LOCUS       sequence-459093-        8274 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of polySUMO3 protein under T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8274)
  AUTHORS   Kim J, Qin S, Zhou HX, Rosen MK
  TITLE     Surface Charge Can Modulate Phase Separation of Multidomain 
            Proteins.
  JOURNAL   J Am Chem Soc. 2024 Feb 7;146(5):3383-3395. doi: 
            10.1021/jacs.3c12789. Epub 2024 Jan 23.
  PUBMED    38262618
REFERENCE   2  (bases 1 to 8274)
  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: 
            "10.1021/jacs.3c12789"; journalName: "J Am Chem Soc"; date: 
            "2024-02-7- 7"; volume: "146"; issue: "5"; pages: "3383-3395"
COMMENT     Sequence Label: polySUMO3
FEATURES             Location/Qualifiers
     source          1..8274
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      7..34
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        53..144
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(145..213,214..1005)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(363..382)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(1047..1560)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1242..1263)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1454..1473
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      1671..2259
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2160..2179
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2413..2430
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2536..2555)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2696..2718
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2712..2903)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        3399..3476
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3477..4559
                     /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(3496..3515)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4802..4807,4808..4823,4824..4830)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    4838..4854
                     /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     4843..4865
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             4924..6024
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      5881..5890
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     5998..6021
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             6073..6084
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             6097..6117
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             7687..7707
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             7708..7725
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7738..7755)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7738..7754)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7747..7769)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(7933..7950)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      8103..8189
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttaagcagaa ggccatcctg acggatggcc tttttgcgtt tctacaaact cttttgttta
       61 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
      121 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
      181 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
      241 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
      301 aagatccttg agagttttcg ccccgaagaa cgtttcccaa tgatgagcac ttttaaagtt
      361 ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc
      421 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
      481 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
      541 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
      601 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
      661 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
      721 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
      781 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
      841 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
      901 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
      961 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     1021 tactcatata tactttagat tgatttaccc cggttgataa tcagaaaagc cccaaaaaca
     1081 ggaagattgt ataagcaaat atttaaattg taaacgttaa tattttgtta aaattcgcgt
     1141 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     1201 ataaatcaaa agaatagccc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     1261 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     1321 gcccactacg tgaaccatca cccaaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     1381 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     1441 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     1501 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     1561 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     1621 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     1681 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     1741 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     1801 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     1861 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     1921 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     1981 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2041 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2101 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2161 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2221 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2281 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     2341 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     2401 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     2461 ctccttacgc atctgtgcgg tatttcacac cgcatgtggt gcactctcag tacaccgcat
     2521 gtggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg
     2581 ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg
     2641 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     2701 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa
     2761 agctcatcag cgtggtcgtg cagcgattca cagatgtctg cctgttcatc cgcgtccagc
     2821 tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg
     2881 gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg
     2941 gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat
     3001 gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag
