LOCUS       sequence-82751-e        5577 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E.coli expression of GST tagged human RanBP2 fragment with SUMO 
            ligase activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5577)
  AUTHORS   Flotho A, Werner A, Winter T, Frank AS, Ehret H, Melchior F
  TITLE     Recombinant reconstitution of sumoylation reactions in vitro.
  JOURNAL   Methods Mol Biol. 2012;832:93-110. doi: 10.1007/978-1-61779-474-2_5.
  PUBMED    22350878
REFERENCE   2  (bases 1 to 5577)
  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: "Methods Mol
            Biol."; date: "2012"; volume: "832:93-110. doi"; pages: " 
            10.1007/978-1-61779-474-2_5"
COMMENT     Sequence Label: pGEX3x-hRanBP2-(IR1+M)
FEATURES             Location/Qualifiers
     source          1..5577
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             921..932
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1627..1649)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1794..1812)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1880..1984
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1985..2053,2054..2845)
                     /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(2203..2222)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3016..3604
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3505..3524
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3758..3775
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3848..3925
                     /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             3926..5008
                     /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(3945..3964)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5021..5042
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5057..5062,5063..5080,5081..5087)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5095..5111
                     /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     5119..5135
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5119..5135
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5131..5577,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5151..5168)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5151..5167)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5160..5182)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg atcgaaggtc gtgggatccc cgatgttctc attgtatatg
      961 aactaactcc aaccgctgag cagaaagccc ttgcaaccaa acttaaactt cctccaactt
     1021 tcttctgcta caagaataga ccagattatg ttagtgaaga agaggaggat gatgaagatt
     1081 tcgaaacagc tgtcaagaaa cttaatggaa aactatattt ggatggctca gaaaaatgta
     1141 gacccttgga agaaaataca gcagataatg agaaagaatg tatttgagtt tgggaaaaga
     1201 aaccaacagt tgaagagaag gcaaaagcag atacgttaaa acttccacct acattttttt
     1261 gtggagtctg tagtgatact gatgaagaca atggaaatgg ggaagacttt caatcagagc
     1321 ttcaaaaagt tcaggaagct caaaaatctc agacagaaga aataactagc acaactgaca
     1381 gtgtatatac aggtgggact gaagtgatgg taccttcttt ctgtaaatct gaagaacctg
     1441 attctattac caaatccatt agttcaccat ctgtttcctc tgaaactatg gacaaacctg
     1501 tagatttgtc aactagaaag gaaattgata cagattctac aagccaaggg gaaagcaaga
     1561 tagtttagga attcatcgtg actgactgac gatctgcctc gcgcgtttcg gtgatgacgg
     1621 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     1681 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc
     1741 catgacccag tcacgtagcg atagcggagt gtataattct tgaagacgaa agggcctcgt
     1801 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     1861 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     1921 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     1981 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     2041 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     2101 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     2161 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     2221 atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     2281 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     2341 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     2401 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     2461 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     2521 gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     2581 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     2641 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     2701 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     2761 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     2821 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     2881 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     2941 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     3001 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     3061 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     3121 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     3181 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     3241 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     3301 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     3361 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     3421 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     3481 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     3541 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     3601 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     3661 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     3721 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     3781 ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     3841 aaattccgac accatcgaat ggtgcaaaac ctttcgcggt atggcatgat agcgcccgga
     3901 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta
     3961 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc
     4021 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt
     4081 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc
     4141 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc
     4201 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca
     4261 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga
     4321 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga
     4381 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga
     4441 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
     4501 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
     4561 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
     4621 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
     4681 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg
     4741 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
     4801 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
     4861 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
     4921 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     4981 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     5041 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     5101 gcggataaca atttcacaca ggaaacagct atgaccatga ttacggattc actggccgtc
     5161 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     5221 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     5281 cagttgcgca gcctgaatgg cgaatggcgc tttgcctggt ttccggcacc agaagcggtg
     5341 ccggaaagct ggctggagtg cgatcttcct gaggccgata ctgtcgtcgt cccctcaaac
     5401 tggcagatgc acggttacga tgcgcccatc tacaccaacg taacctatcc cattacggtc
     5461 aatccgccgt ttgttcccac ggagaatccg acgggttgtt actcgctcac atttaatgtt
     5521 gatgaaagct ggctacagga aggccagacg cgaattattt ttgatggcgt tggaatt
//
