LOCUS       sequence-464108-        6501 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of the MERS-CoV Mpro domain for biochemical 
            assay..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6501)
  TITLE     ASAP Discovery Plasmids 2024
REFERENCE   2  (bases 1 to 6501)
  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: MERS-CoV Mpro strain HCoV-EMC 3CL protease domain
FEATURES             Location/Qualifiers
     source          1..6501
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    39..63
                     /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             78..100
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             111..128
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1385..1402
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1455..1473)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1469..1516
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1553..2008
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1640..1659)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1850..1871
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2100..2915)
                     /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(2100..2915)
                     /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     2823..2842
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2907..2926
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3037..3625
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3526..3545
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3779..3796
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3811..3950
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3876..3895)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4036..4058
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4052..4243)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5052..6134)
                     /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     6096..6115
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6135..6212)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6418..6437
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat
       61 tcccctctag aaataatttt gtttaacttt aagaaggaga tatacatatg caccaccatc
      121 atcaccatgg ctcaggagat caggaagcga aaccttcgac cgaggacctg ggcgacaaga
      181 aagaagggga gtatattaag ttaaaggtga tcggccagga ctcaagcgag atccacttca
      241 aagtcaaaat gaccacgcac ctcaagaagt tgaaggagag ttactgtcaa cggcaagggg
      301 ttcctatgaa ctcgcttcgc ttcctgttcg agggccagcg aattgcggac aaccacacac
      361 ctaaggaact tggtatggaa gaggaagact tcattgaggt gtaccaggag cagacaggcg
      421 gctctgggct cgttaagatg tcccatccta gcggagacgt cgaggcgtgt atggtacaag
      481 ttacgtgcgg gagcatgacg ttaaacggat tgtggctcga caatacagtt tggtgcccac
      541 gacatgtcat gtgtccggct gatcaattaa gtgacccaaa ttacgacgcg ttgctcatat
      601 ccatgacgaa ccacagcttt agcgtccaaa agcacattgg tgcccctgct aaccttcggg
      661 tcgttggcca tgctatgcag ggcactcttt taaagctgac agttgacgtt gctaacccta
      721 gtacgcctgc ctatacattc actacggtta aacccggcgc cgcgttctcc gttttggcat
      781 gttacaacgg tcgtccgaca ggtacgttca cagtagttat gcggccaaat tataccatca
      841 aaggtagctt cctttgcggt tcatgtggta gtgtgggcta caccaaagag ggctcggtga
      901 taaatttctg ttacatgcac caaatggaac ttgcaaacgg gacccacacc ggtagtgcgt
      961 ttgatggtac catgtatgga gctttcatgg acaagcaagt tcaccaagtg caactgaccg
     1021 ataagtactg ctctgttaat gtcgtggctt ggctgtatgc cgccatcttg aacggctgcg
     1081 cctggtttgt caagccaaat cgcacgtccg ttgtaagctt caacgagtgg gccctggcaa
     1141 accaattcac tgagttcgtt gggacccagt ctgtcgacat gttggccgtt aagacgggtg
     1201 tagcgataga gcagctgctt tacgcaatcc aacaattata cacaggtttc cagggaaagc
     1261 agattctcgg ttctacaatg ttagaggacg aattcacgcc cgaggacgtg aacatgcaga
     1321 tcatgggcgt tgtcatgcaa taatgactcg agcacccgtc gacaagcttg cggccgcact
     1381 cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa aggaagctga
     1441 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     1501 cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg ggacgcgccc
     1561 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     1621 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     1681 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     1741 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     1801 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     1861 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     1921 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     1981 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     2041 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct
     2101 tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata
     2161 ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat
     2221 aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct
     2281 attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact
     2341 gaatccggtg agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag
     2401 ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc
     2461 gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa
     2521 tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat
     2581 tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca
     2641 tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt
     2701 agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac
     2761 aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca
     2821 ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc
     2881 ctagagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg
     2941 taagcagaca gttttattgt tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3001 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3061 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     3121 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     3181 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     3241 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3301 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3361 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3421 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3481 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3541 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3601 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3661 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     3721 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     3781 cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct
     3841 gtgcggtatt tcacaccgca atggtgcact ctcagtacaa tctgctctga tgccgcatag
     3901 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
     3961 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     4021 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     4081 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg
     4141 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
     4201 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag
     4261 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
     4321 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
     4381 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata
     4441 cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa
     4501 tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca
     4561 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc
     4621 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca
     4681 acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata atggcctgct
     4741 tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga
     4801 ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag tggtcctcgc
     4861 cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag
     4921 tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga
     4981 aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa
     5041 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     5101 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     5161 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     5221 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     5281 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca
     5341 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     5401 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     5461 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     5521 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     5581 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     5641 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     5701 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     5761 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     5821 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     5881 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     5941 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     6001 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     6061 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     6121 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     6181 ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga
     6241 ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag
     6301 gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
     6361 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
     6421 gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat
     6481 gcgtccggcg tagaggatcg a
//
