LOCUS       sequence-471869-        8822 bp    DNA     circular SYN 16-JUN-2025
DEFINITION  Express NLuc-YAP1_TEAD4-HaloTag(R) Fusion Proteins in Mammalian 
            Cells under a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8822)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 8822)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 16, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: Nluc-YAP1_TEAD4-HaloTag Bidirectional(Balanced...
FEATURES             Location/Qualifiers
     source          1..8822
                     /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(2371..2389)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2372..2389)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2509..2812
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2813..3016
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2967..2987
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          3152..3284
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3267..3286
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     3328..3347
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3328..3346
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4683..4703
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             4719..5603
                     /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    5690..5811
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5701..5720)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(5926..5959)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5969..6016
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6048..6842
                     /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(6102..6121)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6712..6731
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      7059..7604
                     /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     7505..7524
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7721..7738
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    7888..7936
                     /note="synthetic polyadenylation signal"
     CDS             complement(7960..8358)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      8355..8364
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(8389..8746)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      8403..8538
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        8555..8746
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     8703..8723
                     /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 atggtgagcg
     2341 ctagctgcaa aaagaacaag tagcttgtat tctatagtgt cacctaaatg ggcgatctga
     2401 cggttcacta aaccagctct gcttatatag gcctccctgg cgcgccacta gttattaata
     2461 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     2521 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     2581 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
     2641 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc
     2701 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg
     2761 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
     2821 gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
     2881 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
     2941 atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt
     3001 ctatataagc agagctggtt tagtgaaccg tcagatcact agaagcttta ttgcggtagt
     3061 ttatcacagt taaattgcta acgcagtcag tgcttctgac acaacagtct cgaacttaag
     3121 ctgcagaagt tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta
     3181 aggagaccaa tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc
     3241 acctattggt cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccagtt
     3301 caattacagc tcttaaggct agagtattaa tacgactcac tatagggcta gcgatcgcca
     3361 tggagggcac ggccggcacc attacctcca acgagtggag ctctcccacc tcccctgagg
     3421 ggagcaccgc ctctgggggc agtcaggcac tggacaagcc catcgacaat gacgcagagg
     3481 gcgtgtggag cccggatatt gagcagagtt tccaggaggc cctcgccatc tacccgccct
     3541 gtggcaggcg caaaatcatc ctgtcggacg agggcaagat gtatggtcgg aacgagctga
     3601 ttgcccgcta catcaagctc cggacaggga agacccgcac caggaagcag gtctccagcc
     3661 acatccaggt gctggctcgt cgcaaagctc gcgagatcca ggccaagcta aaggaccagg
     3721 cagctaagga caaggccctg cagagcatgg ctgccatgtc gtctgcacag atcatctccg
     3781 ccacggcctt ccacagtagc atggccctcg cccggggccc cggccgccca gcagtctcag
     3841 ggttttggca aggagctttg ccaggccaag ccggaacgtc ccatgatgtg aagcctttct
     3901 ctcagcaaac ctatgctgtc cagcctccgc tgcctctgcc agggtttgag tctcctgcag
     3961 ggcccgcccc atcgccctct gcgcccccgg cacccccatg gcagggccgc agcgtggcca
     4021 gctccaagct ctggatgttg gagttctctg ccttcctgga gcagcagcag gacccggaca
     4081 cgtacaacaa gcacctgttc gtgcacattg gccagtccag cccaagctac agcgacccct
     4141 acctcgaagc cgtggacatc cgccaaatct atgacaaatt cccggagaaa aagggtggac
     4201 tcaaggatct cttcgaacgg ggaccctcca atgccttttt tcttgtgaag ttctgggcag
     4261 acctcaacac caacatcgag gatgaaggca gctccttcta tggggtctcc agccagtatg
     4321 agagccccga gaacatgatc atcacctgct ccacgaaggt ctgctctttc ggcaagcagg
     4381 tggtggagaa agttgagaca gagtatgctc gctatgagaa tggacactac tcttaccgca
     4441 tccaccggtc cccgctctgt gagtacatga tcaacttcat ccacaagctc aagcacctcc
