LOCUS       sequence-291945-        4720 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of SENP_EuB protease.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4720)
  AUTHORS   Vera Rodriguez A, Frey S, Gorlich D
  TITLE     Engineered SUMO/protease system identifies Pdr6 as a bidirectional 
            nuclear transport receptor.
  JOURNAL   J Cell Biol. 2019 Jun 3;218(6):2006-2020. doi: 
            10.1083/jcb.201812091. Epub 2019 Apr 25.
  PUBMED    31023724
REFERENCE   2  (bases 1 to 4720)
  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: 
            "10.1083/jcb.201812091"; journalName: "J Cell Biol"; date: 
            "2019-06-3- 3"; volume: "218"; issue: "6"; pages: "2006-2020"
COMMENT     Sequence Label: pAV0286
FEATURES             Location/Qualifiers
     source          1..4720
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             193..213
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     terminator      962..1056
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(1106..1125)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1117..1932
                     /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             1117..1932
                     /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     complement(1190..1209)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2151..2237
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2331..3413)
                     /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     3375..3394
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3414..3491)
                     /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."
     primer_bind     complement(3575..3597)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3683..3823
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3738..3757
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3838..3855)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4009..4597)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4089..4108)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4688..4706
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aaccatgagc
      121 aagcatcacc atcattcagg ccatcaccat accggacacc accatcattc aggcagtcat
      181 caccataccg gcgagaacct gtattttcag ggtaccaagg aagaagtgcc ggaaccgttt
      241 gtgccgttga ccgacgaaga cgaagataac gttcgtcacg cactgggtgg ccgcaaacgt
      301 tccgaaaccc tgtccgtgca cgaagcgtca tctattgtga tcacgcgcga gatcctgcag
      361 tgtctgaacg acaaagaatg gctgaatgat gaggttatca acctgtacct ggaactgctg
      421 aaagagcgtg aactgcgtga accgaacaaa tttctgaaat gtcatttctt caatacgttt
      481 ttctacaaaa agctgattaa cggcggctat gattacaaaa gcgttcgtga gtggactacg
      541 aagcgcaaac tgggttacaa cctgattgat tgcgacaaaa tcttcgtgcc gattcacaaa
      601 gatgttcatt ggtgcctggc tgtaattaac atcaaagaaa agaaattcca gtatctcgat
      661 agcctgggct atatggatat gaaagcgctg cgcattctgg ctaaatatct cgtggacgaa
      721 gtaaaagaca aatccggtaa gcagatcgac gttcacgcgt ggaaacagga gggtgttcag
      781 aacctgccgc tgcaggaaaa cggttgggat tgcggtatgt tcatgctgaa gtatatcgat
      841 ttctattctc gtgacatgga actggtcttt ggccagaaac acatgtcgta ctttcgtcgt
      901 cgtaccgcta aagaaatcct ggatctgaag gccggttagt aagcttaatt agctgagctt
      961 ggactcctgt tgatagatcc agtaatgacc tcagaactcc atctggattt gttcagaacg
     1021 ctcggttgcc gccgggcgtt ttttattggt gagaatccaa gctagccatg aacaataaaa
     1081 ctgtctgctt acataaacag taatacaagg ggtgttatga gccatattca acgggaaacg
     1141 tcttgctcta ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg
     1201 gctcgcgata atgtcgggca atcaggtgcg acaatctatc gattgtatgg gaagcccgat
     1261 gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag
     1321 atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc
     1381 cgtactcctg atgatgcttg gttactcacg actgcgatcc ccggcaaaac agcattccag
     1441 gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg
     1501 cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt
     1561 ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac
     1621 gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcacaaact tttgccattc
     1681 tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag
     1741 gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat
     1801 cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt
     1861 caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat
     1921 gagtttttct aagaattaat tcatgggcaa atattatacg caaggcgaca aggtgctgat
     1981 gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg gcagaatgct
     2041 taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaatttt tttaaggcag
     2101 ttattggtgc ccttaaacgc ctggggtaat gactctctag cttgaggcat caaataaaac
     2161 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     2221 tcctgagtag gacaaatccg ccgctctaga ttacgtgcag tcgatgataa gctgtcaaac
     2281 atgagaattg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg
     2341 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     2401 gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac
     2461 agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt
     2521 tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg
     2581 tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg
     2641 gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga
     2701 acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg
     2761 ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc
     2821 agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga
     2881 cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata
     2941 ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca
     3001 gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg
     3061 cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc
     3121 atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt
     3181 tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg
     3241 cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc
     3301 actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc
     3361 tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc
     3421 accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcaccat
     3481 tcgatggtgt cggaatttcg ggcagcgttg ggtcctggcc acgggtgcgc atgatctaga
     3541 gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     3601 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     3661 cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt
     3721 atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc atatgcggtg
     3781 tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt ccgcttcctc
     3841 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3901 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3961 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4021 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     4081 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     4141 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     4201 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     4261 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     4321 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     4381 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     4441 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     4501 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     4561 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     4621 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgacattaac
     4681 ctataaaaat aggcgtatca cgaggccctt tcgtcttcac
//
