LOCUS       sequence-475139-        8812 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express NanoLuc-YAP1(biased)-TEAD4-HaloTag Fusion Protein in 
            Mammalian Cells under a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8812)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8812)
  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: 
            "Unpublished"
COMMENT     Sequence Label: NanoLuc-YAP1(biased)-TEAD4-HaloTag Bidirection...
FEATURES             Location/Qualifiers
     source          1..8812
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    3..84
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(1820..2332)
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     promoter        complement(2361..2379)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2362..2379)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2499..2802
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2803..3006
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2957..2977
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          3142..3274
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3257..3276
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     3318..3337
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3318..3336
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4673..4693
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             4709..5593
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     polyA_signal    5680..5801
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5691..5710)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(5916..5949)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5959..6006
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6038..6832
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6092..6111)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6702..6721
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      7049..7594
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     7495..7514
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7711..7728
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    7878..7926
                     /note="synthetic polyadenylation signal"
     CDS             complement(7950..8348)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      8345..8354
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(8379..8736)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      8393..8528
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        8545..8736
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     8693..8713
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 cggcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
       61 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
      121 cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtatggct
      181 gattatgatc cgcggccgca atacgtcgac gttatcagct gacttcgtac gagagcctag
      241 gtgtttctag aggatccccg ggtaccgagc ccgaattcgt ttaaactaac catgtaagaa
      301 agctttcttt atctagcttg gtggcagcca aaacagactc catgtcatta aggatgtcag
      361 aactcaaagc ttcctgcaga cttggcatca gctcctctcc ttctatgttc attccatctc
      421 cttccagtgt tccaaggtcc acatttgtcc caggaatggc ttcaaggtag tctgggaaac
      481 ggttctgctg tgagggcagg gtgctttggt tgatagtatc acctgtatcc atctcatcca
      541 cactgttcag gaagtcatct ggggttcgag ggacactgta gctgctcatg cttagtccac
      601 tgtctgtact ctcatctcga gagtgatagg tgccactgtt aaggaaagga tctgagctat
      661 tggtcgtcat tgttctcaat tcctgagaca tcccgggaga agacactgga ttttgagtcc
      721 caccatcctg ctccagtgtt ggtaactggc tacgcagggc taactcctga cattttggag
      781 aatttgctgt gctgggattg atattccgca ttgcctgccg aagcagttct tgctgtttca
      841 gccgcagcct ctccttctcc atctgcagtt gctgcagtcg catctgttgc tgctggttgg
      901 agttgctgcc acccatgacg cctccctgtg ggctctgggg agccaggggt ggtggctgtt
      961 tcactggagc actctgactg attctctggt tcatggcaaa acgagggtca agccttgggt
     1021 ctagccaaga ggtggtcttg ttcttatggt ttatatagta aatttctcca tcctgagtca
     1081 tggcttgttc ccatccatca ggaagaggac ctgaagccga gttcatcata ttctgctgca
     1141 ctggtggact ggtgggggct gtgacgttca tctgggacag catggccttc ctggggtcct
     1201 gccatgttgt tgtctgatcg atgtgattta agaagtatct ctgaccagaa gatgtctttg
     1261 ccatctccca acctgctggc agaggtacat catcaggtat ctcaaaagaa gactgtcgaa
     1321 gatgctgagc tgtgggtgta gctgctgggc cagagactac tccagtgggg gtcagtgtcc
     1381 caggagaaac agctcccaac tgcagagaag ctggagagga atgagctcga acatgctgtg
     1441 gagtcagggc tcctgcagtg cctgcatcag tactggcctg tcgggagtgg gatttgggct
