LOCUS       sequence-351687-        7143 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the N-terminal His6-tagged SARS-CoV helicase in E coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7143)
  AUTHORS   Tanner JA, Watt RM, Chai YB, Lu LY, Lin MC, Peiris JS, Poon LL, Kung
            HF, Huang JD
  TITLE     The severe acute respiratory syndrome (SARS) coronavirus 
            NTPase/helicase belongs to a distinct class of 5' to 3' viral 
            helicases.
  JOURNAL   J Biol Chem. 2003 Oct 10;278(41):39578-82. doi: 
            10.1074/jbc.C300328200. Epub 2003 Aug 13.
  PUBMED    12917423
REFERENCE   2  (bases 1 to 7143)
  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"; doi: 
            "10.1074/jbc.C300328200"; journalName: "J Biol Chem"; date: 
            "2003-10-10- 10"; volume: "278"; issue: "41"; pages: "39578-82"
COMMENT     Sequence Label: pHelA12
FEATURES             Location/Qualifiers
     source          1..7143
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(36..83)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     79..97
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(150..167)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1995..2027)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             complement(2031..2048)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(2058..2075)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2094..2116
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2131..2155
                     /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     complement(2155..2174)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2156..2174)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2258..2277)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2483..2560
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2561..3643
                     /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(2580..2599)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4452..4643
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4637..4659)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4745..4884
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4800..4819
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4899..4916)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5070..5658)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5150..5169)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5769..5788)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5780..6595
                     /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             5780..6595
                     /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     complement(5853..5872)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(6687..7142)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6824..6845)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7036..7055
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ccattcgcca atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta
       61 gaggccccaa ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc
      121 tttcgggctt tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtcattgt
      181 aatgtagcca cattgcgacg tggtatttct agacttgtaa attgcagttt gtcataaaga
      241 tctctatcag acattatgca caaaatgcca atttttgccc ttgtgatagc cacattgaag
      301 cggttgacat tacaagagtg tgctgtttca gtagtttgtg tgaatatgac atagtcatat
      361 tcagaaccct gtgatgaatc aacagtctgc gtaggcaatc ctaagatttt tgaagctacg
      421 gcgttctgtg aattataagg tgagataaaa acagcttttc tccaagcagg attgcgtgta
      481 agaaattctc ttacaacgcc tatttgaggt ctgttgattg cagatgaaac atcatgtgta
      541 ataacacctt tgtagaacat tttgaagcat tgagctgact tatccttgtg tgcttttagc
      601 ttattgtcat aaactaaagc actcacagtg tcaacaattt cagcaggaca acggcgacaa
      661 gttccaagga acatgtctgg acctattgtt ttcataagtc tgcacactga attaaaatat
      721 tctggttcta gtgtgccttt agtcagcaat gtgcgggggg ctggtaattg agcaggatcg
      781 ccaatataga cgtagtgttt tgcacgaagt ctagcattga caacactcaa gtcataatta
      841 gtagccatag agatttcatc aaagactaca atgtcagcag ttgtttctgg caatgcattt
      901 acagtgcaga aaacatactg ttctagtgtt gaattcactt tgaatttatc aaaacactct
      961 acgcgcgcac gcgcaggtat gattctacta catttatcta tgggcaaata ttttaatgcc
     1021 ttttcacata gggcatcaac agctgcatga gagcatgccg tatacactat gcgagcagat
     1081 gggtaataga gagcaagtcc gatggcaaaa tgactcttac cagtaccagg tggtccttgg
     1141 agtgtagagt acttttgcat gccgaccttt tgataatttg caacattgct agaaaactca
     1201 tctgagatgt tgagtgttgg gtacaagcca gtaattctca catagtgctc ttgtggcact
     1261 agagtaggtg cactaagtgg cattacagtg tgagatgtca acacaaagta atcaccaaca
     1321 ttcaacttgt atgtcgtagt acctctgtac acaacagcat caccatagtc acctttttca
     1381 aaggtgtact ctccaatctg tactttacta tttttagtta cacggtaacc agtaaagaca
     1441 tagtttctgt tcaatggtgg tctaggtttt ccaacctccc atgaaagatg caattctctg
     1501 tcagagagta cttcgcgtac agtggcaata ccatatgaca gcttaaatgt ttcctcagtg
     1561 gctttgagcg tttctgctgc gaaaagcttg agtctctcag tacaagtgtt ggcaagtatg
     1621 taatcgccag cattagtcca atcacatgtt gctatcgcat tgaagtcagt gacattgtca
     1681 ctgcctacac atgtgttttt gtataaacca aaaacctgac cattagcaca taatggaaaa
     1741 ctaatgggag gcttatgtga cttgcaataa tagctcatac ctcctagata cagttgtgtc
     1801 acatcagtga catcacaacc cggggcattg caaacatagg gattaacaga caacactaat
     1861 ttgtgtgatg ttgaaatgac atggtcatag cagcacttgc aacataggaa tggtctccta
     1921 atacaggcac cgcaacgaag tgaagtctgt gaattgcaca atacacaagc acctacagcc
     1981 atggcggatc cgcgacccat ttgctgtcca ccagtcatgc tagccatatg gctgccgcgc
     2041 ggcaccaggc cgctgctgtg atgatgatga tgatggctgc tgcccatggt atatctcctt
     2101 cttaaagtta aacaaaatta tttctagagg ggaattgtta tccgctcaca attcccctat
