LOCUS       sequence-473622-        7415 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  transient overexpression of KEAP1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7415)
  AUTHORS   Marzio A, Kurz E, Sahni JM, Di Feo G, Puccini J, Jiang S, Hirsch CA,
            Arbini AA, Wu WL, Pass HI, Bar-Sagi D, Papagiannakopoulos T, Pagano 
            M
  TITLE     EMSY inhibits homologous recombination repair and the interferon 
            response, promoting lung cancer immune evasion.
  JOURNAL   Cell. 2022 Jan 6;185(1):169-183.e19. doi: 
            10.1016/j.cell.2021.12.005. Epub 2021 Dec 27.
  PUBMED    34963055
REFERENCE   2  (bases 1 to 7415)
  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"; doi: 
            "10.1016/j.cell.2021.12.005"; journalName: "Cell"; date: "2022-01-6-
            6"; volume: "185"; issue: "1"; pages: "169-183.e19"
COMMENT     Sequence Label: pcDNA3.1 2xFLAG-2xSTREP-KEAP1 (human)
FEATURES             Location/Qualifiers
     source          1..7415
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1250..1267)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1256..1480
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1526..1954
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1613..1632)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1823..1844
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1963..1983)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1968..2297
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2148..2283
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2210..2229
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2364..3158
                     /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(2418..2437)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3028..3047
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3332..3453
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3369..3388)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3423..3442
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3502..3518)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3502..3518)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3515..3537)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3526..3542
                     /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(3550..3556,3557..3574,3575..3580))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3595..3616
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3733..3750)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3904..4492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3984..4003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4663..5454,5455..5523))
                     /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     5286..5305
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5524..5628)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5703..5722)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5894..6273
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6274..6477
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6428..6448
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     6522..6541
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6522..6540
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6582..6605
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             6621..6644
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             6660..6683
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             6720..6743
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
ORIGIN
        1 aacctgtccc actgccaact ggtgaccctc atcagccggg acgacctgaa cgtgcgctgc
       61 gagtccgagg tcttccacgc ctgcatcaac tgggtcaagt acgactgcga acagcgacgg
      121 ttctacgtcc aggcgctgct gcgggccgtg cgctgccact cgttgacgcc gaacttcctg
      181 cagatgcagc tgcagaagtg cgagatcctg cagtccgact cccgctgcaa ggactacctg
      241 gtcaagatct tcgaggagct caccctgcac aagcccacgc aggtgatgcc ctgccgggcg
      301 cccaaggtgg gccgcctgat ctacaccgcg ggcggctact tccgacagtc gctcagctac
      361 ctggaggctt acaaccccag tgacggcacc tggctccggt tggcggacct gcaggtgccg
      421 cggagcggcc tggccggctg cgtggtgggc gggctgttgt acgccgtggg cggcaggaac
      481 aactcgcccg acggcaacac cgactccagc gccctggact gttacaaccc catgaccaat
      541 cagtggtcgc cctgcgcccc catgagcgtg ccccgtaacc gcatcggggt gggggtcatc
      601 gatggccaca tctatgccgt cggcggctcc cacggctgca tccaccacaa cagtgtggag
      661 aggtatgagc cagagcggga tgagtggcac ttggtggccc caatgctgac acgaaggatc
      721 ggggtgggcg tggctgtcct caatcgtctc ctttatgccg tggggggctt tgacgggaca
      781 aaccgcctta attcagctga gtgttactac ccagagagga acgagtggcg aatgatcaca
      841 gcaatgaaca ccatccgaag cggggcaggc gtctgcgtcc tgcacaactg tatctatgct
      901 gctgggggct atgatggtca ggaccagctg aacagcgtgg agcgctacga tgtggaaaca
      961 gagacgtgga ctttcgtagc ccccatgaag caccggcgaa gtgccctggg gatcactgtc
     1021 caccagggga gaatctacgt ccttggaggc tatgatggtc acacgttcct ggacagtgtg
     1081 gagtgttacg acccagatac agacacctgg agcgaggtga cccgaatgac atcgggccgg
     1141 agtggggtgg gcgtggctgt caccatggag ccctgccgga agcagattga ccagcagaac
     1201 tgtacctgtt gactcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg
     1261 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     1321 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     1381 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     1441 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     1501 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     1561 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     1621 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     1681 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     1741 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     1801 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     1861 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     1921 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     1981 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2041 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2101 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     2161 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     2221 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     2281 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag
     2341 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     2401 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     2461 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     2521 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     2581 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     2641 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     2701 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     2761 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     2821 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     2881 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     2941 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     3001 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     3061 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     3121 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     3181 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     3241 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     3301 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     3361 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     3421 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     3481 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     3541 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     3601 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     3661 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     3721 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     3781 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     3841 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     3901 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     3961 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4021 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4081 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4141 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4201 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4261 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4321 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4381 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4441 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4501 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4561 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4621 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4681 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     4741 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     4801 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     4861 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     4921 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     4981 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5041 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5101 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5161 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5221 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5281 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5341 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5401 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     5461 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     5521 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     5581 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     5641 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc
     5701 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     5761 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     5821 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
     5881 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
     5941 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     6001 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     6061 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     6121 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     6181 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     6241 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
     6301 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     6361 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     6421 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc
     6481 tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac tcactatagg
     6541 gagacccaag ctggctagcg tttaaactta agcttaccat ggactacaag gacgacgatg
     6601 acaaggtacc aggatctatg gattataaag atgatgatga taaagggtcg gccgccagct
     6661 ggagccaccc tcagttcgag aagggaggag gaagcggcgg aggcagcgga ggaggaagct
     6721 ggagccaccc gcagttcgag aaaggagctg gatcccagcc agatcccagg cctagcgggg
     6781 ctggggcctg ctgccgattc ctgcccctgc agtcacagtg ccctgagggg gcaggggacg
     6841 cggtgatgta cgcctccact gagtgcaagg cggaggtgac gccctcccag catggcaacc
     6901 gcaccttcag ctacaccctg gaggatcata ccaagcaggc ctttggcatc atgaacgagc
     6961 tgcggctcag ccagcagctg tgtgacgtca cactgcaggt caagtaccag gatgcaccgg
     7021 ccgcccagtt catggcccac aaggtggtgc tggcctcatc cagccctgtc ttcaaggcca
     7081 tgttcaccaa cgggctgcgg gagcagggca tggaggtggt gtccattgag ggtatccacc
     7141 ccaaggtcat ggagcgcctc attgaattcg cctacacggc ctccatctcc atgggcgaga
     7201 agtgtgtcct ccacgtcatg aacggtgctg tcatgtacca gatcgacagc gttgtccgtg
     7261 cctgcagtga cttcctggtg cagcagctgg accccagcaa tgccatcggc atcgccaact
     7321 tcgctgagca gattggctgt gtggagttgc accagcgtgc ccgggagtac atctacatgc
     7381 attttgggga ggtggccaag caagaggagt tcttc
//