     1501 ccggcggctt gaagaaggag tcgggcagct tccggagcct catgggcacg gtctggggca
     1561 cgttggccgt cttggggttc atgacggcgt tgaagagcgc ctccaggtcg gtctccgagt
     1621 ccccgcggac gtgcacgatc tgatgcccgg cggggggtgc ctgcggcgcc gcctgggtcg
     1681 ccgcgggtgc cggttgcccg ggtccggacg gcgggccctg cccctgcggg ggctgcgaag
     1741 gcggctgccc ttggccctgg ggggccggtt gaggcggcgg ctgctgcccg ggatccatgg
     1801 cgatcgcgcc gctcgagccc gccagaatgc gttcgcacag ccgccagccg gtcactccgt
     1861 tgatggttac tcggaacagc agggagccgt cggggttgat caggcgctcg tcgataattt
     1921 tgttgccgtt ccacagggtc cctgttacag tgatcttttt gccgtcgaac acggcgatgc
     1981 cttcatacgg ccgtccgaaa tagtcgatca tgttcggcgt aaccccgtcg attaccagtg
     2041 tgccatagtg caggatcacc ttaaagtgat gatcatccac agggtacacc accttaaaaa
     2101 ttttttcgat ctggcccatt tggtcgccgc tcagaccttc atacgggatg atgacatgga
     2161 tgtcgatctt cagcccattt tcaccgctca ggacaatcct ttggatcgga gttacggaca
     2221 ccccgagatt ctgaaacaaa ctggacacac ctccctgttc aaggacttgg tccaggttgt
     2281 agccggctgt ctgtcgccag tccccaacga aatcttcgag tgtgaagacc atggttgcaa
     2341 aaagaacaag tagcttgtat tctatagtgt cacctaaatg ggcgatctga cggttcacta
     2401 aaccagctct gctgatatag gcctccctgg cgcgccacta gttattaata gtaatcaatt
     2461 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     2521 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     2581 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     2641 actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc tattgacgtc
     2701 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg ggactttcct
     2761 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     2821 tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     2881 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaat
     2941 aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     3001 agagctggtt tagtgaaccg tcagatcact agaagcttta ttgcggtagt ttatcacagt
     3061 taaattgcta acgcagtcag tgcttctgac acaacagtct cgaacttaag ctgcagaagt
     3121 tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta aggagaccaa
     3181 tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc acctattggt
     3241 cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccagtt caattacagc
     3301 tcttaaggct agagtattaa tacgactcac tatagggcta gcgatcgcca tggagggcac
     3361 ggccggcacc attacctcca acgagtggag ctctcccacc tcccctgagg ggagcaccgc
     3421 ctctgggggc agtcaggcac tggacaagcc catcgacaat gacgcagagg gcgtgtggag
     3481 cccggatatt gagcagagtt tccaggaggc cctcgccatc tacccgccct gtggcaggcg
     3541 caaaatcatc ctgtcggacg agggcaagat gtatggtcgg aacgagctga ttgcccgcta
     3601 catcaagctc cggacaggga agacccgcac caggaagcag gtctccagcc acatccaggt
     3661 gctggctcgt cgcaaagctc gcgagatcca ggccaagcta aaggaccagg cagctaagga
     3721 caaggccctg cagagcatgg ctgccatgtc gtctgcacag atcatctccg ccacggcctt
     3781 ccacagtagc atggccctcg cccggggccc cggccgccca gcagtctcag ggttttggca
     3841 aggagctttg ccaggccaag ccggaacgtc ccatgatgtg aagcctttct ctcagcaaac
     3901 ctatgctgtc cagcctccgc tgcctctgcc agggtttgag tctcctgcag ggcccgcccc
     3961 atcgccctct gcgcccccgg cacccccatg gcagggccgc agcgtggcca gctccaagct
     4021 ctggatgttg gagttctctg ccttcctgga gcagcagcag gacccggaca cgtacaacaa
     4081 gcacctgttc gtgcacattg gccagtccag cccaagctac agcgacccct acctcgaagc
     4141 cgtggacatc cgccaaatct atgacaaatt cccggagaaa aagggtggac tcaaggatct
     4201 cttcgaacgg ggaccctcca atgccttttt tcttgtgaag ttctgggcag acctcaacac
     4261 caacatcgag gatgaaggca gctccttcta tggggtctcc agccagtatg agagccccga
     4321 gaacatgatc atcacctgct ccacgaaggt ctgctctttc ggcaagcagg tggtggagaa
     4381 agttgagaca gagtatgctc gctatgagaa tggacactac tcttaccgca tccaccggtc
     4441 cccgctctgt gagtacatga tcaacttcat ccacaagctc aagcacctcc ctgagaagta
     4501 catgatgaac agcgtgctgg agaacttcac catcctgcag gtggtcacca acagagacac
     4561 acaggagacc ttgctgtgca ttgcctatgt ctttgaggtg tcagccagtg agcacggggc
     4621 tcagcaccac atctacaggc tggtgaaaga agtttctctc gagccaacca ctgaggatct
     4681 gtactttcag agcgataacg atggatccga aatcggtact ggctttccat tcgaccccca
     4741 ttatgtggaa gtcctgggcg agcgcatgca ctacgtcgat gttggtccgc gcgatggcac
     4801 ccctgtgctg ttcctgcacg gtaacccgac ctcctcctac gtgtggcgca acatcatccc
     4861 gcatgttgca ccgacccatc gctgcattgc tccagacctg atcggtatgg gcaaatccga
     4921 caaaccagac ctgggttatt tcttcgacga ccacgtccgc ttcatggatg ccttcatcga
     4981 agccctgggt ctggaagagg tcgtcctggt cattcacgac tggggctccg ctctgggttt
     5041 ccactgggcc aagcgcaatc cagagcgcgt caaaggtatt gcatttatgg agttcatccg
