LOCUS       sequence-408951-        5847 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of VP1 protein from rhinovirus B14 (RV-B14). 
            Contains N-terminal GST-tag and C-terminal His6-tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5847)
  AUTHORS   Jouppila NVV, Lehtonen J, Seppala E, Puustinen L, Oikarinen S, 
            Laitinen OH, Knip M, Hyoty H, Hytonen VP
  TITLE     Assessment of Enterovirus Antibodies during Early Childhood Using a 
            Multiplex Immunoassay.
  JOURNAL   Microbiol Spectr. 2023 Jun 15;11(3):e0535222. doi: 
            10.1128/spectrum.05352-22. Epub 2023 May 25.
  PUBMED    37227147
REFERENCE   2  (bases 1 to 5847)
  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"; journalName: "Microbiol 
            Spectr."; date: "2023-06-15"; volume: "11(3):e0535222. doi"; pages: 
            " 10.1128/spectrum.05352-22. Epub 2023 May 25"
COMMENT     Sequence Label: pGEX-2T_RV-B14_VP1
FEATURES             Location/Qualifiers
     source          1..5847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(182..187,188..203,204..210)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    218..234
                     /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)."
     CDS             257..910
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     868..890
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             917..934
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1808..1825
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1896..1918)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2063..2081)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2149..2253
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2254..2322,2323..3114)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2472..2491)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3285..3873
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3774..3793
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4027..4044
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4117..4194
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4195..5277
                     /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     complement(4214..4233)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5326..5331,5332..5349,5350..5356)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5364..5380
                     /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)."
     primer_bind     5388..5404
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5388..5404
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5400..5847,1..80)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5420..5437)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5420..5436)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5429..5451)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gcttatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg
       61 tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt
      121 ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct
      181 gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata acaatttcac
      241 acaggaaaca gtattcatgt cccctatact aggttattgg aaaattaagg gccttgtgca
      301 acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg
      361 cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct
      421 tccttattat attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat
      481 agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct
      541 tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt
      601 tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga
      661 tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt
      721 gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa
      781 attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc
      841 cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca
      901 tcctccaaaa tcggatctgg ttccgcgtgg atccggctta ggtgatgaat tagaagaagt
      961 catcgttgag aaaacgaaac agacggtggc ctcaatctca tctggtccaa aacacacaca
     1021 aaaagtcccc atactaactg caaacgaaac aggggccaca atgcctgttc ttccatcaga
     1081 cagcatagaa accagaacta cctacatgca ctttaatggt tcagaaactg atgtagaatg
     1141 ctttttgggt cgtgcagctt gtgtgcatgt aactgaaata caaaacaaag atgctactgg
     1201 aatagataat cacagagaag caaaattgtt caatgattgg aaaatcaacc tgtccagcct
     1261 tgtccaactt agaaagaaac tagaactctt cacttatgtt aggtttgatt ctgagtatac
     1321 catactggcc actgcatctc aacctgattc agcaaactat tcaagcaatt tggtggtcca
     1381 agccatgtat gttccacctg gtgccccgaa tccaaaagag tgggacgatt acacatggca
     1441 aagtgcttca aaccccagtg tattcttcaa ggtgggggat acatccaggt ttagtgtgcc
     1501 ttatgtagga ttggcatcag catataattg tttttatgat ggttactcac atgatgatgc
     1561 agaaactcag tatggcataa ctgttctaaa ccatatgggt agtatggcat tcagaatagt
     1621 aaatgaacat gatgaacata aaactcttgt caagatcaga gtttatcaca gggcaaagca
     1681 cgttgaagca tggattccaa gagcacccag agcactaccc tacacatcaa tagggcgcac
     1741 aaattatcct aagaatacag aaccagtaat taagaagagg aaaggtgaca ttaaatccta
     1801 tcccgggcac catcatcacc accatcccgg gtaataggaa ttcatcgtga ctgactgacg
     1861 atctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag
     1921 acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca
     1981 gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga tagcggagtg
     2041 tataattctt gaagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     2101 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     2161 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     2221 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     2281 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     2341 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     2401 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     2461 tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg acgccgggca agagcaactc
     2521 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     2581 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     2641 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     2701 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     2761 gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac aacgttgcgc
     2821 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     2881 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     2941 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     3001 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     3061 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     3121 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     3181 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     3241 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     3301 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     3361 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     3421 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     3481 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     3541 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     3601 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     3661 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     3721 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     3781 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     3841 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     3901 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     3961 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     4021 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt
     4081 acgcatctgt gcggtatttc acaccgcata aattccgaca ccatcgaatg gcgcaaaacc
     4141 tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt gaatgtgaaa
     4201 ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac cgtttcccgc
     4261 gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg
     4321 gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg
     4381 ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg
     4441 attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc
     4501 ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg
     4561 atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc ctgcactaat
     4621 gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat tattttctcc
     4681 catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca ccagcaaatc
     4741 gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc tggctggcat
     4801 aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga ctggagtgcc
     4861 atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg
     4921 ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg
     4981 cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag ctcatgttat
     5041 atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac cagcgtggac
     5101 cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt gcccgtctca
     5161 ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     5221 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
     5281 caacgcaatt aatgtaagtt agctcactca ttaggcaccc caggctttac actttatgct
     5341 tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta
     5401 tgaccatgat tacggattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     5461 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     5521 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgct
     5581 ttgcctggtt tccggcacca gaagcggtgc cggaaagctg gctggagtgc gatcttcctg
     5641 aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca cggttacgat gcgcccatct
     5701 acaccaacgt gacctatccc attacggtca atccgccgtt tgttcccacg gagaatccga
     5761 cgggttgtta ctcgctcaca tttaatgttg atgaaagctg gctacaggaa ggccagacgc
     5821 gaattatttt tgatggcgtt ggaatta
//
