LOCUS       sequence-374531-        7767 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid ensures constitutive expression of the N-terminal fragment 
            of split PPARgamma (aa 200-429), fused to the VPR activation domain,
            a flexible GS linker and the SV40 NLS signal at the C-terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7767)
  AUTHORS   Rihtar E, Lebar T, Lainscek D, Kores K, Lesnik S, Bren U, Jerala R
  TITLE     Chemically inducible split protein regulators for mammalian cells.
  JOURNAL   Nat Chem Biol. 2022 Sep 26. doi: 10.1038/s41589-022-01136-x.
  PUBMED    36163385
REFERENCE   2  (bases 1 to 7767)
  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 Chem 
            Biol. 2022 Sep 26. doi: 10.1038/s41589-022-01136-x."
COMMENT     Sequence Label: nPPARg429-gs10-NLS-VPR
FEATURES             Location/Qualifiers
     source          1..7767
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(54..73)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        245..624
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        625..828
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     779..799
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     873..892
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        873..891
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      954..963
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1686..1706
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1731..1880
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             2349..2705
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALL"
     CDS             2724..3293
                     /codon_start=1
                     /product="transcriptional activation domain from the human 
                     herpesvirus 4 (Epstein-Barr virus) replication and 
                     transcription activator Rta/BRLF1 (Hardwick et al., 1992; 
                     Chavez et al., 2015)"
                     /label=Rta AD
                     /translation="RDSREGMFLPKPEAGSAISDVFEGREVCQPKRIRPFHPPGSPWAN
                     RPLPASLAPTPTGPVHEPVGSLTPAPVPQPLDPAPAVTPEASHLLEDPDEETSQAVKAL
                     REMADTVIPQKEEAAICGQMDLSHPPPRGHLDELTTTLESMTEDLNLDSPLTPELNEIL
                     DTFLNDECLLHAMHISTGLSIFDTSLF"
     promoter        complement(3330..3348)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3331..3348)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3368..3385)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3374..3598
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3644..4072
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3731..3750)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3941..3962
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4081..4101)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4086..4415
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4266..4401
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4328..4347
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4482..5276
                     /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(4536..4555)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5146..5165
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5450..5571
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5487..5506)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5541..5560
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5620..5636)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5620..5636)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5633..5655)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5644..5660
                     /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(5668..5674,5675..5692,5693..5698))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5713..5734
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5851..5868)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6022..6610)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6102..6121)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6781..7572,7573..7641))
                     /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     7404..7423
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7642..7746)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc
       61 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct
      121 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg
      181 aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg
      241 cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
      301 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      361 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      421 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      481 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      541 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      601 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      661 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      721 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      781 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
      841 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
      901 gcttagtacc gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcgccacca
      961 tgggcagcca catgttgaat cctgaaagcg ctgatttgcg cgcacttgct aaacatctct
     1021 acgattctta tatcaaatca ttccccttga ccaaagcaaa agcgagagct atacttaccg
     1081 gtaagacaac ggataagtcc ccattcgtta tttatgacat gaattcactt atgatgggtg
     1141 aagataagat aaaattcaaa catataactc ctctccaaga gcaatcaaag gaagttgcga
     1201 ttagaatctt ccagggctgt caattcagaa gcgtagaagc ggtccaggag attacggagt
     1261 atgcaaaatc tatccccggc tttgtcaacc tcgaccttaa cgaccaagtc actcttctta
     1321 aatacggggt acacgaaatt atatacacca tgcttgcgtc cctgatgaac aaagacggtg
     1381 tgctgattag tgaaggtcag gggtttatga cgcgggaatt ccttaagagt cttagaaaac
     1441 ctttcgggga ctttatggag ccaaagttcg agtttgcagt gaaatttaat gcactggaac
     1501 tcgacgactc tgacctggct atattcatag ccgttattat tctcagcgga gaccggcctg
     1561 gactcctcaa cgtgaagccg attgaggaca tacaggataa cctgctccaa gcccttgagc
     1621 tgcaactgaa gctcaatcac cccgaatcta gcggcggctc tggcggcggc tccggaggct
     1681 ctgatccaaa aaagaagaga aaggtagagg ccagcggttc gggacgggct gacgcattgg
     1741 acgattttga tctggatatg ctgggaagtg acgccctcga tgattttgac cttgacatgc
     1801 ttggttcgga tgcccttgat gactttgacc tcgacatgct cggcagtgac gcccttgatg
