LOCUS       sequence-153094-        3706 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 3706)
  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 3706)
  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-NanoLuc-PuroR
FEATURES             Location/Qualifiers
     source          1..3706
                     /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..727
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     CDS             737..790
                     /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    1414..1535
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1425..1444)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1479..1498
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1783..1800)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1954..2542)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2034..2053)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2742..3533,3534..3602))
                     /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 gggatccgtc ttcacactcg aagatttcgt
      241 tggggactgg cgacagacag ccggctacaa cctggaccaa gtccttgaac agggaggtgt
      301 gtccagtttg tttcagaatc tcggggtgtc cgtaactccg atccaaagga ttgtcctgag
      361 cggtgaaaat gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag gtctgagcgg
      421 cgaccaaatg ggccagatcg aaaaaatttt taaggtggtg taccctgtgg atgatcatca
      481 ctttaaggtg atcctgcact atggcacact ggtaatcgac ggggttacgc cgaacatgat
      541 cgactatttc ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa agatcactgt
      601 aacagggacc ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca accccgacgg
      661 ctccctgctg ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt gcgaacgcat
      721 tctggcgtcc ggaggagagg gcagaggaag tctgctaaca tgcggtgacg tcgaggagaa
      781 tcctggccca atgactgaat acaagcctac tgtcaggttg gctacaagag acgacgttcc
      841 tagagccgtg agaactctgg ctgcagcctt cgccgactac cccgccacga gacacaccgt
      901 tgacccagat cggcatattg agagagtgac tgaactgcag gagctgtttc ttacaagagt
      961 tggcctcgac ataggcaagg tgtgggtggc ggacgacggc gccgccgtgg ccgtctggac
     1021 cactcccgaa tcagttgagg ctggcgccgt attcgctgag atcggcccga gaatggctga
     1081 gctcagcggg agtaggctcg cggcacagca gcaaatggag ggactgctgg caccacacag
     1141 gcccaaagaa cccgcctggt tcctggcaac cgtcggtgta tctcccgatc atcaggggaa
     1201 aggtctgggc tctgccgtag tgctccctgg cgtggaggca gctgagagag caggagtacc
     1261 tgccttcttg gagacctccg ctccaaggaa tcttcccttc tatgaacggt tgggcttcac
     1321 cgtgacagcc gacgtggaag tccccgaagg cccccgcact tggtgcatga cgaggaagcc
     1381 tggagcgtga gggcccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
     1441 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     1501 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     1561 tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat
     1621 gtggtaaaat cgataaagat ccgtcgaccg atgcccttga gagccttcaa cccagtcagc
     1681 tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc
     1741 atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct
     1801 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     1861 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     1921 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     1981 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2041 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2101 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2161 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2221 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2281 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2341 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     2401 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2461 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2521 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2581 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     2641 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     2701 ttggtctgac agcggccgca aatgctaaac cactgcagtg gttaccagtg cttgatcagt
     2761 gaggcaccga tctcagcgat ctgcctattt cgttcgtcca tagtggcctg actccccgtc
     2821 gtgtagatca ctacgattcg tgagggctta ccatcaggcc ccagcgcagc aatgatgccg
     2881 cgagagccgc gttcaccggc ccccgatttg tcagcaatga accagccagc agggagggcc
     2941 gagcgaagaa gtggtcctgc tactttgtcc gcctccatcc agtctatgag ctgctgtcgt
     3001 gatgctagag taagaagttc gccagtgagt agtttccgaa gagttgtggc cattgctact
     3061 ggcatcgtgg tatcacgctc gtcgttcggt atggcttcgt tcaactctgg ttcccagcgg
     3121 tcaagccggg tcacatgatc acccatatta tgaagaaatg cagtcagctc cttagggcct
     3181 ccgatcgttg tcagaagtaa gttggccgcg gtgttgtcgc tcatggtaat ggcagcacta
     3241 cacaattctc ttaccgtcat gccatccgta agatgctttt ccgtgaccgg cgagtactca
     3301 accaagtcgt tttgtgagta gtgtatacgg cgaccaagct gctcttgccc ggcgtctata
     3361 cgggacaaca ccgcgccaca tagcagtact ttgaaagtgc tcatcatcgg gaatcgttct
     3421 tcggggcgga aagactcaag gatcttgccg ctattgagat ccagttcgat atagcccact
     3481 cttgcaccca gttgatcttc agcatctttt actttcacca gcgtttcggg gtgtgcaaaa
     3541 acaggcaagc aaaatgccgc aaagaaggga atgagtgcga cacgaaaatg ttggatgctc
     3601 atactcgtcc tttttcaata ttattgaagc atttatcagg gttactagta cgtctctcaa
     3661 ggataagtaa gtaatattaa ggtacgggag gtattggaca ggccgc
//
