LOCUS       sequence-334854-        6578 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of mouse SPOP-BTB (L186D, L190D, L193D, I217K).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6578)
  AUTHORS   Bouchard JJ, Otero JH, Scott DC, Szulc E, Martin EW, Sabri N, 
            Granata D, Marzahn MR, Lindorff-Larsen K, Salvatella X, Schulman BA,
            Mittag T
  TITLE     Cancer Mutations of the Tumor Suppressor SPOP Disrupt the Formation 
            of Active, Phase-Separated Compartments.
  JOURNAL   Mol Cell. 2018 Oct 4;72(1):19-36.e8. doi: 
            10.1016/j.molcel.2018.08.027. Epub 2018 Sep 20.
  PUBMED    30244836
REFERENCE   2  (bases 1 to 6578)
  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.molcel.2018.08.027"; journalName: "Mol Cell"; date: 
            "2018-10-4- 4"; volume: "72"; issue: "1"; pages: "19-36.e8"
COMMENT     Sequence Label: SPOP-V5 (BTB)
FEATURES             Location/Qualifiers
     source          1..6578
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(25..42)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(196..784)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(276..295)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(955..1746,1747..1815))
                     /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     1578..1597
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1816..1920)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1995..2014)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2186..2565
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2566..2769
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2720..2740
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2814..2833
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2814..2832
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2922..2963
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(4120..4137)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4126..4350
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4396..4824
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4483..4502)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4693..4714
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4833..4853)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4838..5167
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5018..5153
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5080..5099
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5234..6028
                     /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(5288..5307)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5898..5917
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6202..6323
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6239..6258)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6293..6312
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6372..6388)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6372..6388)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6385..6407)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6396..6412
                     /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(6420..6426,6427..6444,6445..6450))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6465..6486
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
       61 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
      121 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
      181 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
      241 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
      301 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
      361 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
      421 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
      481 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
      541 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
      601 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
      661 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
      721 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
      781 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
      841 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
      901 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
      961 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     1021 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     1081 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     1141 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     1201 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     1261 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     1321 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     1381 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     1441 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     1501 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     1561 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     1621 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     1681 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     1741 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     1801 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     1861 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     1921 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
     1981 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     2041 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     2101 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
     2161 gatgtacggg ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca
     2221 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
     2281 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     2341 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     2401 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     2461 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     2521 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
     2581 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     2641 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     2701 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     2761 agcagagctc tctggctaac tagagaaccc actgcttact ggcttatcga aattaatacg
     2821 actcactata gggagaccca agctggctag cgtttaaact taagcttggt accgagctcg
     2881 gatccactag tccagtgtgg tggaattctg cagatatcat gggtaagcct atccctaacc
     2941 ctctcctcgg tctcgattct acgatgtcaa gggttccaag tcctccacct ccggcagaaa
     3001 tgtcgagtgg tcccgttgcc gagagctggt gctatacaca gatcaaggta gtgaagttct
     3061 cctacatgtg gaccatcaat aactttagtt tttgccggga ggaaatgggt gaagtcataa
     3121 aaagttcaac tttttcatca ggagccaatg ataaactgaa atggtgtttg cgagtaaacc
     3181 cgaaagggct agatgaagaa agcaaagatt acctgtcgct ttacctgttg ttggtcagct
     3241 gtccaaagag tgaagttcgg gcaaagttca agttctccat cctgaatgcc aagggagaag
     3301 aaaccaaagc tatggagagt cagcgagctt ataggttcgt gcaaggcaaa gactggggat
     3361 tcaaaaaatt catccgcaga gattttctct tggacgaggc caatgggctt ctccctgatg
     3421 acaagctcac cctcttctgt gaggtgagtg tagtacaaga ctctgtcaat atttctgggc
     3481 agaatactat gaacatggta aaggttcctg aatgccggga cgcagatgag gacggagggg
     3541 attgggagaa ctctcgattc accgactgct gcttgtgtgt ggctggtcaa gaattccagg
     3601 ctcacaaggc taagttagca gctcgttctc cagtcttcag tgccatgttt gaacatgaaa
     3661 tggaggagag caaaaagaat cgggttgaga ttaacgacgt ggagcctgag gtgtttaagg
     3721 agatgatgtg cttcatctac acggggaagg cgccgaacct cgacaagatg gctgacgact
     3781 tgctggcagc tgccgataag tacgccctgg agcgtctgaa ggtcatgtgt gaggacgccc
     3841 tctgcagtaa cctctctgtg gagaacgctg cagagattct catcctggct gacctccaca
     3901 gcgcagatca gctgaagacg caggcagttg atttcatcaa ctatcatgct tcggatgtct
     3961 tggagacctc tgggtggaag tcaatggtgg tgtcacatcc ccacttagtg gcagaagcat
     4021 accgttctct ggcttcagca cagtgccctt tcctggggcc cccacgcaaa cgcctgaagc
     4081 aatcctaatc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     4141 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     4201 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     4261 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     4321 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     4381 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     4441 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     4501 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     4561 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     4621 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     4681 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     4741 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4801 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     4861 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4921 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4981 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     5041 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     5101 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     5161 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     5221 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     5281 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     5341 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     5401 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     5461 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     5521 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     5581 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     5641 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     5701 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     5761 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     5821 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     5881 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     5941 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     6001 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     6061 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     6121 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     6181 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     6241 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     6301 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     6361 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     6421 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     6481 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     6541 cattaatgaa tcggccaacg cgcggggaga ggcggttt
//
