LOCUS       sequence-387118-        6814 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  protein expression of the GST-tagged catalytic domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6814)
  TITLE     Knapp lab plasmids
REFERENCE   2  (bases 1 to 6814)
  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"
COMMENT     Sequence Label: pGTVL2-SENP3
FEATURES             Location/Qualifiers
     source          1..6814
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     41..63
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(57..248)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1057..2139)
                     /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     2101..2120
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2140..2217)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2423..2442
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2526..2545
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2526..2544
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2545..2569
                     /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             2584..2606
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2617..2634
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2641..3294
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGADHPPK"
     CDS             3322..3342
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             4206..4223
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4276..4294)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4290..4337
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4374..4829
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4461..4480)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4671..4692
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4919..5734)
                     /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(4919..5734)
                     /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     5642..5661
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5726..5745
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5856..6444
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6345..6364
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6598..6615
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6630..6769
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6695..6714)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
       61 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
      121 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
      181 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
      241 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
      301 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
      361 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
      421 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
      481 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
      541 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
      601 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
      661 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc
      721 gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg
      781 aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc
      841 gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct
      901 acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc
      961 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     1021 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     1081 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     1141 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     1201 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     1261 aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc
     1321 gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     1381 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt
     1441 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     1501 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc
     1561 cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag
     1621 atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt
     1681 ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat
     1741 ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag
     1801 attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac
     1861 gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg
     1921 cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg
     1981 tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt
     2041 tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc
     2101 ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact
     2161 ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg
     2221 gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga
     2281 ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc
     2341 ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt
     2401 ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg
     2461 tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc tcgatcccgc
     2521 gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc ctctagaaat
     2581 aattttgttt aactttaaga aggagatata catatgcacc atcatcatca tcattcttct
     2641 atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt
     2701 ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa
     2761 tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat
     2821 ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac
     2881 atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg
     2941 gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt
     3001 gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa
     3061 acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat
     3121 gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa
     3181 aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca
     3241 tggcctttgc agggctggca agccacgttt ggtggtgccg accatcctcc aaaatcgagc
     3301 tcaggtgtag atctgggtac cgagaacctg tacttccaat ccatgagccc cctgcgagag
     3361 gagcatgtga cctgcgtaca gagcatcttg gacgaattcc ttcaaacgta tggcagcctc
     3421 atacccctca gcactgatga ggtagtagag aagctggagg acattttcca gcaggagttt
     3481 tccacccctt ccaggaaggg cctggtgttg cagctgatcc agtcttacca gcggatgcca
     3541 ggcaatgcca tggtgagggg cttccgagtg gcttataagc ggcacgtgct gaccatggat
     3601 gacttgggga ccttgtatgg acagaactgg ctcaatgacc aggtgatgaa catgtatgga
     3661 gacctggtca tggacacagt ccctgaaaag gtgcatttct tcaatagttt cttctatgat
     3721 aaactccgta ccaagggtta tgatggggtg aaaaggtgga ccaaaaacgt ggacatcttc
     3781 aataaggagc tactgctaat ccccatccac ctggaggtgc attggtccct catctctgtt
     3841 gatgtgaggc gacgcaccat cacctatttt gactcgcagc gtaccctaaa ccgccgctgc
     3901 cctaagcata ttgccaagta tctacaggca gaggcggtaa agaaagaccg actggatttc
     3961 caccagggct ggaaaggtta cttcaaaatg aatgtggcca ggcagaataa tgacagtgac
     4021 tgtggtgctt ttgtgttgca gtactgcaag catctggccc tgtctcagcc attcagcttc
     4081 acccagcagg acatgcccaa acttcgtcgg cagatctaca aggagctgtg tcactgcaaa
     4141 ctcacttaaa ggtggatacg gatccgaatt cgagctccgt cgacaagctt gcggccgcac
     4201 tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg
     4261 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     4321 tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc
     4381 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     4441 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     4501 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     4561 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     4621 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     4681 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     4741 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     4801 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
     4861 acccctattt gtttattttt ctaaakwmww tmwrakwwrw mtysgyytrw sarttamctt
     4921 agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac
     4981 catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata
     5041 ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta
     5101 ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg
     5161 aatccggtga gaatggcaaa agcttatgca tttctttcca gacttgttca acaggccagc
     5221 cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg
     5281 cctgagcgag acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat
     5341 gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt
     5401 cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat
     5461 caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta
     5521 gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca
     5581 actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat
     5641 tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc
     5701 tggagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt
     5761 aagcagacag ttttattgtt catgaysawa wwyyyttawc ktgwgywwts kwymcaswga
     5821 gcgtcagacc ccgtagaaar gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     5881 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5941 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6001 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6061 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6121 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6181 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6241 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6301 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6361 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6421 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6481 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6541 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6601 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     6661 tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     6721 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     6781 gccaacaccc gctgacgcgc cctgacgggc ttgt
//
