LOCUS       sequence-420358-        4493 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes EV-D68 MO/14/18947 3ABC with active site mutation optimized 
            for expression in mammalian transcription/translation extracts.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4493)
  TITLE     Differential Inhibition of Intra- and Inter-molecular Protease 
            Cleavages by Antiviral Compounds
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4493)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pT7CFE1_EVD68_3ABC_C147R
FEATURES             Location/Qualifiers
     source          1..4493
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(789..836)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     832..850
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(906..923)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1841..1867)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             complement(1841..1867)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    1875..2336
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     2263..2280
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     complement(2386..2405)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2387..2405)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2412..2429)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2412..2428)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2421..2443)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2569..3024
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2656..2675)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2866..2887
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3093..3112)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3212..3234
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3272..3290)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3358..3462
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3463..3531,3532..4323)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3681..3700)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcccgga tatagttcct
      781 cctttcagca aaaaacccct caagacccgt ttagaggccc caaggggtta tgctagttat
      841 tgctcagcgg actagtgttt aaactttttt tttttttttt tttttttttt ttcagtcaga
      901 tctcagtggt ggtggtggtg gtgctcgagt gtgtatcgga aaagtaagag tgtagtagca
      961 ttgctgcaaa tccttgagct ccatttccac ctacatgtat tcctatcacc ttacctgtag
     1021 ttgtcaccac accaccacgc tggccagctc ttgttgggaa gttatacatt aaaatgcggt
     1081 gcgtcggtgt accacctagg ttcaagaagc cataattggt gacttgtcca actgggatat
     1141 acatatttgg gaattttgat gtatgtacgc taagcacagc gtcattgtaa tcatcctcat
     1201 atctgggcag aaaatgtctg atatctctaa atttttgatt acggtctaat ttgactatgg
     1261 ttatctctaa gtttgtatca gtcaagtctc taagtgcaca cgcatctaaa actttagtct
     1321 ctacatcatt aatgtaaatg gtttctccaa cagatgcgtg tgtggggatg accgctaccc
     1381 tatcatatac acccagcatg gtgaactcac ccttttcagt ccttgcaatg acggtatttt
     1441 tcttcattat ggcttgtgca aaatcgaacc ctggtccttg cactttagct gttctgagag
     1501 agggtacttt aggtttagga ttggggattc ctgtgtatgg accctgaatg ccagcaaaaa
     1561 gtttgtatat aacataaact acaccagcta ttgatacaaa ggtggcaatg gcttgtagag
     1621 taataaaagc tctactaatg tgtttttcta ctattagctc atttgatggg tgtactacaa
     1681 tccatccttt cttttggcaa taatccctaa cttcttgaga atccactgac ctaagaaggt
     1741 catttatagc atcaggagct ggtgtatctg gggtgactga aattttgatc tctttaaact
     1801 gtggaggccc gaattcggat cccatatggg tggtggccag ggcgtagtcg ggcacgtcgt
     1861 aggggtaggc catattatca tcgtgttttt caaaggaaaa ccacgtcccc gtggttcggg
     1921 gggcctagac gttttttaac ctcgactaaa cacatgtaaa gcatgtgcac cgaggcccca
     1981 gatcagatcc catacaatgg ggtaccttct gggcatcctt cagccccttg ttgaatacgc
     2041 ttgaggtgag ccatttgact ctttccacaa ctatccaact cacaacgtgg cactggggtt
     2101 gtgccgcctt tgcaggtgta tcttatacac gtggcttttg gccgcagagg cacctgtcgc
     2161 caggtggggg gttccgctgc ctgcaaaggg tcgctacaga cgttgtttgt cttcaagaag
     2221 cttccagagg aactgcttcc ttcacgacat tcaacagacc ttgcattcct ttggcgagag
     2281 gggaaagacc cctaggaatg ctcgtcaaga agacagggcc aggtttccgg gccctcacat
     2341 tgccaaaaga cggcaatatg gtggaaaata accggaatta attcgcccta tagtgagtcg
     2401 tattacaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     2461 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     2521 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggac gcgccctgta
     2581 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     2641 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     2701 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     2761 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     2821 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     2881 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     2941 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     3001 acaaaatatt aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg
     3061 tatttcacac cgcatatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3121 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     3181 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     3241 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     3301 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     3361 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3421 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     3481 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     3541 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     3601 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     3661 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     3721 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     3781 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     3841 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     3901 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     3961 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     4021 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     4081 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     4141 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     4201 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     4261 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     4321 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     4381 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4441 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
