LOCUS       sequence-212070-        5185 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for expression of protein in E. coli as TEV cleavable 
            N-terminal Avi-His8-tag and GST fusions.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5185)
  TITLE     Hyvonen Lab pOP plasmids
REFERENCE   2  (bases 1 to 5185)
  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: pOP3GT
FEATURES             Location/Qualifiers
     source          1..5185
                     /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    19..43
                     /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             56..78
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             92..136
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             146..169
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             185..835
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPP"
     primer_bind     796..818
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             863..883
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     promoter        complement(930..948)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(931..948)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1006..1024)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1020..1067
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1303..1324)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1474..2556)
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     2518..2537
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2557..2634)
                     /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     2780..2799
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2955..2974)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3074..3096
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3134..3152)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3220..3324
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3325..3393,3394..4185)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCHTLLSRIDAGQEQLGRRARYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4356..4944
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4845..4864
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5098..5115
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatacgact cactataggg aattgtgagc ggataacaat tcctctagaa ataattttgt
       61 ttaactttaa gaaggagata tatccatgaa tggactgaat gatatctttg aagcgcagaa
      121 aattgaatgg catgaatccg gatctcatca ccatcaccat caccatcaca ctagtagcaa
      181 ttccatgtcc cctatactag gttattggaa aattaagggc cttgtgcaac ccactcgact
      241 tcttttggaa tatcttgaag aaaaatatga agagcatttg tatgagcgcg atgaaggtga
      301 taaatggcga aacaaaaagt ttgaattggg tttggagttt cccaatcttc cttattatat
      361 tgatggtgat gttaaattaa cacagtctat ggccatcata cgttatatag ctgacaagca
      421 caacatgttg ggtggttgtc caaaagagcg tgcagagatt tcaatgcttg aaggagcggt
      481 tttggatatt agatacggtg tttcgagaat tgcatatagt aaagactttg aaactctcaa
      541 agttgatttt cttagcaagc tacctgaaat gctgaaaatg ttcgaagatc gtttatgtca
      601 taaaacatat ttaaatggtg atcatgtaac ccatcctgac ttcatgttgt atgacgctct
      661 tgatgttgtt ttatacatgg acccaatgtg cctggatgcg ttcccaaaat tagtttgttt
      721 taaaaaacgt attgaagcta tcccacaaat tgataagtac ttgaaatcca gcaagtatat
      781 agcatggcct ttgcagggct ggcaagccac gtttggtggt ggcgaccatc ctccaaccgg
      841 tagtggcacc agtgggtcga cagaaaacct gtacttccag ggatccatgg aattcgcggc
      901 cgccctaggc tcgagctaag cttgagtatt ctatagtgtc acctaaatcc cagcttgatc
      961 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
     1021 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
     1081 ctatatccgg ataacctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     1141 gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta atattttgtt aaaattcgcg
     1201 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     1261 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     1321 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     1381 ggcccacagc ttgcatgcct gcaggtcgga agcataaagt gtaaagcctg gggtgcctaa
     1441 tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     1501 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     1561 gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac
     1621 cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa
     1681 atcctgtttg atggtggttg acggcgggat ataacatgag ctgtcttcgg tatcgtcgta
     1741 tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc
     1801 gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag
     1861 catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat
     1921 cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga
     1981 gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg
     2041 ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg
     2101 gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc
     2161 atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt
     2221 gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct
     2281 ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag
     2341 ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc
     2401 cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt
     2461 cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc
     2521 atactctgcg acatcgtata acgttactgg tttcacattc accaccctga attgactctc
     2581 ttccgggcgc tatcatgcca taccgcgaaa ggttttgcac cattcgatgg tgtcgggtac
     2641 cgagctcgaa tttacgtgaa ccatcaccca aatcaagttt tttgcggtcg aggtgccgta
     2701 aagctctaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg
     2761 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa
     2821 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg
     2881 gcgcgtcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2941 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     3001 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     3061 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     3121 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     3181 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     3241 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     3301 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     3361 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     3421 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     3481 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     3541 ctgctatgtc atacactatt atcccgtatt gacgccgggc aagagcaact cggtcgccgg
     3601 gcgcggtatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     3661 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     3721 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     3781 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     3841 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     3901 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     3961 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     4021 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     4081 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     4141 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     4201 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     4261 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     4321 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     4381 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     4441 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     4501 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     4561 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     4621 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     4681 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     4741 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     4801 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     4861 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     4921 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     4981 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5041 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     5101 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     5161 tggccgattc attaatgcag ctgat
//
