LOCUS       sequence-220235-        5186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used for expression of protein in E. coli as PreScission 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 5186)
  TITLE     Hyvonen Lab pOP plasmids
REFERENCE   2  (bases 1 to 5186)
  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: pOP3GP
FEATURES             Location/Qualifiers
     source          1..5186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     39..58
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        39..57
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    57..81
                     /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             94..116
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             130..174
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             184..207
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             223..873
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPP"
     primer_bind     834..856
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             901..924
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     promoter        complement(974..992)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(975..992)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1050..1068)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1064..1111
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1347..1368)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1518..2600)
                     /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     2562..2581
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2601..2678)
                     /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     2821..2840
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2994..3013)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3113..3135
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3173..3191)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3259..3363
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3364..3432,3433..4224)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3582..3601)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4395..4983
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4884..4903
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5137..5154
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tctccccgcg cgttggccga ttcattaatg cagctgatta atacgactca ctatagggaa
       61 ttgtgagcgg ataacaattc ctctagaaat aattttgttt aactttaaga aggagatata
      121 tccatgaatg gactgaatga tatctttgaa gcgcagaaaa ttgaatggca tgaatccgga
      181 tctcatcacc atcaccatca ccatcacact agtagcaatt ccatgtcccc tatactaggt
      241 tattggaaaa ttaagggcct tgtgcaaccc actcgacttc ttttggaata tcttgaagaa
      301 aaatatgaag agcatttgta tgagcgcgat gaaggtgata aatggcgaaa caaaaagttt
      361 gaattgggtt tggagtttcc caatcttcct tattatattg atggtgatgt taaattaaca
      421 cagtctatgg ccatcatacg ttatatagct gacaagcaca acatgttggg tggttgtcca
      481 aaagagcgtg cagagatttc aatgcttgaa ggagcggttt tggatattag atacggtgtt
      541 tcgagaattg catatagtaa agactttgaa actctcaaag ttgattttct tagcaagcta
      601 cctgaaatgc tgaaaatgtt cgaagatcgt ttatgtcata aaacatattt aaatggtgat
      661 catgtaaccc atcctgactt catgttgtat gacgctcttg atgttgtttt atacatggac
      721 ccaatgtgcc tggatgcgtt cccaaaatta gtttgtttta aaaaacgtat tgaagctatc
      781 ccacaaattg ataagtactt gaaatccagc aagtatatag catggccttt gcagggctgg
      841 caagccacgt ttggtggtgg cgaccatcct ccaaccggta gtggcaccag tgggtcgaca
      901 ctggaagttc tgtttcaggg cccaggatcc atggaattcg cggccgccct aggctcgagc
      961 taagcttgag tattctatag tgtcacctaa atcccagctt gatccggctg ctaacaaagc
     1021 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1081 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggataacc
     1141 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     1201 ggcgaatgga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     1261 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     1321 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     1381 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca cagcttgcat
     1441 gcctgcaggt cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactta
     1501 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     1561 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt
     1621 tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag
     1681 agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg
     1741 gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata
     1801 tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga
     1861 tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt
     1921 tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg
     1981 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg
     2041 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc
     2101 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga
     2161 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc
     2221 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta
     2281 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg
     2341 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg
     2401 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg
     2461 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg
     2521 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg
     2581 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat
     2641 gccataccgc gaaaggtttt gcaccattcg atggtgtcgg gtaccgagct cgaattgtga
     2701 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2761 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2821 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2881 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcct gatgcggtat
     2941 tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct cagtacaatc
     3001 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     3061 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     3121 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     3181 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     3241 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     3301 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     3361 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     3421 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     3481 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     3541 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     3601 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     3661 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     3721 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     3781 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     3841 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     3901 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     3961 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     4021 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     4081 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     4141 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     4201 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     4261 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4321 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4381 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4441 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4501 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     4561 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     4621 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     4681 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4741 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4801 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4861 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4921 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4981 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5041 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     5101 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     5161 gaagagcgcc caatacgcaa accgcc
//