     2161 agtgagtcgt attaatttcg cgggatcgag atctcgatcc tctacgccgg acgcatcgtg
     2221 gccggcatca ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat
     2281 ggggaagatc gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg
     2341 gcaggccccg tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg
     2401 gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat
     2461 aagggagagc gtcgagatcc cggacaccat cgaatggcgc aaaacctttc gcggtatggc
     2521 atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata
     2581 cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc
     2641 cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta
     2701 cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc
     2761 cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc
     2821 cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg
     2881 taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc
     2941 gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt
     3001 tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac
     3061 gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg
     3121 cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg
     3181 caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca
     3241 acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga
     3301 tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga
     3361 tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac
     3421 caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact
     3481 ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa
     3541 aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     3601 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     3661 taagttagct cactcattag gcaccgggat ctcgaccgat gcccttgaga gccttcaacc
     3721 cagtcagctc cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct
     3781 tctttatcat gcaactcgta ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg
     3841 accgctttcg ctggagcgcg acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc
     3901 acgccctcgc tcaagccttc gtcactggtc ccgccaccaa acgtttcggc gagaagcagg
     3961 ccattatcgc cggcatggcg gccccacggg tgcgcatgat cgtgctcctg tcgttgagga
     4021 cccggctagg ctggcggggt tgccttactg gttagcagaa tgaatcaccg atacgcgagc
     4081 gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca tgaatggtct
     4141 tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc agcgccctgc accattatgt
     4201 tccggatctg catcgcagga tgctgctggc taccctgtgg aacacctaca tctgtattaa
     4261 cgaagcgctg gcattgaccc tgagtgattt ttctctggtc ccgccgcatc cataccgcca
     4321 gttgtttacc ctcacaacgt tccagtaacc gggcatgttc atcatcagta acccgtatcg
     4381 tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat cccccttaca
     4441 cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc tttatcagaa
     4501 gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa caggcagaca
     4561 tctgtgaatc gcttcacgac cacgctgatg agctttaccg cagctgcctc gcgcgtttcg
     4621 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     4681 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     4741 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     4801 ggcatcagag cagattgtac tgagagtgca ccattgcggt gtgaaatacc gcacagatgc
     4861 gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4921 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4981 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5041 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5101 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5161 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5221 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5281 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5341 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5401 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5461 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
     5521 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5581 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5641 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5701 aacgaaaact cacgttaagg gattttggtc atgaacaata aaactgtctg cttacataaa
     5761 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcttgct ctaggccgcg
     5821 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
     5881 gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag agttgtttct
     5941 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg
     6001 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
     6061 atggttactc accactgcga tccccgggaa aacagcattc caggtattag aagaatatcc
     6121 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
     6181 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc
     6241 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
     6301 tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg attcagtcgt
     6361 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
     6421 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     6481 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     6541 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaagaatt
     6601 aattcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     6661 gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc
     6721 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     6781 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     6841 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     6901 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     6961 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     7021 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     7081 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     7141 gtc
//
