LOCUS       sequence-304876-        8266 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Reporter integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8266)
  AUTHORS   Yoshimatsu S, Sone T, Nakajima M, Sato T, Okochi R, Ishikawa M, 
            Nakamura M, Sasaki E, Shiozawa S, Okano H
  TITLE     A versatile toolbox for knock-in gene targeting based on the 
            Multisite Gateway technology.
  JOURNAL   PLoS One. 2019 Aug 27;14(8):e0221164. doi: 
            10.1371/journal.pone.0221164. eCollection 2019.
  PUBMED    31454364
REFERENCE   2  (bases 1 to 8266)
  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: "PLoS One.";
            date: "2019-08-27"; volume: "14(8):e0221164. doi"; pages: " 
            10.1371/journal.pone.0221164. eCollection 2019"
COMMENT     Sequence Label: pUC-DEST-cjBLIMP1-Venus-HygTK
FEATURES             Location/Qualifiers
     source          1..8266
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..23
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     15..32
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     16..32
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    73..93
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     regulatory      96..105
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             719..775
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(776..778,779..1489)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(818..839)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1079..1098)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     repeat_region   1499..1561
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    1732..1756
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        1763..2262
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1798..1821)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2195..2214
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3366..4361
                     /codon_start=1
                     /product="truncated herpes simplex virus thymidine kinase 
                     (Salomon et al., 1995)"
                     /label=Delta-TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWRV
                     LGASETIANIYTTQHRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGS
                     SHAPPPALTLIFDRHPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGAL
                     PEDRHIDRLAKRQRPGERLDLAMLAAIRRVYGLLANTVRYLQCGGSWREDWGQLSGTAV
                     PPQGAEPQSNAGPRPHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRSMHVFI
                     LDYDQSPAGCRDALLQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     protein_bind    complement(4727..4751)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     repeat_region   5025..5059
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    complement(5601..5621)
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     complement(5682..5698)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5682..5698)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5695..5717)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5706..5722
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5730..5736,5737..5754,5755..5760))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5775..5796
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5913..5930)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6084..6672)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6164..6183)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6843..7634,7635..7703))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7466..7485
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7704..7808)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7876..7894
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7932..7954)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8054..8073
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cccagtcacg acgttgtaaa acgacggcca gtgccaagct tgcatgcctg caggtcggga
       61 tatctgaatt atcaactttg tataataaag ttgttgccac catgggcgtc tcccttccct
      121 gctgtctctc tgctctgcac tgtaggtctg ccacaagcga tttagtagca ccagcaatct
      181 caagacccac ctgcgactcc attctggaga gaaaccgtac cagtgcaagg tgtgccctgc
      241 caagttcacc cagttcgtgc acctgaaact gcacaagcgc ctgcacaccc gggagcggcc
      301 ccacaagtgc gcccagtgcc acaagaacta catccatctc tgtagcctca aggttcacct
      361 gaaggggaac tgccctgtag ccccagcccc tgggctgccc ttggaagacc tgacccgaat
      421 caatgaagaa attgagaagt ttgacatcag cgacaatgcc gaccggcttg aggacgtgga
      481 ggatgacatc agtgtggtct ctgtcgtgga gaaggaaatt ctggccatgg tcagaaaaga
      541 gaaagaagaa actggcctga aagtgtcttt gcaaagaaac atggggaatg gactcctctc
      601 ctcagggtgt ggcctttacg agtcatcaga tctgcccctc atgaagttgc ctcccagcaa
      661 cccgctacct ctggtacctg taaaggtcaa acaagaaaca gtcgaaccaa tggatcctgc
      721 cacgaacttc tctctgttaa agcaagcagg agacgtggaa gaaaaccccg gtcctgttag
      781 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
      841 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
      901 gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
      961 caccctgggc tacggcctgc agtgcttcgc ccgctacccc gaccacatga agcagcacga
     1021 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     1081 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     1141 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     1201 gtacaactac aacagccaca acgtctatat caccgccgac aagcagaaga acggcatcaa
     1261 ggccaacttc aagatccgcc acaacatcga ggacggcggc gtgcagctcg ccgaccacta
     1321 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     1381 ctaccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     1441 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg aattttaacc
     1501 ctagaaagat aatcatattg tgacgtacgt taaagataat catgcgtaaa attgacgcat
     1561 gtgttttatc ggtctgtata tcgaggtccc gtgaggcgtt tgaatagata ttaagtttta
     1621 ttatatttac acttacatac taataataaa ttcaacaaac aattctttta cgtgactttt
     1681 aagatttaac tcaaacaaaa actcaaaatt tcttctataa agtaacaaaa ctcaagtttg
     1741 tacaaaaaag cagaattcta ccgggtaggg gaggcgcttt tcccaaggca gtctggagca
     1801 tgcgctttag cagccccgct gggcacttgg cgctacacaa gtggcctctg gcctcgcaca
     1861 cattccacat ccaccggtag gcgccaaccg gctccgttct ttggtggccc cttcgcgcca
     1921 ccttctactc ctcccctagt caggaagttc ccccccgccc cgcagctcgc gtcgtgcagg
     1981 acgtgacaaa tggaagtagc acgtctcact agtctcgtgc agatggacag caccgctgag
     2041 caatggaagc gggtaggcct ttggggcagc ggccaatagc agctttgctc cttcgctttc
     2101 tgggctcaga ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc
     2161 ggggcgggcg cccgaaggtc ctccggaggc ccggcattct gcacgcttca aaagcgcacg
     2221 tctgccgcgc tgttctcctc ttcctcatct ccgggccttt cgaccgatca tcaagcttga
