LOCUS       sequence-389608-        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 (WT)
FEATURES             Location/Qualifiers
     source          1..6995
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(59..78)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     269..290
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(409..429)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        414..743
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      594..729
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     656..675
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        791..838
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             857..1231
                     /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    1361..1482
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1398..1417)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1452..1471
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1531..1547)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1531..1547)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1544..1566)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1555..1571
                     /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(1579..1585,1586..1603,1604..1609))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1624..1645
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1762..1779)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1933..2521)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2013..2032)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2692..3483,3484..3552))
                     /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     3315..3334
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3553..3657)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3732..3751)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3923..4302
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4303..4506
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4457..4477
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    4508..4526
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4529..4547
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     4557..4581
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(4725..4727,4728..4730,4731..5441)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4770..4791)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5031..5050)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5378..5399
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             6606..6635
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             6651..6668
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6691..6708)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6697..6921
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(6967..6995,1..400)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
       61 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
      121 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
      181 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
      241 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
      301 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
      361 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
      421 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
      481 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
      541 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
      601 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
      661 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
      721 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
      781 atcagcacgt gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa
      841 ggtgaggaac taaaccatgg ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga
      901 cgtcgccgga gcggtcgagt tctggaccga ccggctcggg ttctcccggg acttcgtgga
      961 ggacgacttc gccggtgtgg tccgggacga cgtgaccctg ttcatcagcg cggtccagga
     1021 ccaggtggtg ccggacaaca ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta
     1081 cgccgagtgg tcggaggtcg tgtccacgaa cttccgggac gcctccgggc cggccatgac
     1141 cgagatcggc gagcagccgt gggggcggga gttcgccctg cgcgacccgg ccggcaactg
     1201 cgtgcacttc gtggccgagg agcaggactg acacgtgcta cgagatttcg attccaccgc
     1261 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     1321 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     1381 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     1441 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     1501 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     1561 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     1621 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     1681 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     1741 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     1801 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     1861 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     1921 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     1981 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     2041 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     2101 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     2161 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     2221 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     2281 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     2341 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     2401 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     2461 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     2521 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     2581 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     2641 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     2701 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     2761 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     2821 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     2881 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     2941 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     3001 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     3061 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     3121 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     3181 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     3241 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     3301 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     3361 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     3421 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     3481 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     3541 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     3601 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     3661 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
     3721 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
     3781 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
     3841 ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
     3901 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     3961 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     4021 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     4081 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     4141 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     4201 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     4261 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     4321 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     4381 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     4441 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     4501 agagctctcc ctatcagtga tagagatctc cctatcagtg atagagatcg tcgacgagct
     4561 cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga
     4621 agacaccggg accgatccag cctccggact ctagcgttta aacttaagct tggtaccgag
     4681 ctcggatcca ctagtccagt gtggtggaat tctgcagata tccaatggtg agcaagggcg
     4741 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     4801 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     4861 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttca
     4921 cctacggcgt gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
     4981 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     5041 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     5101 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     5161 acaacagcca caaggtctat atcaccgccg acaagcagaa gaacggcatc aaggtgaact
     5221 tcaagacccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     5281 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     5341 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     5401 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gagatatcca atgtcaaggg
     5461 ttccaagtcc tccacctccg gcagaaatgt cgagtggtcc cgttgccgag agctggtgct
     5521 atacacagat caaggtagtg aagttctcct acatgtggac catcaataac tttagttttt
     5581 gccgggagga aatgggtgaa gtcataaaaa gttcaacttt ttcatcagga gccaatgata
     5641 aactgaaatg gtgtttgcga gtaaacccga aagggctaga tgaagaaagc aaagattacc
     5701 tgtcgcttta cctgttgttg gtcagctgtc caaagagtga agttcgggca aagttcaagt
     5761 tctccatcct gaatgccaag ggagaagaaa ccaaagctat ggagagtcag cgagcttata
     5821 ggttcgtgca aggcaaagac tggggattca aaaaattcat ccgcagagat tttctcttgg
     5881 acgaggccaa tgggcttctc cctgatgaca agctcaccct cttctgtgag gtgagtgtag
     5941 tacaagactc tgtcaatatt tctgggcaga atactatgaa catggtaaag gttcctgaat
     6001 gccggttggc agatgagtta ggagggctgt gggagaactc tcgattcacc gactgctgct
     6061 tgtgtgtggc tggtcaagaa ttccaggctc acaaggctat cttagcagct cggtctccag
     6121 tcttcagtgc catgtttgaa catgaaatgg aggagagcaa aaagaatcgg gttgagatta
     6181 acgacgtgga gcctgaggtg tttaaggaga tgatgtgctt catctacacg gggaaggcgc
     6241 cgaacctcga caagatggct gacgacttgc tggcagctgc cgataagtac gccctggagc
     6301 gtctgaaggt catgtgtgag gacgccctct gcagtaacct ctctgtggag aacgctgcag
     6361 agattctcat cctggctgac ctccacagcg cagatcagct gaagacgcag gcagttgatt
     6421 tcatcaacta tcatgcttcg gatgtcttgg agacctctgg gtggaagtca atggtggtgt
     6481 cacatcccca cttagtggca gaagcatacc gttctctggc ttcagcacag tgccctttcc
     6541 tggggccccc acgcaaacgc ctgaagcaat cctagtggcg gccgctcgag tctagagggc
     6601 ccttcgaaca aaaactcatc tcagaagagg atctgaatat gcataccggt catcatcacc
     6661 atcaccattg agtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat
     6721 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     6781 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     6841 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     6901 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     6961 atccccacgc gccctgtagc ggcgcattaa gcgcg
//
