LOCUS       sequence-194708-        6898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6898)
  TITLE     SGC Toronto Q2 Q3 2016
REFERENCE   2  (bases 1 to 6898)
  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: 5JJW (USP15_SART3)
FEATURES             Location/Qualifiers
     source          1..6898
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             91..108
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             121..141
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1763..1780
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1833..1851)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1847..1894
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1931..2386
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2018..2037)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2228..2249
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2478..3293)
                     /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(2478..3293)
                     /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     3201..3220
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3285..3304
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3415..4003
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3904..3923
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4157..4174
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4189..4328
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4254..4273)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4414..4436
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4430..4621)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5430..6512)
                     /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     6474..6493
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6513..6590)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6796..6815
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg catcatcatc atcatcacag cagcggcaga
      121 gaaaacttgt atttccaggg catggcggaa ggcggagcgg cggatctgga cacccagcgg
      181 tctgacatcg cgacgctgct caaaacctcg ctccggaaag gggacacctg gtacctagtc
      241 gatagtcgct ggttcaaaca gtggaaaaaa tatgttggct ttgacagttg ggacaaatac
      301 cagatgggag atcaaaatgt gtatcctgga cccattgata actctggact tctcaaagat
      361 ggtgatgccc agtcacttaa ggaacacctt attgatgaat tggattacat actgttgcca
      421 actgaaggtt ggaataaact tgtcagctgg tacacattga tggaaggtca agagccaata
      481 gcacgaaagg tggttgaaca gggtatgttt gtaaagcact gcaaagtaga agtatatctc
      541 acagaattga agctatgtga aaatggaaac atgaataatg ttgtaactcg aagatttagc
      601 aaagctgaca caatagatac aattgaaaag gaaataagaa aaatcttcag tattccagat
      661 gaaaaggaga ccagattgtg gaacaaatac atgagtaaca catttgaacc actgaataaa
      721 ccagacagca ccattcagga tgctggttta taccaaggac aggtattagt gatagaacag
      781 aaaaatgaag atggaacatg gccaaggggt tgactcgacg gaggaataat catatgtcag
      841 taattcagaa ctataacaaa gcactacagc agctggagaa atataaaccc tatgaagaag
      901 cactgttgca ggcagaggca ccaaggctgg cagaatatca agcatatatc gattttgaga
      961 tgaaaattgg cgatcctgct cgcattcagt tgatctttga gcgcgccctg gtcgagaact
     1021 gccttgtccc agacttatgg atccgttaca gtcagtacct agatcgacaa ctgaaagtaa
     1081 aggatttggt tttatctgta cataaccgcg ctattagaaa ctgcccctgg acagttgcct
     1141 tatggagtcg gtacctcttg gccatggaga gacatggagt tgatcatcaa gtaatttctg
     1201 taaccttcga gaaagctttg aatgccggct tcatccaggc cactgattat gtggagattt
     1261 ggcaggcata ccttgattac ctgaggagaa gggttgattt caaacaagac tccagtaaag
     1321 agctggagga gttgagggcc gcctttactc gtgccttgga gtatctgaag caggaggtgg
     1381 aagagcgttt caatgagagt ggtgatccaa gctgcgtgat tatgcagaac tgggctagga
     1441 ttgaggctcg actgtgcaat aacatgcaga aagctcggga actctgggat agcatcatga
     1501 ccagaggaaa tgccaagtac gccaacatgt ggctagagta ttacaacctg gaaagagctc
     1561 atggtgacac ccagcactgc cggaaggctc tgcaccgggc cgtccagtgc accagtgact
     1621 acccagagca cgtctgcgaa gtgttactca ccatggagag gacagaaggt tctttagaag
     1681 attgggatat agctgttcag aaaactgaaa cccgattagc tcgtgtcaat gagtgatgac
     1741 gaagcttgcg gccgcactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa
     1801 agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag cataacccct
     1861 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatt
     1921 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1981 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2041 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2101 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2161 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2221 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     2281 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     2341 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     2401 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2461 cgctcatgaa ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc
     2521 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     2581 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     2641 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     2701 tcaccatgag tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag
     2761 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg
     2821 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa
     2881 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     2941 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg
     3001 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     3061 aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct
     3121 ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc
     3181 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     3241 ttggaattta atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc
     3301 cttgtattac tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg
     3361 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3421 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3481 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3541 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     3601 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3661 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3721 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3781 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3841 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3901 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3961 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4021 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4081 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4141 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     4201 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     4261 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     4321 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4381 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4441 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     4501 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     4561 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     4621 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     4681 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     4741 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     4801 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     4861 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     4921 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     4981 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     5041 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     5101 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     5161 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     5221 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     5281 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     5341 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     5401 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5461 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5521 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     5581 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     5641 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     5701 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     5761 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     5821 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     5881 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     5941 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     6001 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     6061 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     6121 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     6181 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     6241 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     6301 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     6361 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     6421 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     6481 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     6541 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     6601 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     6661 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     6721 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     6781 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     6841 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaat
//
