LOCUS       sequence-460017-        6274 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pGex for protein overexpression of human BRD4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6274)
  AUTHORS   Hsia O, Hinterndorfer M, Cowan AD, Iso K, Ishida T, Sundaramoorthy 
            R, Nakasone MA, Imrichova H, Schatz C, Rukavina A, Husnjak K, Wegner
            M, Correa-Saez A, Craigon C, Casement R, Maniaci C, Testa A, Kaulich
            M, Dikic I, Winter GE, Ciulli A
  TITLE     Targeted protein degradation via intramolecular bivalent glues.
  JOURNAL   Nature. 2024 Mar;627(8002):204-211. doi: 10.1038/s41586-024-07089-6.
            Epub 2024 Feb 21.
  PUBMED    38383787
REFERENCE   2  (bases 1 to 6274)
  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: 
            "10.1038/s41586-024-07089-6"; journalName: "Nature"; date: 
            "2024-03"; volume: "627"; issue: "8002"; pages: "204-211"
COMMENT     Sequence Label: His-GST-TEV-BRD4(42-459) BD1-BD2
FEATURES             Location/Qualifiers
     source          1..6274
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        833..910
                     /gene="lacI"
                     /label=lacI promoter
     CDS             911..1993
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(930..949)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(2042..2047,2048..2065,2066..2072)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2080..2096
                     /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)."
     primer_bind     2104..2120
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2104..2120
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2116..2643
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(2136..2153)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2136..2152)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2145..2167)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(2745..2750,2751..2766,2767..2773)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2781..2797
                     /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)."
     CDS             2826..2855
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2829..2858
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2832..2861
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2868..3521
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     3479..3501
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             3528..3548
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(4886..4908)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5053..5071)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5139..5243
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5244..5312,5313..6104)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5462..5481)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt ccgacaccat
      841 cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
      901 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
      961 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
     1021 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
     1081 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
     1141 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
     1201 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
     1261 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
     1321 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
     1381 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
     1441 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
     1501 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
     1561 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
     1621 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
     1681 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
     1741 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     1801 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     1861 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     1921 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     1981 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcaccccagg
     2041 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     2101 acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt acaacgtcgt
     2161 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     2221 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     2281 aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga aagctggctg
     2341 gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca gatgcacggt
     2401 tacgatgcgc ccatctacac caacgtgacc tatcccatta cggtcaatcc gccgtttgtt
     2461 cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga aagctggcta
     2521 caggaaggcc agacgcgaat tatttttgat ggcgttggaa ttagcttatc gactgcacgg
     2581 tgcaccaatg cttctggcgt caggcagcca tcggaagctg tggtatggct gtgcaggtcg
     2641 taaatcactg cataattcgt gtcgctcaag gcgcactccc gttctggata atgttttttg
     2701 cgccgacatc ataacggttc tggcaaatat tctgaaatga gctgttgaca attaatcatc
     2761 ggctcgtata atgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagtattcc
     2821 atatgcatca ccatcaccat caccatcacc accatcatca cgcaggcatg tcccctatac
     2881 taggttattg gaaaattaag ggccttgtgc aacccactcg acttcttttg gaatatcttg
     2941 aagaaaaata tgaagagcat ttgtatgagc gcgatgaagg tgataaatgg cgaaacaaaa
     3001 agtttgaatt gggtttggag tttcccaatc ttccttatta tattgatggt gatgttaaat
     3061 taacacagtc tatggccatc atacgttata tagctgacaa gcacaacatg ttgggtggtt
     3121 gtccaaaaga gcgtgcagag atttcaatgc ttgaaggagc ggttttggat attagatacg
     3181 gtgtttcgag aattgcatat agtaaagact ttgaaactct caaagttgat tttcttagca
     3241 agctacctga aatgctgaaa atgttcgaag atcgtttatg tcataaaaca tatttaaatg
     3301 gtgatcatgt aacccatcct gacttcatgt tgtatgacgc tcttgatgtt gttttataca
     3361 tggacccaat gtgcctggat gcgttcccaa aattagtttg ttttaaaaaa cgtattgaag
     3421 ctatcccaca aattgataag tacttgaaat ccagcaagta tatagcatgg cctttgcagg
     3481 gctggcaagc cacgtttggt ggtggcgacc atcctccaaa atcggatgaa aacctgtatt
     3541 ttcagggatc caccaacccc ccgcccccag agacctccaa ccctaacaag cccaagaggc
     3601 agaccaacca actgcaatac ctgctcagag tggtgctcaa gacactatgg aaacaccagt
     3661 ttgcatggcc tttccagcag cctgtggatg ccgtcaagct gaacctccct gattactata
     3721 agatcattaa aacgcctatg gatatgggaa caataaagaa gcgcttggaa aacaactatt
     3781 actggaatgc tcaggaatgt atccaggact tcaacactat gtttacaaat tgttacatct
     3841 acaacaagcc tggagatgac atagtcttaa tggcagaagc tctggaaaag ctcttcttgc
     3901 aaaaaataaa tgagctaccc acagaagaaa ccgagatcat gatagtccag gcaaaaggaa
     3961 gaggacgtgg gaggaaagaa acagggacag caaaacctgg cgtttccacg gtaccaaaca
     4021 caactcaagc atcgactcct ccgcagaccc agacccctca gccgaatcct cctcctgtgc
     4081 aggccacgcc tcaccccttc cctgccgtca ccccggacct catcgtccag acccctgtca
     4141 tgacagtggt gcctccccag ccactgcaga cgcccccgcc agtgcccccc cagccacaac
     4201 ccccacccgc tccagctccc cagcccgtac agagccaccc acccatcatc gcggccaccc
     4261 cacagcctgt gaagacaaag aagggagtga agaggaaagc agacaccacc acccccacca
     4321 ccattgaccc cattcacgag ccaccctcgc tgcccccgga gcccaagacc accaagctgg
     4381 gccagcggcg ggagagcagc cggcctgtga aacctccaaa gaaggacgtg cccgactctc
     4441 agcagcaccc agcaccagag aagagcagca aggtctcgga gcagctcaag tgctgcagcg
     4501 gcatcctcaa ggagatgttt gccaagaagc acgccgccta cgcctggccc ttctacaagc
     4561 ctgtggacgt ggaggcactg ggcctacacg actactgtga catcatcaag caccccatgg
     4621 acatgagcac aatcaagtct aaactggagg cccgtgagta ccgtgatgct caggagtttg
     4681 gtgctgacgt ccgattgatg ttctccaact gctataagta caaccctcct gaccatgagg
     4741 tggtggccat ggcccgcaag ctccaggatg tgttcgaaat gcgctttgcc aagatgccgg
     4801 actaagaatt cccgggtcga ctcgagcggc cgcatcgtga ctgactgacg atctgcctcg
     4861 cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag acggtcacag
     4921 cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg
     4981 gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga tagcggagtg tataattctt
     5041 gaagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     5101 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     5161 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     5221 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     5281 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     5341 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     5401 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     5461 tgctatgtgg cgcggtatta tcccgtgttg acgccgggca agagcaactc ggtcgccgca
     5521 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     5581 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5641 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5701 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5761 acgacgagcg tgacaccacg atgcctgcag caatggcaac aacgttgcgc aaactattaa
     5821 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5881 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     5941 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     6001 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     6061 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     6121 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     6181 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     6241 cgtcagaccc cgtagaaaag atcaaaggat cttc
//
