LOCUS       sequence-383918-        7102 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Wild type His-SUMO SPOP expression plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7102)
  AUTHORS   Thomasen FE, Cuneo MJ, Mittag T, Lindorff-Larsen K
  TITLE     Conformational and oligomeric states of SPOP from small-angle X-ray 
            scattering and molecular dynamics simulations.
  JOURNAL   Elife. 2023 Mar 1;12:e84147. doi: 10.7554/eLife.84147.
  PUBMED    36856266
REFERENCE   2  (bases 1 to 7102)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife."; 
            date: "2023-03-1"; volume: "12:e84147. doi"; pages: " 
            10.7554/eLife.84147"
COMMENT     Sequence Label: His SUMO TEV WT SPOP 28-359
FEATURES             Location/Qualifiers
     source          1..7102
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1050..1069
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1089..1166)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1372..1391
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1475..1494
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1475..1493
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1494..1518
                     /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)."
     RBS             1533..1555
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1575..1592
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1602..1619
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1626..1919
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     protein_bind    1926..1950
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1953..1973
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     protein_bind    complement(2977..3001)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3078..3096)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3092..3139
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(3314..3319,3320..3336,3337..3342))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(3369..3387)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3455..3559
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3560..3628,3629..4420)
                     /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(3778..3797)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4591..5179
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5080..5099
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5333..5350
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5365..5504
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5430..5449)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5590..5612
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5606..5797)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ttgcgctcac tgcccgcttt ccagtcggga aactgtcgtg ccagctgcat taatgaatcg
       61 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc
      121 agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag
      181 cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg
      241 atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg
      301 cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc
      361 agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac
      421 atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat
      481 ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc
      541 gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca
      601 tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga
      661 acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg
      721 atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg
      781 ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta
      841 atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc
      901 agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc
      961 gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc
     1021 acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact
     1081 ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga
     1141 aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat gcgactcctg
     1201 cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg
     1261 tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac
     1321 gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc
     1381 ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc
     1441 ggcgtagagg atcgagatct cgatcccgcg aaattaatac gactcactat aggggaattg
     1501 tgagcggata acaattcccc tctagaaata attttgttta actttaagaa ggagatatac
     1561 atatgggcag cagccatcat catcatcatc acggcagcgg cctggtgccg cgcggcagcg
     1621 ctagcatgtc ggactcagaa gtcaatcaag aagctaagcc agaggtcaag ccagaagtca
     1681 agcctgagac tcacatcaat ttaaaggtgt ccgatggatc ttcagagatc ttcttcaaga
     1741 tcaaaaagac cactccttta agaaggctga tggaagcgtt cgctaaaaga cagggtaagg
     1801 aaatggactc cttaagattc ttgtacgacg gtattagaat tcaagctgat cagacccctg
     1861 aagatttgga catggaggat aacgatatta ttgaggctca cagagaacag attggtggta
     1921 gtatcacaag tttgtacaaa aaagcaggct tcgaaaacct gtattttcag ggcagcaagg
     1981 tagtgaaatt ctcctacatg tggaccatca ataactttag cttttgccgg gaggaaatgg
     2041 gtgaagtcat taaaagttct acattttcat caggagcaaa tgataaactg aaatggtgtt
     2101 tgcgagtaaa ccccaaaggg ttagatgaag aaagcaaaga ttacctgtca ctttacctgt
     2161 tactggtcag ctgtccaaag agtgaagttc gggcaaaatt caaattctcc atcctgaatg
     2221 ccaagggaga agaaaccaaa gctatggaga gtcaacgggc atataggttt gtgcaaggca
     2281 aagactgggg attcaagaaa ttcatccgta gagattttct tttggatgag gccaacgggc
     2341 ttctccccga tgacaagctt accctcttct gcgaggtgag tgttgtgcaa gattctgtca
     2401 acatttctgg ccagaatacc atgaacatgg taaaggttcc tgagtgccgg ctggcagatg
     2461 agttaggagg actgtgggag aattcccggt tcacagactg ctgcttgtgt gttgccggcc
     2521 aggaattcca ggctcacaag gctatcttag cagctcgttc tccggttttt agtgccatgt
     2581 ttgaacatga aatggaggag agcaaaaaga atcgagttga aatcaatgat gtggagcctg
     2641 aagtttttaa ggaaatgatg tgcttcattt acacggggaa ggctccaaac ctcgacaaaa
     2701 tggctgatga tttgctggca gctgctgaca agtatgccct ggagcgctta aaggtcatgt
     2761 gtgaggatgc cctctgcagt aacctgtccg tggagaacgc tgcagaaatt ctcatcctgg
     2821 ccgacctcca cagtgcagat cagttgaaaa ctcaggcagt ggatttcatc aactatcatg
     2881 cttcggatgt cttggagacc tctgggtgga agtcaatggt ggtgtcacat ccccacttgg
     2941 tggctgaggc ataccgctct ctggcttcag cataaaaccc agctttcttg tacaaagtgg
     3001 tgatactgga tccatgggta ccctcgagga tccggctgct aacaaagccc gaaaggaagc
     3061 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg
     3121 ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggatatcccg caagaggccc
     3181 ggcagtaccg gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg
     3241 acgatgagcg cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt
     3301 gataaactac cgcattaaag cttatcgatg ataagctgtc aaacatgaga attcttgaag
     3361 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     3421 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     3481 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     3541 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     3601 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     3661 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     3721 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     3781 atgtggcgcg gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca
     3841 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     3901 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     3961 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     4021 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     4081 cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg
     4141 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     4201 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     4261 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     4321 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     4381 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     4441 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4501 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     4561 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     4621 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     4681 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     4741 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     4801 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     4861 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     4921 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     4981 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     5041 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     5101 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     5161 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5221 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5281 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     5341 agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg
     5401 tatttcacac cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     5461 cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa
     5521 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     5581 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     5641 ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt
     5701 catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc
     5761 gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat
     5821 ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta
     5881 ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga
     5941 tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg
     6001 taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc
     6061 agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg
     6121 ttgttgctca ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg
     6181 gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca
     6241 ggagcacgat catgcgcacc cgtggccagg acccaacgct gcccgagatg cgccgcgtgc
     6301 ggctgctgga gatggcggac gcgatggata tgttctgcca agggttggtt tgcgcattca
     6361 cagttctccg caagaattga ttggctccaa ttcttggagt ggtgaatccg ttagcgaggt
     6421 gccgccggct tccattcagg tcgaggtggc ccggctccat gcaccgcgac gcaacgcggg
     6481 gaggcagaca aggtataggg cggcgcctac aatccatgcc aacccgttcc atgtgctcgc
     6541 cgaggcggca taaatcgccg tgacgatcag cggtccaatg atcgaagtta ggctggtaag
     6601 agccgcgagc gatccttgaa gctgtccctg atggtcgtca tctacctgcc tggacagcat
     6661 ggcctgcaac gcgggcatcc cgatgccgcc ggaagcgaga agaatcataa tggggaaggc
     6721 catccagcct cgcgtcgcga acgccagcaa gacgtagccc agcgcgtcgg ccgccatgcc
     6781 ggcgataatg gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg
     6841 agcgagggcg tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca
     6901 gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg
     6961 catgataaag aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa
     7021 ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt
     7081 gagctaactt acattaattg cg
//
