LOCUS       sequence-419641-        6153 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of a fusion of Zika NS2B-NS3 proteins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6153)
  TITLE     The SARS-Cov-2 main protease (Mpro) is responsible for viral 
            polyprotein processing and therefore a therapeutic target.
REFERENCE   2  (bases 1 to 6153)
  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: Fusion of Zika NS2B-NS3 proteins
FEATURES             Location/Qualifiers
     source          1..6153
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     77..96
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     180..199
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        180..198
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    199..223
                     /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             238..260
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             271..288
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(304..327,328..363,364..387)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             412..432
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1197..1214
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1267..1285)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1281..1328
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1365..1820
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1452..1471)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1662..1683
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1912..2727)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1912..2727)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2635..2654
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2719..2738
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2849..3437
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3338..3357
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3591..3608
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3623..3762
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3688..3707)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3848..3870
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3864..4055)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4864..5946)
                     /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     5908..5927
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5947..6024)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
       61 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
      121 cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc ccgcgaaatt
      181 aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag aaataatttt
      241 gtttaacttt aagaaggaga tatacatatg caccatcatc atcatcattc ttctggtgcg
      301 agctggagtc acccacaatt cgagaaaggt ggtggctctg gtggcggttc aggcggctcc
      361 gcctggtccc atccacagtt tgaaaagggt tctggtgtag atctgggtac cgagaacctg
      421 tacttccaat ccatgagtgg aaagtctgtc gatatgtaca tcgagcgagc cggcgacatc
      481 acatgggaga aagacgcaga ggttaccggg aatagcccgc ggcttgacgt tgcgttggac
      541 gagagtggtg acttctctct ggtcgaggac gacggaccac ctatgcgtga gggaggaggt
      601 gggtcaggtg gtggtggtgg ctcgggtgct ctttgggatg tccctgcccc caaagaggtg
      661 aagaagggtg agacgaccga cggagtgtat cgcgttatga caagacgttt attagggtca
      721 acacaagtag gtgttggtgt aatgcaagag ggcgttttcc acacaatgtg gcacgttact
      781 aagggctcag ctcttcggtc aggtgaggga cgccttgacc cgtactgggg cgacgttaag
      841 caagatctgg tgtcttattg tggtccctgg aagttagacg ctgcatggga cggtcatagc
      901 gaggtccaat tgctggcagt gcccccgggt gagagagcgc gaaatattca aacgcttcct
      961 gggattttca agaccaagga cggtgacatc ggtgcagtgg cgctggatta cccggcggga
     1021 acgtccggca gtccgatttt ggataagtgc ggtcgggtaa tcggactgta tggtaatggg
     1081 gtggtgatta agaacggctc atacgtgtct gcaataaccc agggccgtcg agaggaagaa
     1141 actcctgttg agtaatgact cgagcacccg tcgacaagct tgcggccgca ctcgagcacc
     1201 accaccacca ccactgagat ccggctgcta acaaagcccg aaaggaagct gagttggctg
     1261 ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg
     1321 gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc cctgtagcgg
     1381 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     1441 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     1501 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     1561 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     1621 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     1681 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     1741 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     1801 aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     1861 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgaattaatt cttagaaaaa
     1921 ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa taccatattt
     1981 ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc ataggatggc
     2041 aagatcctgg tatcggtctg ctattccgac tcgtccaaca tcaatacaac ctattaattt
     2101 cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga ctgaatccgg
     2161 tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc agccattacg
     2221 ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt gcgcctgagc
     2281 gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg aatgcaaccg
     2341 gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat attcttctaa
     2401 tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat catcaggagt
     2461 acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt ttagtctgac
     2521 catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa acaactctgg
     2581 cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga cattatcgcg
     2641 agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg gcctagagca
     2701 agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta tgtaagcaga
     2761 cagttttatt gttcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     2821 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     2881 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     2941 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     3001 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     3061 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3121 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3181 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     3241 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     3301 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     3361 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     3421 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     3481 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     3541 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     3601 tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     3661 tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     3721 gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca
     3781 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     3841 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg
     3901 cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca
     3961 tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg
     4021 gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt
     4081 ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact
     4141 gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg
     4201 cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta
     4261 ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag
     4321 ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt
     4381 gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt
     4441 gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg
     4501 agcacgatca tgcgcacccg tggggccgcc atgccggcga taatggcctg cttctcgccg
     4561 aaacgtttgg tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa gattccgaat
     4621 accgcaagcg acaggccgat catcgtcgcg ctccagcgaa agtggtcctc gccgaaaatg
     4681 acccagagcg ctgccggcac ctgtcctacg agttgcatga taaagaagac agtcataagt
     4741 gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc
     4801 aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg
     4861 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     4921 caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag
     4981 tgagacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg
     5041 gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat
     5101 ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatatccg caccaacgcg
     5161 cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag
     5221 catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat
     5281 ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt
     5341 atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc
     5401 gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg
     5461 ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac
     5521 attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat
     5581 cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc
     5641 gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat
     5701 cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag
     5761 caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc
     5821 catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac
     5881 gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg
     5941 tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa
     6001 ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc gactcctgca
     6061 ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg
     6121 catgcaagga gatggcgccc aacagtcccc cgg
//
