LOCUS       sequence-390451-        6995 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian inducible expression vector for GFP-tagged SPOP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6995)
  AUTHORS   Sabri N, Cuneo MJ, Marzahn MR, Lee J, Bouchard JJ, Gullulu O, 
            Vaithiyalingam S, Borgia MB, Schmit J, Mittag T
  TITLE     Reduction of oligomer size modulates the competition between cluster
            formation and phase separation of the tumor suppressor SPOP.
  JOURNAL   J Biol Chem. 2023 Dec;299(12):105427. doi: 
            10.1016/j.jbc.2023.105427. Epub 2023 Nov 4.
  PUBMED    37926283
REFERENCE   2  (bases 1 to 6995)
  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: "J Biol 
            Chem."; date: "2023-12"; volume: "299(12):105427. doi"; pages: " 
            10.1016/j.jbc.2023.105427. Epub 2023 Nov 4"
COMMENT     Sequence Label: pCDNA4/TO-GFP-SPOP (R354H)
FEATURES             Location/Qualifiers
     source          1..6995
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(1037..1039,1040..1042,1043..1753)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1082..1103)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1343..1362)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1690..1711
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2918..2947
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2963..2980
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3003..3020)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3009..3233
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3279..3707
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3366..3385)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3576..3597
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3716..3736)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3721..4050
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3901..4036
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3963..3982
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4098..4145
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4164..4538
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4668..4789
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4705..4724)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4759..4778
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4838..4854)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4838..4854)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4851..4873)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4862..4878
                     /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(4886..4892,4893..4910,4911..4916))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4931..4952
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5069..5086)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5240..5828)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5320..5339)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5999..6790,6791..6859))
                     /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     6622..6641
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6860..6964)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cactagtcca gtgtggtgga
     1021 attctgcaga tatccaatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     1081 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     1141 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     1201 cgtgccctgg cccaccctcg tgaccacctt cacctacggc gtgcagtgct tcgcccgcta
     1261 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     1321 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     1381 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     1441 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaaggtct atatcaccgc
     1501 cgacaagcag aagaacggca tcaaggtgaa cttcaagacc cgccacaaca tcgaggacgg
     1561 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     1621 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
     1681 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
     1741 cgagctgtac aagagatatc caatgtcaag ggttccaagt cctccacctc cggcagaaat
     1801 gtcgagtggt cccgttgccg agagctggtg ctatacacag atcaaggtag tgaagttctc
     1861 ctacatgtgg accatcaata actttagttt ttgccgggag gaaatgggtg aagtcataaa
     1921 aagttcaact ttttcatcag gagccaatga taaactgaaa tggtgtttgc gagtaaaccc
     1981 gaaagggcta gatgaagaaa gcaaagatta cctgtcgctt tacctgttgt tggtcagctg
     2041 tccaaagagt gaagttcggg caaagttcaa gttctccatc ctgaatgcca agggagaaga
     2101 aaccaaagct atggagagtc agcgagctta taggttcgtg caaggcaaag actggggatt
     2161 caaaaaattc atccgcagag attttctctt ggacgaggcc aatgggcttc tccctgatga
     2221 caagctcacc ctcttctgtg aggtgagtgt agtacaagac tctgtcaata tttctgggca
     2281 gaatactatg aacatggtaa aggttcctga atgccggttg gcagatgagt taggagggct
     2341 gtgggagaac tctcgattca ccgactgctg cttgtgtgtg gctggtcaag aattccaggc
     2401 tcacaaggct atcttagcag ctcggtctcc agtcttcagt gccatgtttg aacatgaaat
     2461 ggaggagagc aaaaagaatc gggttgagat taacgacgtg gagcctgagg tgtttaagga
     2521 gatgatgtgc ttcatctaca cggggaaggc gccgaacctc gacaagatgg ctgacgactt
     2581 gctggcagct gccgataagt acgccctgga gcgtctgaag gtcatgtgtg aggacgccct
     2641 ctgcagtaac ctctctgtgg agaacgctgc agagattctc atcctggctg acctccacag
     2701 cgcagatcag ctgaagacgc aggcagttga tttcatcaac tatcatgctt cggatgtctt
     2761 ggagacctct gggtggaagt caatggtggt gtcacatccc cacttagtgg cagaagcata
     2821 ccattctctg gcttcagcac agtgcccttt cctggggccc ccacgcaaac gcctgaagca
     2881 atcctagtgg cggccgctcg agtctagagg gcccttcgaa caaaaactca tctcagaaga
     2941 ggatctgaat atgcataccg gtcatcatca ccatcaccat tgagtttaaa cccgctgatc
     3001 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3061 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3121 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     3181 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     3241 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3301 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3361 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3421 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3481 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3541 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3601 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3661 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     3721 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3781 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3841 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3901 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3961 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4021 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4081 ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat
     4141 acgacaaggt gaggaactaa accatggcca agttgaccag tgccgttccg gtgctcaccg
     4201 cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg gctcgggttc tcccgggact
     4261 tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt gaccctgttc atcagcgcgg
     4321 tccaggacca ggtggtgccg gacaacaccc tggcctgggt gtgggtgcgc ggcctggacg
     4381 agctgtacgc cgagtggtcg gaggtcgtgt ccacgaactt ccgggacgcc tccgggccgg
     4441 ccatgaccga gatcggcgag cagccgtggg ggcgggagtt cgccctgcgc gacccggccg
     4501 gcaactgcgt gcacttcgtg gccgaggagc aggactgaca cgtgctacga gatttcgatt
     4561 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     4621 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     4681 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4741 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     4801 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     4861 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     4921 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     4981 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5041 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5101 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5161 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5221 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5281 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5341 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5401 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5461 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5521 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5581 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5641 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5701 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5761 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5821 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5881 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5941 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6001 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6061 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6121 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6181 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6241 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6301 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6361 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6421 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6481 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6541 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6601 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6661 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6721 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     6781 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     6841 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6901 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     6961 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
