LOCUS       sequence-329046-        5894 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial codon optimized expression vector for SARS-CoV-2 3CL 
            protease..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5894)
  AUTHORS   Iketani S, Forouhar F, Liu H, Hong SJ, Lin FY, Nair MS, Zask A, 
            Huang Y, Xing L, Stockwell BR, Chavez A, Ho DD
  TITLE     Lead compounds for the development of SARS-CoV-2 3CL protease 
            inhibitors.
  JOURNAL   Nat Commun. 2021 Apr 1;12(1):2016. doi: 10.1038/s41467-021-22362-2.
  PUBMED    33795671
REFERENCE   2  (bases 1 to 5894)
  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: "Nat 
            Commun."; date: "2021-04-1"; volume: "12(1):2016. doi"; pages: " 
            10.1038/s41467-021-22362-2"
COMMENT     Sequence Label: pGEX-5X-3-SARS-CoV-2-3CL
FEATURES             Location/Qualifiers
     source          1..5894
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             921..932
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1944..1966)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2111..2129)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2197..2301
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2302..2370,2371..3162)
                     /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(2520..2539)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3333..3921
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3822..3841
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4075..4092
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4165..4242
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4243..5325
                     /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(4262..4281)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5374..5379,5380..5397,5398..5404)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5412..5428
                     /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     5436..5452
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5436..5452
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5448..5894,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5468..5485)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5468..5484)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5477..5499)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg atcgaaggtc gttcagggtt ccgcaagatg gcgtttccgt
      961 cgggaaaggt ggaaggctgc atggtccagg tgacctgcgg cactacgacg ttgaatggcc
     1021 tgtggttaga cgatgtcgta tattgcccgc gtcacgtcat ttgtacgtca gaggatatgc
     1081 tgaatcccaa ctacgaggac ttgttaatcc gcaaaagtaa ccacaacttt ctggtgcaag
     1141 cagggaacgt acagctgcgc gtgatcgggc actcgatgca gaactgcgtt ctcaaactga
     1201 aagtagacac tgcaaacccc aaaacgccga aatacaaatt cgtccgtatc cagccgggcc
     1261 aaaccttctc cgttttggcg tgctataacg ggagcccctc gggcgtgtat cagtgcgcga
     1321 tgcgcccgaa ctttacaatc aaagggtcgt tcttgaacgg cagttgcgga agcgtgggct
     1381 tcaatatcga ctacgattgc gttagtttct gctatatgca tcacatggag ctcccgaccg
     1441 gcgtgcacgc cggtactgat cttgagggga atttctacgg gccattcgta gatcgtcaga
     1501 ccgcgcaggc ggcagggacc gataccacca tcaccgtgaa cgtactcgcg tggctgtatg
     1561 cagcagtgat caacggtgat cgctggttcc tgaaccgctt cacaacgacc ctcaacgatt
     1621 tcaatttagt cgccatgaaa tataactacg agcccttgac gcaggatcac gtggatatcc
     1681 tcggcccgct gagtgcgcag acggggatcg cggtgctgga catgtgcgcg agcctgaagg
     1741 agcttttaca gaacggcatg aacggtcgca ctatcttagg cagcgcactg ctggaagacg
     1801 agttcacccc gttcgacgta gtacgccagt gtagcggagt aacctttcag tgaggatccc
     1861 caggaattcc cgggtcgact cgagcggccg catcgtgact gactgacgat ctgcctcgcg
     1921 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     1981 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     2041 gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta taattcttga
     2101 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
     2161 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     2221 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     2281 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     2341 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     2401 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     2461 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     2521 ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata
     2581 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     2641 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     2701 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     2761 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     2821 gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact
     2881 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     2941 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     3001 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     3061 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     3121 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     3181 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     3241 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     3301 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     3361 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3421 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     3481 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3541 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3601 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3661 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3721 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3781 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     3841 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3901 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3961 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     4021 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     4081 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     4141 ggtatttcac accgcataaa ttccgacacc atcgaatggc gcaaaacctt tcgcggtatg
     4201 gcatgatagc gcccggaaga gagtcaattc agggtggtga atgtgaaacc agtaacgtta
     4261 tacgatgtcg cagagtatgc cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag
     4321 gccagccacg tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc ggagctgaat
     4381 tacattccca accgcgtggc acaacaactg gcgggcaaac agtcgttgct gattggcgtt
     4441 gccacctcca gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc
     4501 gccgatcaac tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc
     4561 tgtaaagcgg cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat cattaactat
     4621 ccgctggatg accaggatgc cattgctgtg gaagctgcct gcactaatgt tccggcgtta
     4681 tttcttgatg tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt
     4741 acgcgactgg gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg
     4801 ggcccattaa gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact
     4861 cgcaatcaaa ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt
     4921 caacaaacca tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac
     4981 gatcagatgg cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg
     5041 gatatctcgg tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtta
     5101 accaccatca aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa
     5161 ctctctcagg gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga
     5221 aaaaccaccc tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     5281 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     5341 tgtaagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     5401 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta
     5461 cggattcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     5521 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     5581 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcttt gcctggtttc
     5641 cggcaccaga agcggtgccg gaaagctggc tggagtgcga tcttcctgag gccgatactg
     5701 tcgtcgtccc ctcaaactgg cagatgcacg gttacgatgc gcccatctac accaacgtga
     5761 cctatcccat tacggtcaat ccgccgtttg ttcccacgga gaatccgacg ggttgttact
     5821 cgctcacatt taatgttgat gaaagctggc tacaggaagg ccagacgcga attatttttg
     5881 atggcgttgg aatt
//
