LOCUS       sequence-220240-        5186 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 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: pOP5GP
FEATURES             Location/Qualifiers
     source          1..5186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     43..62
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        43..61
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    61..85
                     /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             98..120
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             140..163
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             179..829
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPP"
     primer_bind     790..812
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             857..880
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             917..961
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     promoter        complement(978..996)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(979..996)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1054..1072)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1068..1115
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1351..1372)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1522..2604)
                     /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     2566..2585
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2605..2682)
                     /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     2825..2844
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2998..3017)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3117..3139
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3177..3195)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3263..3367
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3368..3436,3437..4228)
                     /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(3586..3605)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4399..4987
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4888..4907
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5141..5158
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg attaatacga ctcactatag
       61 ggaattgtga gcggataaca attcctctag aaataatttt gtttaacttt aagaaggaga
      121 tatatccatg aatggatctc atcaccatca ccatcaccat cacactagta gcaattccat
      181 gtcccctata ctaggttatt ggaaaattaa gggccttgtg caacccactc gacttctttt
      241 ggaatatctt gaagaaaaat atgaagagca tttgtatgag cgcgatgaag gtgataaatg
      301 gcgaaacaaa aagtttgaat tgggtttgga gtttcccaat cttccttatt atattgatgg
      361 tgatgttaaa ttaacacagt ctatggccat catacgttat atagctgaca agcacaacat
      421 gttgggtggt tgtccaaaag agcgtgcaga gatttcaatg cttgaaggag cggttttgga
      481 tattagatac ggtgtttcga gaattgcata tagtaaagac tttgaaactc tcaaagttga
      541 ttttcttagc aagctacctg aaatgctgaa aatgttcgaa gatcgtttat gtcataaaac
      601 atatttaaat ggtgatcatg taacccatcc tgacttcatg ttgtatgacg ctcttgatgt
      661 tgttttatac atggacccaa tgtgcctgga tgcgttccca aaattagttt gttttaaaaa
      721 acgtattgaa gctatcccac aaattgataa gtacttgaaa tccagcaagt atatagcatg
      781 gcctttgcag ggctggcaag ccacgtttgg tggtggcgac catcctccaa ccggtagtgg
      841 caccagtggg tcgacactgg aagttctgtt tcagggccca ggatccatgg aattcgcggc
      901 cgccctaggc tcgagcggac tgaatgacat tttcgaagca cagaagatcg aatggcatga
      961 agcctaagct tgagtattct atagtgtcac ctaaatccca gcttgatccg gctgctaaca
     1021 aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc
     1081 ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat
     1141 aacctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     1201 gaatggcgaa tggaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     1261 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     1321 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     1381 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccacagctt
     1441 gcatgcctgc aggtcggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     1501 cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     1561 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt
     1621 ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg
     1681 agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat
     1741 ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga
     1801 gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat
     1861 ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt
     1921 ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg
     1981 attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa
     2041 tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
     2101 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
     2161 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
     2221 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
     2281 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
     2341 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
     2401 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
     2461 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
     2521 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
     2581 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
     2641 tcatgccata ccgcgaaagg ttttgcacca ttcgatggtg tcgggtaccg agctcgaatt
     2701 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     2761 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     2821 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     2881 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcctgatgcg
     2941 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca ctctcagtac
     3001 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     3061 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     3121 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     3181 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     3241 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     3301 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     3361 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     3421 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     3481 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     3541 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     3601 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     3661 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     3721 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     3781 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     3841 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     3901 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     3961 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     4021 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     4081 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     4141 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     4201 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     4261 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     4321 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     4381 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     4441 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     4501 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     4561 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     4621 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     4681 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     4741 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     4801 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     4861 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     4921 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     4981 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5041 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     5101 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     5161 agcggaagag cgcccaatac gcaaac
//
