LOCUS       sequence-153096-        3517 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 3517)
  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 3517)
  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-ProteoTuner DD-PuroR
FEATURES             Location/Qualifiers
     source          1..3517
                     /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..538
                     /codon_start=1
                     /product="destabilization domain that can be stabilized by 
                     Shield1 in the ProteoTuner(TM) system"
                     /label=DD-C
                     /note="FKBP12 mutant optimized for use as a C-terminal tag"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLGDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQGAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     ELE"
     CDS             548..601
                     /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    1225..1346
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1236..1255)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1290..1309
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1594..1611)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1765..2353)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1845..1864)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2553..3344,3345..3413))
                     /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 gggatccgga gtgcaggtgg aaaccatctc
      241 cccaggagac gggcgcacct tccccaagcg cggccagacc tgcgtggtgc actacaccgg
      301 gatgcttgga gatggaaaga aagttgactc ctcccgggac agaaacaagc cctttaagtt
      361 tatgctaggc aagcaggagg tgatccgagg ctgggaagaa ggggttgccc agatgagtgt
      421 gggtcaggga gccaaactga ctatatctcc agattatgcc tatggtgcca ctgggcaccc
      481 aggcatcatc ccaccacatg ccactctcgt cttcgatgtg gagcttctag aactggaatc
      541 cggaggagag ggcagaggaa gtctgctaac atgcggtgac gtcgaggaga atcctggccc
      601 aatgactgaa tacaagccta ctgtcaggtt ggctacaaga gacgacgttc ctagagccgt
      661 gagaactctg gctgcagcct tcgccgacta ccccgccacg agacacaccg ttgacccaga
      721 tcggcatatt gagagagtga ctgaactgca ggagctgttt cttacaagag ttggcctcga
      781 cataggcaag gtgtgggtgg cggacgacgg cgccgccgtg gccgtctgga ccactcccga
      841 atcagttgag gctggcgccg tattcgctga gatcggcccg agaatggctg agctcagcgg
      901 gagtaggctc gcggcacagc agcaaatgga gggactgctg gcaccacaca ggcccaaaga
      961 acccgcctgg ttcctggcaa ccgtcggtgt atctcccgat catcagggga aaggtctggg
     1021 ctctgccgta gtgctccctg gcgtggaggc agctgagaga gcaggagtac ctgccttctt
     1081 ggagacctcc gctccaagga atcttccctt ctatgaacgg ttgggcttca ccgtgacagc
     1141 cgacgtggaa gtccccgaag gcccccgcac ttggtgcatg acgaggaagc ctggagcgtg
     1201 agggcccgct tcgagcagac atgataagat acattgatga gtttggacaa accacaacta
     1261 gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa
     1321 ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg
     1381 ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa
     1441 tcgataaaga tccgtcgacc gatgcccttg agagccttca acccagtcag ctccttccgg
     1501 tgggcgcggg gcatgactat cgtcgccgca cttatgactg tcttctttat catgcaactc
     1561 gtaggacagg tgccggcagc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     1621 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     1681 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     1741 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     1801 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     1861 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     1921 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     1981 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     2041 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     2101 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     2161 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     2221 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     2281 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     2341 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     2401 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     2461 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     2521 cagcggccgc aaatgctaaa ccactgcagt ggttaccagt gcttgatcag tgaggcaccg
     2581 atctcagcga tctgcctatt tcgttcgtcc atagtggcct gactccccgt cgtgtagatc
     2641 actacgattc gtgagggctt accatcaggc cccagcgcag caatgatgcc gcgagagccg
     2701 cgttcaccgg cccccgattt gtcagcaatg aaccagccag cagggagggc cgagcgaaga
     2761 agtggtcctg ctactttgtc cgcctccatc cagtctatga gctgctgtcg tgatgctaga
     2821 gtaagaagtt cgccagtgag tagtttccga agagttgtgg ccattgctac tggcatcgtg
     2881 gtatcacgct cgtcgttcgg tatggcttcg ttcaactctg gttcccagcg gtcaagccgg
     2941 gtcacatgat cacccatatt atgaagaaat gcagtcagct ccttagggcc tccgatcgtt
     3001 gtcagaagta agttggccgc ggtgttgtcg ctcatggtaa tggcagcact acacaattct
     3061 cttaccgtca tgccatccgt aagatgcttt tccgtgaccg gcgagtactc aaccaagtcg
     3121 ttttgtgagt agtgtatacg gcgaccaagc tgctcttgcc cggcgtctat acgggacaac
     3181 accgcgccac atagcagtac tttgaaagtg ctcatcatcg ggaatcgttc ttcggggcgg
     3241 aaagactcaa ggatcttgcc gctattgaga tccagttcga tatagcccac tcttgcaccc
     3301 agttgatctt cagcatcttt tactttcacc agcgtttcgg ggtgtgcaaa aacaggcaag
     3361 caaaatgccg caaagaaggg aatgagtgcg acacgaaaat gttggatgct catactcgtc
     3421 ctttttcaat attattgaag catttatcag ggttactagt acgtctctca aggataagta
     3481 agtaatatta aggtacggga ggtattggac aggccgc
//
