LOCUS       sequence-171339-        7230 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of full length human Uba2 with Cterminal His-tag in 
            E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7230)
  AUTHORS   Werner A, Moutty MC, Moller U, Melchior F
  TITLE     Performing in vitro sumoylation reactions using recombinant enzymes.
  JOURNAL   Methods Mol Biol. 2009;497:187-99. doi: 
            10.1007/978-1-59745-566-4_12.
  PUBMED    19107418
REFERENCE   2  (bases 1 to 7230)
  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: "2009"; volume: "497:187-99. doi"; pages: " 
            10.1007/978-1-59745-566-4_12"
COMMENT     Sequence Label: pET28b-hUba2-His
FEATURES             Location/Qualifiers
     source          1..7230
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     71..90
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        71..89
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    90..114
                     /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             129..151
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2087..2116
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="ASMTGGQQMG"
     CDS             2165..2182
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2235..2253)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2249..2296
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2333..2788
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2420..2439)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2630..2651
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2880..3695)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2880..3695)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3603..3622
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3687..3706
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3817..4405
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4306..4325
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4559..4576
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4591..4730
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4656..4675)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4816..4838
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4832..5023)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5832..6914)
                     /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     6876..6895
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6915..6992)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7198..7217
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat
       61 cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta
      121 gaaataattt tgtttaactt taagaaggag atataccatg gggatggcac tgtcgcgggg
      181 gctgccccgg gagctggctg aggcggtggc cgggggccgg gtgctggtgg tgggggcggg
      241 cggcatcggc tgcgagctcc tcaagaatct cgtgctcacc ggtttctccc acatcgacct
      301 gattgatctg gatactattg atgtaagcaa cctcaacaga cagtttttgt ttcaaaagaa
      361 acatgttgga agatcaaagg cacaggttgc caaggaaagt gtactgcagt tttacccgaa
      421 agctaatatc gttgcctacc atgacagcat catgaaccct gactataatg tggaattttt
      481 ccgacagttt atactggtta tgaatgcttt agataacaga gctgcccgaa accatgttaa
      541 tagaatgtgc ctggcagctg atgttcctct tattgaaagt ggaacagctg ggtatcttgg
      601 acaagtaact actatcaaaa agggtgtgac cgagtgttat gagtgtcatc ctaagccgac
      661 ccagagaacc tttcctggct gtacaattcg taacacacct tcagaaccta tacattgcat
      721 cgtttgggca aagtacttgt tcaaccagtt gtttggggaa gaagatgctg atcaagaagt
      781 atctcctgac agagctgacc ctgaagctgc ctgggaacca acggaagccg aagccagagc
      841 tagagcatct aatgaagatg gtgacattaa acgtatttct actaaggaat gggctaaatc
      901 aactggatat gatccagtta aactttttac caagcttttt aaagatgaca tcaggtatct
      961 gttgacaatg gacaaactat ggcggaaaag gaaacctcca gttccgttgg actgggctga
     1021 agtacaaagt caaggagaag aaacgaatgc atcagatcaa cagaatgaac cccagttagg
     1081 cctgaaagac cagcaggttc tagatgtaaa gagctatgca cgtctttttt caaagagcat
     1141 cgagactttg agagttcatt tagcagaaaa gggggatgga gctgagctca tatgggataa
     1201 ggatgaccca tctgcaatgg attttgtcac ctctgctgca aacctcagga tgcatatttt
     1261 cagtatgaat atgaagagta gatttgatat caaatcaatg gcagggaaca ttattcctgc
     1321 tattgctact actaatgcag taattgctgg gttgatagta ttggaaggat tgaagatttt
     1381 atcaggaaaa atagaccagt gcagaacaat ttttttgaat aaacaaccaa acccaagaaa
     1441 gaagcttctt gtgccttgtg cactggatcc tcccaacccc aattgttatg tatgtgccag
     1501 caagccagag gtgactgtgc ggctgaatgt ccataaagtg actgttctca ccttacaaga
     1561 caagatagtg aaagaaaaat ttgctatggt agcaccagat gtccaaattg aagatgggaa
     1621 aggaacaatc ctaatatctt ccgaagaggg agagacggaa gctaataatc acaagaagtt
     1681 gtcagaattt ggaattagaa atggcagccg gcttcaagca gatgacttcc tccaggacta
     1741 tactttattg atcaacatcc ttcatagtga agacctagga aaggacgttg aatttgaagt
     1801 tgttggtgat gccccggaaa aagtggggcc caaacaagct gaagatgctg ccaaaagcat
     1861 aaccaatggc agtgatgatg gagctcagcc ctccacctcc acagctcaag agcaagatga
     1921 cgttctcata gttgattcgg atgaagaaga ttcttcaaat aatgccgacg tcagtgaaga
     1981 agagagaagc cgcaagagga aattagatga gaaagagaat ctcagtgcaa agaggtcacg
     2041 tatagaacag aaggaagagc ttgatgatgt catagcatta gatggagcta gcatgactgg
     2101 tggacagcaa atgggtcggg atccgaattc gagctccgtc gacaagcttg cggccgcact
     2161 cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa aggaagctga
     2221 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     2281 cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg ggacgcgccc
     2341 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2401 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2461 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2521 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     2581 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     2641 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2701 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2761 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     2821 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct
     2881 tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata
     2941 ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat
     3001 aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct
     3061 attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact
     3121 gaatccggtg agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag
     3181 ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc
     3241 gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa
     3301 tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat
     3361 tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca
     3421 tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt
     3481 agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac
     3541 aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca
     3601 ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc
     3661 ctagagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg
     3721 taagcagaca gttttattgt tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3781 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3841 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     3901 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     3961 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     4021 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     4081 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     4141 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     4201 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     4261 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     4321 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     4381 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4441 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     4501 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     4561 cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct
     4621 gtgcggtatt tcacaccgca atggtgcact ctcagtacaa tctgctctga tgccgcatag
     4681 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
     4741 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     4801 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     4861 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg
     4921 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
     4981 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag
     5041 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
     5101 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
     5161 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata
     5221 cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa
     5281 tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca
     5341 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc
     5401 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca
     5461 acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata atggcctgct
     5521 tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga
     5581 ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc
     5641 cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag
     5701 tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga
     5761 aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa
     5821 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     5881 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     5941 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     6001 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     6061 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca
     6121 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     6181 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     6241 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     6301 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     6361 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     6421 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     6481 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     6541 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     6601 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     6661 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     6721 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     6781 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     6841 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     6901 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     6961 ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga
     7021 ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag
     7081 gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
     7141 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
     7201 gatgtcggcg atataggcgc cagcaaccgc
//