     2281 tcctcatgaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt
     2341 tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct
     2401 tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca
     2461 aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg
     2521 acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca
     2581 cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca
     2641 tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc
     2701 aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg
     2761 tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg
     2821 atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt
     2881 tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg
     2941 aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt
     3001 tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat
     3061 cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg
     3121 ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat
     3181 ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg
     3241 atggctgtgt agaagtcgcg tctgcgttcg accaggctgc gcgttctcgc ggccatagca
     3301 accgacgtac ggcgttgcgc cctcgccggc agcaagaagc cacggaagtc cgcccggagc
     3361 agaaaatgcc cacgctactg cgggtttata tagacggtcc ccacgggatg gggaaaacca
     3421 ccaccacgca actgctggtg gccctgggtt cgcgcgacga tatcgtctac gtacccgagc
     3481 cgatgactta ctggcgggtg ctgggggctt ccgagacaat cgcgaacatc tacaccacac
     3541 aacaccgcct cgaccagggt gagatatcgg ccggggacgc ggcggtggta atgacaagcg
     3601 cccagataac aatgggcatg ccttatgccg tgaccgacgc cgttctggct cctcatatcg
     3661 ggggggaggc tgggagctca catgccccgc ccccggccct caccctcatc ttcgaccgcc
     3721 atcccatcgc cgccctcctg tgctacccgg ccgcgcggta ccttatgggc agcatgaccc
     3781 cccaggccgt gctggcgttc gtggccctca tcccgccgac cttgcccggc accaacatcg
     3841 tgcttggggc ccttccggag gacagacaca tcgaccgcct ggccaaacgc cagcgccccg
     3901 gcgagcggct ggacctggct atgctggctg cgattcgccg cgtttacggg ctacttgcca
     3961 atacggtgcg gtatctgcag tgcggcgggt cgtggcggga ggactgggga cagctttcgg
     4021 ggacggccgt gccgccccag ggtgccgagc cccagagcaa cgcgggccca cgaccccata
     4081 tcggggacac gttatttacc ctgtttcggg cccccgagtt gctggccccc aacggcgacc
     4141 tgtataacgt gtttgcctgg gccttggacg tcttggccaa acgcctccgt tccatgcacg
     4201 tctttatcct ggattacgac caatcgcccg ccggctgccg ggacgccctg ctgcaactta
     4261 cctccgggat ggtccagacc cacgtcacca cccccggctc cataccgacg atatgcgacc
     4321 tggcgcgcac gtttgcccgg gagatggggg aggctaactg agctctagct tatcgactcg
     4381 ccgatagtgg aaaccgacgc cccagcactc gtccgagggc aaaggaatag tcgagaaatt
     4441 gatgatctat taaacaataa agatgtccac atggaagttt ttcctgtcat actttgttaa
     4501 gaagggtgag aacagagtac ctacattttg aatggaagga ttggagctac gggggtgggg
     4561 gtggggtggg attagataaa tgcctgctct ttactgaagg ctctttacta ttgctttatg
     4621 ataatgtttc atagttggat atcataattt aaacaagcaa aaccaaatta agggccagct
     4681 cattcctccc actcatgatc tatagatcct ctagagtcga gggctgcagc agctttcttg
     4741 tacaaagtgg gatatctata acaagaaaat atatatataa taagttatca cgtaagtaga
     4801 acatgaaata acaatataat tatcgtatga gttaaatctt aaaagtcacg taaaagataa
     4861 tcatgcgtca ttttgactca cgcggtcgtt atagttcaaa atcagtgaca cttaccgcat
     4921 tgacaagcac gcctcacggg agctccaagc ggcgactgag atgtcctaaa tgcacagcga
     4981 cggattcgcg ctatttagaa agagagagca atatttcaag aatgcatgcg tcaattttac
     5041 gcagactatc tttctagggt taataagatt ttccgaaaac acttgttttg tttcttaagt
     5101 tatgacttgg tgagtcaggg tgcctgtagg aagttgcttg tacataattc cagctctgca
     5161 aagctctctt gatagcaaat ggtttcccct cacctctgga attaaagaag gaactccaaa
     5221 gttactgaaa tctcagggca tgaacaaggc aaaggccata tatatatata cttatttact
     5281 cccatgtcta tatatttgcc cctgtgtatt ttgaatattt gtgtggatat gtttgcatag
     5341 ccttcccatt actgagacta ttgcctagtc ataattattt tttcaatgat aatccttcat
     5401 aatttattat acaatttatc attcagaaag caataattaa aaaagtttac aataactgga
     5461 aagattcctt gtaatttgag tatcaatgta tttttgtctt gtggccattc tttgtagata
     5521 atttctgcac atctctataa gtacctaaga tttagttaaa caaatctatg acttcagtca
     5581 acctctctct ctctaataaa caacttttct atacaaagtt gatagcttga tatcaatact
     5641 ctagaggatc cccgggtacc gagctcgaat tcgtaatcat ggtcatagct gtttcctgtg
     5701 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5761 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5821 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5881 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5941 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     6001 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     6061 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     6121 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     6181 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6241 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6301 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6361 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6421 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6481 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6541 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6601 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6661 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6721 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6781 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6841 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6901 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6961 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     7021 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     7081 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     7141 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7201 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     7261 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7321 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7381 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7441 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7501 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7561 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7621 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7681 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7741 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7801 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     7861 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     7921 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     7981 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     8041 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     8101 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg
     8161 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     8221 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggtttt
//
