LOCUS       sequence-419269-        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: ZIKV NS2B-NS3 protease (H115A- catalytically
            i...
FEATURES             Location/Qualifiers
     source          1..6153
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     150..169
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     253..272
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        253..271
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    272..296
                     /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             311..333
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             344..361
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(377..400,401..436,437..460)
                     /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             485..505
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1270..1287
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1340..1358)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1354..1401
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1438..1893
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1525..1544)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1735..1756
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1985..2800)
                     /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(1985..2800)
                     /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     2708..2727
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2792..2811
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2922..3510
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3411..3430
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3664..3681
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3696..3835
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3761..3780)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3921..3943
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3937..4128)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4937..6019)
                     /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     5981..6000
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6020..6097)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
       61 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
      121 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
      181 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatctcg
      241 atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac aattcccctc
      301 tagaaataat tttgtttaac tttaagaagg agatatacat atgcaccatc atcatcatca
      361 ttcttctggt gcgagctgga gtcacccaca attcgagaaa ggtggtggct ctggtggcgg
      421 ttcaggcggc tccgcctggt cccatccaca gtttgaaaag ggttctggtg tagatctggg
      481 taccgagaac ctgtacttcc aatccatgag cggtaagtct gtcgatatgt acattgaacg
      541 tgccggtgac atcacctggg agaaggatgc agaagtgacg ggtaactcgc caagacttga
      601 tgttgcactc gacgagtcgg gcgattttag cttagttgaa gatgacggtc cgcctatgcg
      661 agagggcggc ggcggtagcg gcggtggtgg tggtagcgga gcattatggg atgtgccggc
      721 acccaaggaa gtaaagaaag gggagacgac agacggtgta tacagagtta tgacgagaag
      781 actgttgggt agcacacaag ttggtgttgg cgtaatgcag gaaggagtct tccacaccat
      841 gtgggcggtc accaagggca gtgcgcttcg cagcggtgag ggacgtctgg atccgtactg
      901 gggagacgtg aagcaagacc tggtcagtta ttgtggtccg tggaagcttg acgcagcctg
      961 ggacggccat agcgaggtgc aactcctcgc cgtaccgccg ggagagcgcg cccgtaatat
     1021 tcaaactctg cccggtatct ttaaaaccaa ggacggagat ataggcgcag tggctcttga
     1081 ttatcccgca gggactagcg gtagtcctat tctggataag tgcggccgag tgattggctt
     1141 atacggaaat ggagtggtta tcaagaatgg aagctatgtc tcagccataa cccaaggtcg
     1201 tcgcgaagag gagacacccg tagaataatg actcgagcac ccgtcgacaa gcttgcggcc
     1261 gcactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa
     1321 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     1381 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg
     1441 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     1501 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     1561 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     1621 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     1681 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     1741 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     1801 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     1861 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     1921 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgaatta
     1981 attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     2041 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     2101 tccataggat ggcaagatcc tggtatcggt ctgctattcc gactcgtcca acatcaatac
     2161 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     2221 cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag
     2281 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     2341 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     2401 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     2461 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
     2521 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     2581 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     2641 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     2701 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     2761 gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
     2821 ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga gttttcgttc
     2881 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     2941 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     3001 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     3061 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     3121 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     3181 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     3241 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     3301 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     3361 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     3421 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     3481 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     3541 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     3601 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     3661 cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg
     3721 catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg
     3781 catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg
     3841 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     3901 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     3961 gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat
     4021 gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct
     4081 tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt
     4141 gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac
     4201 gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact
     4261 ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt
     4321 taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa
     4381 cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa
     4441 gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg
     4501 ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt
     4561 cctcaacgac aggagcacga tcatgcgcac ccgtggggcc gccatgccgg cgataatggc
     4621 ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg
     4681 caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagtggtc
     4741 ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa
     4801 gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg
     4861 gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac
     4921 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4981 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt
     5041 ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga
     5101 gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg
     5161 ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat
     5221 ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat
     5281 cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt
     5341 gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc
     5401 gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc
     5461 ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg
     5521 taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa
     5581 ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg
     5641 gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac
     5701 aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg
     5761 cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg
     5821 caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt
     5881 aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg
     5941 cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt
     6001 ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg
     6061 ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta
     6121 tgcgactcct gcattaggaa gcagcccagt agt
//
