LOCUS       sequence-212073-        5206 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for expression of protein in E. coli as PreScission cleavable 
            N-terminual 8His and GST tag fusions and with C-terminal Avi-tag .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5206)
  TITLE     Hyvonen Lab pOP plasmids
REFERENCE   2  (bases 1 to 5206)
  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: pOP5GP
FEATURES             Location/Qualifiers
     source          1..5206
                     /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             98..121
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             137..787
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPP"
     primer_bind     748..770
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             815..838
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             875..919
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     promoter        complement(951..969)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(952..969)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1027..1045)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1041..1088
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1324..1345)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1495..2577)
                     /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     2539..2558
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2578..2655)
                     /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     2801..2820
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2976..2995)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3095..3117
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3155..3173)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3241..3345
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3346..3414,3415..4206)
                     /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      4377..4965
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4866..4885
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5119..5136
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatacgact cactataggg aattgtgagc ggataacaat tcctctagaa ataattttgt
       61 ttaactttaa gaaggagata tatccatgaa tggatctcat caccatcacc atcaccatca
      121 cactagtagc aattccatgt cccctatact aggttattgg aaaattaagg gccttgtgca
      181 acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg
      241 cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct
      301 tccttattat attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat
      361 agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct
      421 tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt
      481 tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga
      541 tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt
      601 gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa
      661 attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc
      721 cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca
      781 tcctccaacc ggtagtggca ccagtgggtc gacactggaa gttctgtttc agggcccagg
      841 atccatggaa ttcgcggccg ccctaggctc gagcggactg aatgacattt tcgaagcaca
      901 gaagatcgaa tggcatgaag cctatgaatg gatctcataa gcttgagtat tctatagtgt
      961 cacctaaatc ccagcttgat ccggctgcta acaaagcccg aaaggaagct gagttggctg
     1021 ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg
     1081 gttttttgct gaaaggagga actatatccg gataacctgg cgtaatagcg aagaggcccg
     1141 caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggaaat tgtaagcgtt
     1201 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     1261 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     1321 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     1381 aaaaccgtct atcagggcga tggcccacag cttgcatgcc tgcaggtcgg aagcataaag
     1441 tgtaaagcct ggggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg
     1501 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     1561 gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg
     1621 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
     1681 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt gacggcggga tataacatga
     1741 gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga
     1801 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
     1861 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
     1921 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
     1981 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg
     2041 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
     2101 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
     2161 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
     2221 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
     2281 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
     2341 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
     2401 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
     2461 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
     2521 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
     2581 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgca
     2641 ccattcgatg gtgtcgggta ccgagctcga atttacgtga accatcaccc aaatcaagtt
     2701 ttttgcggtc gaggtgccgt aaagctctaa atcggaaccc taaagggagc ccccgattta
     2761 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     2821 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     2881 cgcttaatgc gccgctacag ggcgcgtcgc ctgatgcggt attttctcct tacgcatctg
     2941 tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag
     3001 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     3061 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     3121 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     3181 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     3241 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     3301 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     3361 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     3421 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     3481 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     3541 atgatgagca cttttaaagt tctgctatgt catacactat tatcccgtat tgacgccggg
     3601 caagagcaac tcggtcgccg ggcgcggtat tctcagaatg acttggttga gtactcacca
     3661 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     3721 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     3781 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     3841 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     3901 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     3961 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     4021 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     4081 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     4141 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     4201 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     4261 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     4321 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     4381 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     4441 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     4501 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     4561 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     4621 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     4681 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     4741 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     4801 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     4861 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     4921 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     4981 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5041 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     5101 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     5161 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctgat
//