     4501 ctgagaagta catgatgaac agcgtgctgg agaacttcac catcctgcag gtggtcacca
     4561 acagagacac acaggagacc ttgctgtgca ttgcctatgt ctttgaggtg tcagccagtg
     4621 agcacggggc tcagcaccac atctacaggc tggtgaaaga agtttctctc gagccaacca
     4681 ctgaggatct gtactttcag agcgataacg atggatccga aatcggtact ggctttccat
     4741 tcgaccccca ttatgtggaa gtcctgggcg agcgcatgca ctacgtcgat gttggtccgc
     4801 gcgatggcac ccctgtgctg ttcctgcacg gtaacccgac ctcctcctac gtgtggcgca
     4861 acatcatccc gcatgttgca ccgacccatc gctgcattgc tccagacctg atcggtatgg
     4921 gcaaatccga caaaccagac ctgggttatt tcttcgacga ccacgtccgc ttcatggatg
     4981 ccttcatcga agccctgggt ctggaagagg tcgtcctggt cattcacgac tggggctccg
     5041 ctctgggttt ccactgggcc aagcgcaatc cagagcgcgt caaaggtatt gcatttatgg
     5101 agttcatccg ccctatcccg acctgggacg aatggccaga atttgcccgc gagaccttcc
     5161 aggccttccg caccaccgac gtcggccgca agctgatcat cgatcagaac gtttttatcg
     5221 agggtacgct gccgatgggt gtcgtccgcc cgctgactga agtcgagatg gaccattacc
     5281 gcgagccgtt cctgaatcct gttgaccgcg agccactgtg gcgcttccca aacgagctgc
     5341 caatcgccgg tgagccagcg aacatcgtcg cgctggtcga agaatacatg gactggctgc
     5401 accagtcccc tgtcccgaag ctgctgttct ggggcacccc aggcgttctg atcccaccgg
     5461 ccgaagccgc tcgcctggcc aaaagcctgc ctaactgcaa ggctgtggac atcggcccgg
     5521 gtctgaatct gctgcaagaa gacaacccgg acctgatcgg cagcgagatc gcgcgctggc
     5581 tgtctactct ggagatttcc ggttaataga attctagagt cgacctgcag gcatgcaagc
     5641 tgatttggct gctgccaccg ctgagcaagg ccgcttcgag cagacatgat aagatacatt
     5701 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
     5761 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
     5821 aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt tttaagcaag
     5881 taaaacctct acaaatgtgg taaaatctta attaagaatt ctaatataac ttcgtatagc
     5941 atacattata cgaagttatg gatccactgt tgacaattaa tcatcggcat agtatatcgg
     6001 catagtataa tacgacaagg tgaggaacta aacccaggag gcagatcatg attgaacaag
     6061 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     6121 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     6181 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     6241 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     6301 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     6361 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     6421 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     6481 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     6541 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     6601 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     6661 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     6721 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     6781 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     6841 gattcgaaat gaccgaccaa gcgacgccca accggtatca gctcactcaa aggcggtaat
     6901 acggttatcc acagaatcag gggataacgc aggaaagaac atgtcaaaca tgagaattac
     6961 aacttatatc gtatggggct gacttcaggt gctacatttg aagagataaa ttgcactgaa
     7021 atctagaaat attttatctg attaataaga tgatcttctt gagatcgttt tggtctgcgc
     7081 gtaatctctt gctctgaaaa cgaaaaaacc gccttgcagg gcggtttttc gaaggttctc
     7141 tgagctacca actctttgaa ccgaggtaac tggcttggag gagcgcagtc accaaaactt
     7201 gtcctttcag tttagcctta accggcgcat gacttcaaga ctaactcctc taaatcaatt
     7261 accagtggct gctgccagtg gtgcttttgc atgtctttcc gggttggact caagacgata
     7321 gttaccggat aaggcgcagc ggtcggactg aacggggggt tcgtgcatac agtccagctt
     7381 ggagcgaact gcctacccgg aactgagtgt caggcgtgga atgagacaaa cgcggccata
     7441 acagcggaat gacaccggta aaccgaaagg caggaacagg agagcgcacg agggagccgc
     7501 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccaccac tgatttgagc
     7561 gtcagatttc gtgatgcttg tcaggggggc ggagcctatg gaaaaacggc tttgccgcgg
     7621 ccctctcact tccctgttaa gtatcttcct ggcatcttcc aggaaatctc cgccccgttc
     7681 gtaagccatt tccgctcgcc gcagtcgaac gaccgagcgt agcgagtcag tgagcgagga
     7741 agcggaatat atcctgtatc acatattctg ctgacgcacc ggtgcagcct tttttctcct
     7801 gccacatgaa gcacttcact gacaccctca tcagtgccaa catagtaagc cagtatacac
     7861 tccgctagcg ctgatgtccg gaatcgtcac acaaaaaacc aacacacaga tgtaatgaaa
     7921 ataaagatat tttattgcgg ccatcgtgat ggctagagtt tagccctccc acacataacc
     7981 agagggcagc aattcacgaa tcccaactgc cgtcggctgt ccatcactgt ccttcactat
     8041 ggctttgatc ccaggatgca gatcgagaag cacctgtcgg caccgtccgc aggggctcaa
     8101 gatgcccctg ttctcatttc cgatcgcgac gatacaagtc aggttgccag ctgccgcagc
     8161 agcagcagtg cccagcacca cgagttctgc acaaggtccc ccagtaaaat gatatacatt
     8221 gacaccagtg aagatgcggc cgtcgctaga gagagctgcg ctggcgacgc tgtagtcttc
     8281 agagatgggg atgctgttga ttgtagccgt tgctctttca atgagggtgg attcttcttg
     8341 agacaaaggc ttggccatgg tggcgctagt gtcagaagaa tcgagctttt tgcaaaagcc
     8401 taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggccg aggcggcctc
     8461 ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg
     8521 cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag
     8581 atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac acctggttgc
     8641 tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc
     8701 acaccctaac tgacacacat tccacagctg gttctttccg cctcagaagg tacctaacca
     8761 agttcctctt tcagaggtta tttcaggcca tggtgctgcg caagacgcgt taattaagac
     8821 gt
//