     5101 ccctatcccg acctgggacg aatggccaga atttgcccgc gagaccttcc aggccttccg
     5161 caccaccgac gtcggccgca agctgatcat cgatcagaac gtttttatcg agggtacgct
     5221 gccgatgggt gtcgtccgcc cgctgactga agtcgagatg gaccattacc gcgagccgtt
     5281 cctgaatcct gttgaccgcg agccactgtg gcgcttccca aacgagctgc caatcgccgg
     5341 tgagccagcg aacatcgtcg cgctggtcga agaatacatg gactggctgc accagtcccc
     5401 tgtcccgaag ctgctgttct ggggcacccc aggcgttctg atcccaccgg ccgaagccgc
     5461 tcgcctggcc aaaagcctgc ctaactgcaa ggctgtggac atcggcccgg gtctgaatct
     5521 gctgcaagaa gacaacccgg acctgatcgg cagcgagatc gcgcgctggc tgtctactct
     5581 ggagatttcc ggttaataga attctagagt cgacctgcag gcatgcaagc tgatttggct
     5641 gctgccaccg ctgagcaagg ccgcttcgag cagacatgat aagatacatt gatgagtttg
     5701 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     5761 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     5821 attttatgtt tcaggttcag ggggagatgt gggaggtttt tttaagcaag taaaacctct
     5881 acaaatgtgg taaaatctta attaagaatt ctaatataac ttcgtatagc atacattata
     5941 cgaagttatg gatccactgt tgacaattaa tcatcggcat agtatatcgg catagtataa
     6001 tacgacaagg tgaggaacta aacccaggag gcagatcatg attgaacaag atggattgca
     6061 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     6121 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     6181 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
     6241 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     6301 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     6361 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     6421 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     6481 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     6541 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
     6601 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     6661 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     6721 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     6781 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gattcgaaat
     6841 gaccgaccaa gcgacgccca accggtatca gctcactcaa aggcggtaat acggttatcc
     6901 acagaatcag gggataacgc aggaaagaac atgtcaaaca tgagaattac aacttatatc
     6961 gtatggggct gacttcaggt gctacatttg aagagataaa ttgcactgaa atctagaaat
     7021 attttatctg attaataaga tgatcttctt gagatcgttt tggtctgcgc gtaatctctt
     7081 gctctgaaaa cgaaaaaacc gccttgcagg gcggtttttc gaaggttctc tgagctacca
     7141 actctttgaa ccgaggtaac tggcttggag gagcgcagtc accaaaactt gtcctttcag
     7201 tttagcctta accggcgcat gacttcaaga ctaactcctc taaatcaatt accagtggct
     7261 gctgccagtg gtgcttttgc atgtctttcc gggttggact caagacgata gttaccggat
     7321 aaggcgcagc ggtcggactg aacggggggt tcgtgcatac agtccagctt ggagcgaact
     7381 gcctacccgg aactgagtgt caggcgtgga atgagacaaa cgcggccata acagcggaat
     7441 gacaccggta aaccgaaagg caggaacagg agagcgcacg agggagccgc cagggggaaa
     7501 cgcctggtat ctttatagtc ctgtcgggtt tcgccaccac tgatttgagc gtcagatttc
     7561 gtgatgcttg tcaggggggc ggagcctatg gaaaaacggc tttgccgcgg ccctctcact
     7621 tccctgttaa gtatcttcct ggcatcttcc aggaaatctc cgccccgttc gtaagccatt
     7681 tccgctcgcc gcagtcgaac gaccgagcgt agcgagtcag tgagcgagga agcggaatat
     7741 atcctgtatc acatattctg ctgacgcacc ggtgcagcct tttttctcct gccacatgaa
     7801 gcacttcact gacaccctca tcagtgccaa catagtaagc cagtatacac tccgctagcg
     7861 ctgatgtccg gaatcgtcac acaaaaaacc aacacacaga tgtaatgaaa ataaagatat
     7921 tttattgcgg ccatcgtgat ggctagagtt tagccctccc acacataacc agagggcagc
     7981 aattcacgaa tcccaactgc cgtcggctgt ccatcactgt ccttcactat ggctttgatc
     8041 ccaggatgca gatcgagaag cacctgtcgg caccgtccgc aggggctcaa gatgcccctg
     8101 ttctcatttc cgatcgcgac gatacaagtc aggttgccag ctgccgcagc agcagcagtg
     8161 cccagcacca cgagttctgc acaaggtccc ccagtaaaat gatatacatt gacaccagtg
     8221 aagatgcggc cgtcgctaga gagagctgcg ctggcgacgc tgtagtcttc agagatgggg
     8281 atgctgttga ttgtagccgt tgctctttca atgagggtgg attcttcttg agacaaaggc
     8341 ttggccatgg tggcgctagt gtcagaagaa tcgagctttt tgcaaaagcc taggcctcca
     8401 aaaaagcctc ctcactactt ctggaatagc tcagaggccg aggcggcctc ggcctctgca
     8461 taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg cggagttagg
     8521 ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag atgcatgctt
     8581 tgcatacttc tgcctgctgg ggagcctggg gactttccac acctggttgc tgactaattg
     8641 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acaccctaac
     8701 tgacacacat tccacagctg gttctttccg cctcagaagg tacctaacca agttcctctt
     8761 tcagaggtta tttcaggcca tggtgctgcg caagacgcgt taattaagac gt
//