     1861 atttcgacct ggacatgctg attaacacta gtggcggctc tggcggcggc tccggaggct
     1921 ctagccagta cctgcccgac accgacgaca ggcaccggat cgaggaaaag cggaagcgga
     1981 cctacgagac attcaagagc atcatgaaga agtccccctt cagcggcccc accgacccta
     2041 gacctccacc tagaagaatc gccgtgccca gcagatccag cgccagcgtg ccaaaacctg
     2101 ccccccagcc ttaccccttc accagcagcc tgagcaccat caactacgac gagttcccta
     2161 ccatggtgtt ccccagcggc cagatctctc aggcctctgc tctggctcca gcccctcctc
     2221 aggtgctgcc tcaggctcct gctcctgcac cagctccagc catggtgtct gcactggctc
     2281 aggcaccagc acccgtgcct gtgctggctc ctggacctcc acaggctgtg gctccaccag
     2341 cccctaaacc tacacaggcc ggcgagggca cactgtctga agctctgctg cagctgcagt
     2401 tcgacgacga ggatctggga gccctgctgg gaaacagcac cgatcctgcc gtgttcaccg
     2461 acctggccag cgtggacaac agcgagttcc agcagctgct gaaccagggc atccctgtgg
     2521 cccctcacac caccgagccc atgctgatgg aataccccga ggccatcacc cggctcgtga
     2581 caggcgctca gaggcctcct gatccagctc ctgcccctct gggagcacca ggcctgccta
     2641 atggactgct gtctggcgac gaggacttca gctctatcgc cgatatggat ttctcagcct
     2701 tgctgggctc tggcagcggc agccgggatt ccagggaagg gatgtttttg ccgaagcctg
     2761 aggccggctc cgctattagt gacgtgtttg agggccgcga ggtgtgccag ccaaaacgaa
     2821 tccggccatt tcatcctcca ggaagtccat gggccaaccg cccactcccc gccagcctcg
     2881 caccaacacc aaccggtcca gtacatgagc cagtcgggtc actgaccccg gcaccagtcc
     2941 ctcagccact ggacccagcg cccgcagtga ctcccgaggc cagtcacctg ttggaggacc
     3001 ccgatgaaga gacgagccag gctgtcaaag cccttcggga gatggccgat actgtgattc
     3061 cccagaagga agaggctgca atctgtggcc aaatggacct ttcccatccg cccccaaggg
     3121 gccatctgga tgagctgaca accacacttg agtccatgac cgaggatctg aacctggact
     3181 cacccctgac cccggaattg aacgagattc tggatacctt cctgaacgac gagtgcctct
     3241 tgcatgccat gcatatcagc acaggactgt ccatcttcga cacatctctg tttgacacat
     3301 acaggtatat ctaatctaga gggccctatt ctatagtgtc acctaaatgc tagagctcgc
     3361 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     3421 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     3481 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     3541 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     3601 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     3661 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3721 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3781 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3841 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3901 gtttttcgcc ttttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3961 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     4021 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     4081 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     4141 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     4201 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     4261 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     4321 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     4381 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     4441 ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     4501 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     4561 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     4621 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     4681 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     4741 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     4801 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     4861 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     4921 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     4981 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     5041 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     5101 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     5161 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     5221 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     5281 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     5341 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     5401 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     5461 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5521 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     5581 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     5641 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     5701 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     5761 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5821 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5881 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5941 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     6001 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     6061 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6121 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6181 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     6241 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6301 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6361 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6421 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     6481 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6541 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6601 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6661 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6721 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6781 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6841 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6901 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6961 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     7021 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     7081 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7141 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7201 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7261 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7321 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7381 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7441 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7501 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7561 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7621 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7681 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7741 tccgcgcaca tttccccgaa aagtgcc
//