     3061 agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag
     3121 ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc
     3181 cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc
     3241 gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc
     3301 taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg
     3361 cgcacccgtg gccaggaccc aacgctgccc gaaattccga caccatcgaa tggtgcaaaa
     3421 cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga
     3481 aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc
     3541 gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga
     3601 tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt
     3661 tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg
     3721 cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa
     3781 gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc
     3841 tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta
     3901 atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt attattttct
     3961 cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa
     4021 tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc
     4081 ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg
     4141 ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga
     4201 tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc
     4261 tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt
     4321 atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg
     4381 accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct
     4441 cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt
     4501 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     4561 cgcaacgcaa ttaatgtaag ttagctcact cattaggcac aattctcatg tttgacagct
     4621 tatcatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg
     4681 tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt
     4741 ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct
     4801 gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata acaatttcac
     4861 acaggaaaca gccagtccgt ttaggtgttt tcacgagcac ttcaccaaca aggaccatag
     4921 attatgaaaa ctgaagaagg taaactggta atctggatta acggcgataa aggctataac
     4981 ggtctcgctg aagtcggtaa gaaattcgag aaagataccg gaattaaagt caccgttgag
     5041 catccggata aactggaaga gaaattccca caggttgcgg caactggcga tggccctgac
     5101 attatcttct gggcacacga ccgctttggt ggctacgctc aatctggcct gttggctgaa
     5161 atcaccccgg acaaagcgtt ccaggacaag ctgtatccgt ttacctggga tgccgtacgt
     5221 tacaacggca agctgattgc ttacccgatc gctgttgaag cgttatcgct gatttataac
     5281 aaagatctgc tgccgaaccc gccaaaaacc tgggaagaga tcccggcgct ggataaagaa
     5341 ctgaaagcga aaggtaagag cgcgctgatg ttcaacctgc aagaaccgta cttcacctgg
     5401 ccgctgattg ctgctgacgg gggttatgcg ttcaagtatg aaaacggcaa gtacgacatt
     5461 aaagacgtgg gcgtggataa cgctggcgcg aaagcgggtc tgaccttcct ggttgacctg
     5521 attaaaaaca aacacatgaa tgcagacacc gattactcca tcgcagaagc tgcctttaat
     5581 aaaggcgaaa cagcgatgac catcaacggc ccgtgggcat ggtccaacat cgacaccagc
     5641 aaagtgaatt atggtgtaac ggtactgccg accttcaagg gtcaaccatc caaaccgttc
     5701 gttggcgtgc tgagcgcagg tattaacgcc gccagtccga acaaagagct ggcaaaagag
     5761 ttcctcgaaa actatctgct gactgatgaa ggtctggaag cggttaataa agacaaaccg
     5821 ctgggtgccg tagcgctgaa gtcttacgag gaagagttgg cgaaagatcc acgtattgcc
     5881 gccaccatgg aaaacgccca gaaaggtgaa atcatgccga acatcccgca gatgtccgct
     5941 ttctggtatg ccgtgcgtac tgcggtgatc aacgccgcca gcggtcgtca gactgtcgat
     6001 gaagccctga aagacgcgca gactaattcg agctcgaaca acaacaacaa taacaataac
     6061 aacaacctcg ggatcgaggg aaggatttca gaatttgaaa acctgtattt tcagggccat
     6121 atgggtggga gttggggagg gtctatgtca gaagagaagc ctaaggaggg cgtgaaaacc
     6181 gagaatgacc acattaactt gaaggttgct ggacaggatg ggtccgtggt tcagttcaaa
     6241 attaaacgcc acacaccgtt gtcgaaactg atgaaagcgt acagcgaacg ccagggtctt
     6301 tctatgcgtc agattcgctt ccgctttgat ggtcaaccga ttaatgaaac ggacacgccg
     6361 gcacagctgg agatggagga tgaagacact atcgacgtat ttcagcagca gaccgtcgta
     6421 gggggctccg gcggaagcgg ggggtcggga ggttccatga gcgaagagaa accaaaagaa
     6481 ggagttaaaa ccgaaaacga ccatatcaac ctgaaggtcg ccggtcagga cggctcagtc
     6541 gtgcagttca agattaaacg tcatacccca ttaagtaagc tgatgaaggc gtatagtgag
     6601 cgccaaggct tgtcgatgcg ccagattcgt tttcgctttg atggtcagcc aatcaacgaa
     6661 accgacaccc cggcgcagct ggaaatggaa gatgaagata ccatcgatgt gttccaacag
     6721 caaacggttg ttggcggaag cggcgggagc ggtggtagcg ggggttctat gtccgaagaa
     6781 aaaccgaaag agggtgtgaa gactgaaaac gaccacatta atctgaaggt ggcgggccag
     6841 gacggttccg ttgttcagtt taaaatcaaa cgtcacaccc cgctgtcgaa actgatgaaa
     6901 gcctatagcg aacgtcaagg gctgagcatg cgtcaaattc gttttcgctt tgatggtcag
     6961 ccaatcaacg aaaccgatac accggcgcag ctggagatgg aagacgagga caccattgac
     7021 gtcttccagc aacaaactgt agtgggtggc tccggtggct ctggtgggtc tggtggctcg
     7081 atgagtgagg aaaaacccaa agaaggcgtc aagactgaga acgatcatat taacctgaaa
     7141 gtcgcgggtc aagatggttc agtggtgcag ttcaaaatta aacgccacac cccgctgtca
     7201 aagctgatga aagcctatag tgagcgtcaa ggtctgtcga tgcgccagat ccgctttcgc
     7261 tttgatggtc agccaatcaa cgagacagac actccggcac aactggagat ggaagatgaa
     7321 gacaccatcg atgtttttca acagcagacc gtggtgggag ggagcggcgg tagcggggga
     7381 tcaggtggta gtatgtctga ggaaaagccg aaggaagggg taaagaccga aaatgatcac
     7441 attaatctga aagttgcggg ccaagatggt agtgtagtgc aatttaaaat caagcgtcac
     7501 accccgttat ctaaactgat gaaagcatat agcgaacgcc aaggcctgtc tatgcgccag
     7561 attcgctttc gctttgacgg gcaaccaatt aatgaaacgg acaccccggc acaactggaa
     7621 atggaagacg aggatactat tgacgtgttt cagcaacaga cagtggtcgg cggctccggt
     7681 ggatccgaaa acctgtattt tcagggccat catcatcatc atcactgaaa gcttggcact
     7741 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     7801 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     7861 ttcccaacag ttgcgcagcc tgaatggcga atggcagctt ggctgttttg gcggatgaga
     7921 taagattttc agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa
     7981 tttgcctggc ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa
     8041 acgccgtagc gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc
     8101 atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt
     8161 cggtgaacgc tctcctgagt aggacaaatc cgccgggagc ggatttgaac gttgcgaagc
     8221 aacggcccgg agggtggcgg gcaggacgcc cgccataaac tgccaggcat caaa
//
