LOCUS       sequence-420352-        4973 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes EV-D68 MO/14/18947 VP12A with telaprevir resistance 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 4973)
  TITLE     Differential Inhibition of Intra- and Inter-molecular Protease 
            Cleavages by Antiviral Compounds
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4973)
  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            "Unpublished"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_VP12A_N84T
FEATURES             Location/Qualifiers
     source          1..4973
                     /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(905..922)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2321..2347)
                     /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(2321..2347)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    2355..2816
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     2743..2760
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     complement(2866..2885)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2867..2885)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2892..2909)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2892..2908)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2901..2923)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3049..3504
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3136..3155)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3346..3367
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3573..3592)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3692..3714
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3752..3770)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3838..3942
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3943..4011,4012..4803)
                     /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(4161..4180)
                     /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 tcagtcagat
      901 ctcagtggtg gtggtggtgg tgctcgagtt gttccatagc atcagtatct aaccatagca
      961 aattcctgat atcagtgaaa gctacaattc caccccctcc tgctgtaaga agaccgatta
     1021 ccccatgtgg acaaactagt aaaccaccgc aatctcctgc ttccgcagga ccaactgcca
     1081 aaagtacatt ggtctggtac cttgctgggt aatattccgt ttgttcaatc cattgaatcc
     1141 ctgggccttc aaagcaaatc gggtaacttc tgtttttgtg cctacaatag taaacccctg
     1201 tgttgcactt gcatcttgct atttggtgcc ttccatgagc agcaatgggg gtaaccaaga
     1261 cgtctgattg ccaatccaca tagatagctg actgtctctc ttctgtagtg gccaagtgat
     1321 agttgattat tttaaaagac cctacaaaaa ctcctccgaa gcctggacca gtgttcacta
     1381 tattatgagg catggttttg acactgtctc tattgccaat tatagcactg agcgcatttg
     1441 gtgctctttg tttacctttg taatttgcat ttgcaatact catatatggc agagttcgtg
     1501 gtggtcgtgg tgcccatgct tttatgtgtt taggcttcat gtaaacccta acggtcactg
     1561 tgaaaccaac tggttggtgt tcattcacta ttctaacaca taagttacca atagtgtcag
     1621 ctggatttat tccatacaga ccgtttttct caaatccggc aaagccatca taaaaaactg
     1681 agtatgctga gttaatgcac ataaaaggta tggttattct ggctgggggg tcggagattt
     1741 taaagaatac actagcatta ctgcctgact gccagtggaa tgagtcctgc ttttctgggg
     1801 taagagcacc agtgggtaca aacattgctt gaagtgtcaa gtcaggaaga cccacgtatg
     1861 tattattacc actaccattc actgctacag ttgtgagtat agtgatctca gcatcaaatc
     1921 taaggtatgt gaataattct aattttcttc ttaactgtac aaaggatctg gtgttaattg
     1981 tccatttgaa aaagttcttg tctgctcgtg ctgtagacga agtatgatct ttgtattcaa
     2041 aacttctctt tgataccaaa gctgctctac tgagaaaatt ctccactaga gtctcggata
     2101 caccgtgctg atttatcact gtgcgagttt gtatggcttc ttctggttca gtgttagaag
     2161 ttacacctgt ttcaactgca tttaagctag ggaccacacc aagttcagca ttaatctcac
     2221 ttttcacagt gtcggtcgct gttttgatga tgctctcgat ctggtaggct gcctctgctg
     2281 catgtaaatg gtctagggat cccatatggg tggtggccag ggcgtagtcg ggcacgtcgt
     2341 aggggtaggc catattatca tcgtgttttt caaaggaaaa ccacgtcccc gtggttcggg
     2401 gggcctagac gttttttaac ctcgactaaa cacatgtaaa gcatgtgcac cgaggcccca
     2461 gatcagatcc catacaatgg ggtaccttct gggcatcctt cagccccttg ttgaatacgc
     2521 ttgaggtgag ccatttgact ctttccacaa ctatccaact cacaacgtgg cactggggtt
     2581 gtgccgcctt tgcaggtgta tcttatacac gtggcttttg gccgcagagg cacctgtcgc
     2641 caggtggggg gttccgctgc ctgcaaaggg tcgctacaga cgttgtttgt cttcaagaag
     2701 cttccagagg aactgcttcc ttcacgacat tcaacagacc ttgcattcct ttggcgagag
     2761 gggaaagacc cctaggaatg ctcgtcaaga agacagggcc aggtttccgg gccctcacat
     2821 tgccaaaaga cggcaatatg gtggaaaata accggaatta attcgcccta tagtgagtcg
     2881 tattacaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     2941 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     3001 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggac gcgccctgta
     3061 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     3121 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     3181 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     3241 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     3301 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     3361 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     3421 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     3481 acaaaatatt aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg
     3541 tatttcacac cgcatatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3601 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     3661 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     3721 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     3781 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     3841 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3901 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     3961 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4021 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4081 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4141 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4201 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     4261 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     4321 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     4381 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     4441 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     4501 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     4561 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     4621 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     4681 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     4741 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     4801 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     4861 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4921 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
