg aacggttggg cttcaccgtg acagccgacg tggaagtccc
      901 cgaaggcccc cgcacttggt gcatgacgag gaagcctgga gcgtgagggc ccgcttcgag
      961 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     1021 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     1081 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     1141 gggaggtttt ttaaagcaag taaaacctct acaaatgtgg taaaatcgat aaagatccgt
     1201 cgaccgatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg
     1261 actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg
     1321 gcagcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     1381 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     1441 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     1501 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     1561 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     1621 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     1681 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     1741 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     1801 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     1861 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     1921 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     1981 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     2041 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     2101 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     2161 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     2221 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagcg gccgcaaatg
     2281 ctaaaccact gcagtggtta ccagtgcttg atcagtgagg caccgatctc agcgatctgc
     2341 ctatttcgtt cgtccatagt ggcctgactc cccgtcgtgt agatcactac gattcgtgag
     2401 ggcttaccat caggccccag cgcagcaatg atgccgcgag agccgcgttc accggccccc
     2461 gatttgtcag caatgaacca gccagcaggg agggccgagc gaagaagtgg tcctgctact
     2521 ttgtccgcct ccatccagtc tatgagctgc tgtcgtgatg ctagagtaag aagttcgcca
     2581 gtgagtagtt tccgaagagt tgtggccatt gctactggca tcgtggtatc acgctcgtcg
     2641 ttcggtatgg cttcgttcaa ctctggttcc cagcggtcaa gccgggtcac atgatcaccc
     2701 atattatgaa gaaatgcagt cagctcctta gggcctccga tcgttgtcag aagtaagttg
     2761 gccgcggtgt tgtcgctcat ggtaatggca gcactacaca attctcttac cgtcatgcca
     2821 tccgtaagat gcttttccgt gaccggcgag tactcaacca agtcgttttg tgagtagtgt
     2881 atacggcgac caagctgctc ttgcccggcg tctatacggg acaacaccgc gccacatagc
     2941 agtactttga aagtgctcat catcgggaat cgttcttcgg ggcggaaaga ctcaaggatc
     3001 ttgccgctat tgagatccag ttcgatatag cccactcttg cacccagttg atcttcagca
     3061 tcttttactt tcaccagcgt ttcggggtgt gcaaaaacag gcaagcaaaa tgccgcaaag
     3121 aagggaatga gtgcgacacg aaaatgttgg atgctcatac tcgtcctttt tcaatattat
     3181 tgaagcattt atcagggtta ctagtacgtc tctcaaggat aagtaagtaa tattaaggta
     3241 cgggaggtat tggacaggcc gc
//
