LOCUS       sequence-461605-        7290 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of the Zika virus NS3 helicase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7290)
  TITLE     ASAP Discovery Plasmids 2024
REFERENCE   2  (bases 1 to 7290)
  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: ZIKV isolate PRVABC59 NS3 helicase domain
FEATURES             Location/Qualifiers
     source          1..7290
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     52..71
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        52..70
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    71..95
                     /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             110..132
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             143..160
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             167..820
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             836..856
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2206..2223
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2276..2294)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2290..2337
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2374..2829
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2461..2480)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2671..2692
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2921..3736)
                     /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(2921..3736)
                     /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     3644..3663
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3728..3747
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3858..4446
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4347..4366
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4600..4617
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4632..4771
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4697..4716)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4857..4879
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4873..5064)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5873..6955)
                     /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     6917..6936
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6956..7033)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7239..7258
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ccggccacga tgcgtccggc gtagaggatc gagatctcga tcccgcgaaa ttaatacgac
       61 tcactatagg ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact
      121 ttaagaagga gatatacata tgcaccatca ccaccatcac tcatcgatgt ctcccatttt
      181 gggttattgg aagataaagg ggttagtaca accgaccaga ttactgttag agtaccttga
      241 agagaagtat gaagagcatt tatatgagcg cgacgaaggc gataagtggc gcaacaagaa
      301 atttgagtta ggcctcgagt ttccgaacct tccttattat atcgacgggg acgtaaagtt
      361 gacgcaatct atggcaataa ttcggtatat agcagacaag cataacatgt tgggtggctg
      421 ccccaaagag cgtgccgaga tctccatgtt agaaggagcc gtcctggaca ttcgctatgg
      481 tgtgtcgcgc attgcctact caaaagattt cgagacttta aaagttgact ttctgagcaa
      541 gctgccggag atgcttaaga tgttcgaaga tcggctgtgt cacaagacgt acctcaacgg
      601 tgaccacgtt acccacccag actttatgct ttacgatgcg ctggacgttg tgctttacat
      661 ggacccgatg tgtctggacg cgtttccgaa gttggtatgt ttcaagaagc ggattgaggc
      721 tattccccaa atagacaaat accttaagag tagtaagtac atcgcgtggc cacttcaagg
      781 ttggcaagct accttcggcg gcggcgacca tcccccaaag tcgggcggcg gcagcgagaa
      841 cctgtacttc caatccatgc tcaagaagaa acaattgacc gtgcttgact tgcaccctgg
      901 cgcgggcaag actcgccgtg tgctgccgga gattgtaaga gaggctatca agaaacgcct
      961 gcgtaccgta attctcgctc cgacgcgtgt ggtagccgca gaaatggaag aagcgctgcg
     1021 cggcctgcct gtacgatata tgactactgc tgttaatgtg actcacagtg gtacggagat
     1081 tgtggactta atgtgtcacg cgacgtttac gagtcgtctg ttgcagccga ttcgcgttcc
     1141 aaactataac ctgaatatta tggacgaggc tcattttacc gatcctagtt ctattgcagc
     1201 tcgcggttat atcagcacta gagtggagat gggagaagcc gccgctatat tcatgacagc
     1261 aactcccccg ggtacacgcg acgccttccc cgatagcaat tctcctataa tggatactga
     1321 ggtagaggtt cccgaacgcg cgtggtcgag tgggttcgac tgggtaaccg atcactccgg
     1381 aaagaccgtg tggtttgtcc cgagtgttcg gaatggtaac gagattgccg cgtgcttaac
     1441 caaagcgggt aaaagagtaa ttcaattgtc tcgaaagacg ttcgaaaccg agttccaaaa
     1501 gaccaagaac caggaatggg atttcgttat cactacggat ataagcgaaa tgggagcaaa
     1561 ttttaaagcg gatcgtgtaa ttgattcacg ccgctgtctg aagcctgtga ttttggacgg
     1621 cgaacgcgta attctggccg gtccaatgcc ggtaacccac gcatctgcgg cacaacgtcg
     1681 cggtcgaatt gggcgcaatc cgaataagcc aggtgacgaa tatatgtacg gtggcggctg
     1741 cgctgaaacc gacgagggtc acgcccattg gttagaggcc cgtatgttac tcgataatat
     1801 ttatctgcaa gacgggctga ttgcaagcct gtaccgcccc gaagcggaca aagtcgcagc
     1861 tatcgaaggc gaattcaaac ttcgtacgga acagcgtaaa acatttgtcg agttaatgaa
     1921 acgtggcgat ctgccggttt ggctcgcgta ccaagtagcg tccgcgggta tcacgtatac
     1981 cgatcgtcga tggtgtttcg acggtacgac aaataataca atcatggaag attcagttcc
     2041 ggctgaagta tggactaaat acggtgagaa acgggttctt aagcctcgct ggatggacgc
     2101 gcgagtgtgc tcggatcacg ccgcgttgaa atcattcaag gagtttgcag caggtaagcg
     2161 ataatgactc gagcacccgt cgacaagctt gcggccgcac tcgagcacca ccaccaccac
     2221 cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct
     2281 gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg
     2341 aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     2401 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     2461 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     2521 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     2581 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     2641 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     2701 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     2761 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     2821 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     2881 ctaaatacat tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca
     2941 tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc
     3001 gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt
     3061 atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa
     3121 aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca
     3181 aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa
     3241 aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata
     3301 cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca
     3361 ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg
     3421 ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat
     3481 gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg
     3541 taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct
     3601 tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat
     3661 acccatataa atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc
     3721 gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg
     3781 ttcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     3841 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     3901 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     3961 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     4021 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     4081 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     4141 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     4201 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     4261 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     4321 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     4381 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     4441 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     4501 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     4561 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     4621 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     4681 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc
     4741 gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc
     4801 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     4861 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta
     4921 aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag
     4981 ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag
     5041 ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg
     5101 ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca
     5161 tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca
     5221 gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca
     5281 gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt
     5341 ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt
     5401 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
     5461 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
     5521 gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt
     5581 ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga
     5641 caggccgatc atcgtcgcgc tccagcgaaa gtggtcctcg ccgaaaatga cccagagcgc
     5701 tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat
     5761 agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg
     5821 tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     5881 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5941 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     6001 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     6061 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     6121 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     6181 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     6241 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     6301 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     6361 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     6421 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     6481 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     6541 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     6601 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     6661 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     6721 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     6781 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     6841 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     6901 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     6961 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     7021 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag
     7081 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     7141 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     7201 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
     7261 ccagcaaccg cacctgtggc gccggtgatg
//
