LOCUS       sequence-350042-        6498 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual luciferase reporter for SARS-CoV-2 programmed -1 ribosomal 
            frameshifting. 3-nucleotide deletion in Stem 1 (negative control)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6498)
  AUTHORS   Sun Y, Abriola L, Niederer RO, Pedersen SF, Alfajaro MM, Silva 
            Monteiro V, Wilen CB, Ho YC, Gilbert WV, Surovtseva YV, Lindenbach 
            BD, Guo JU
  TITLE     Restriction of SARS-CoV-2 replication by targeting programmed -1 
            ribosomal frameshifting.
  JOURNAL   Proc Natl Acad Sci U S A. 2021 Jun 29;118(26). pii: 2023051118. doi:
            10.1073/pnas.2023051118.
  PUBMED    34185680
REFERENCE   2  (bases 1 to 6498)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2021-06-29"; volume: "118(26). pii: 
            2023051118. doi"; pages: " 10.1073/pnas.2023051118"
COMMENT     Sequence Label: pFRLuc-CoV2-delStem1
FEATURES             Location/Qualifiers
     source          1..6498
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             126..1055
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="ASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAENA
                     VIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAWF
                     ELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDIA
                     LIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPREI
                     PLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKVK
                     GLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             1107..1898
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV
                     LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS
                     HLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG
                     LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL
                     ATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1158..1177)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1768..1787
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1962..2010
                     /note="synthetic polyadenylation signal"
     promoter        2166..2270
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2271..2339,2340..3131)
                     /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(2489..2508)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(3289..3877)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3369..3388)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    3993..4276
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     promoter        4333..4832
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4368..4391)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     4765..4784
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      4873..4882
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4879..6498,1..30)
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
ORIGIN
        1 aaggccaaga agggcggcaa gatcgccgtg aataactcgt ttttaaacgg gtttgcggtg
       61 taagtgcagc ccgtctaccg tgcggcacag gcactagtac tgatgtcgta tacagggctt
      121 ttgacgcttc caaggtgtac gaccccgagc aacgcaaacg catgatcact gggcctcagt
      181 ggtgggctcg ctgcaagcaa atgaacgtgc tggactcctt catcaactac tatgattccg
      241 agaagcacgc cgagaacgcc gtgatttttc tgcatggtaa cgctgcctcc agctacctgt
      301 ggaggcacgt cgtgcctcac atcgagcccg tggctagatg catcatccct gatctgatcg
      361 gaatgggtaa gtccggcaag agcgggaatg gctcatatcg cctcctggat cactacaagt
      421 acctcaccgc ttggttcgag ctgctgaacc ttccaaagaa aatcatcttt gtgggccacg
      481 actggggggc ttgtctggcc tttcactact cctacgagca ccaagacaag atcaaggcca
      541 tcgtccatgc tgagagtgtc gtggacgtga tcgagtcctg ggacgagtgg cctgacatcg
      601 aggaggatat cgccctgatc aagagcgaag agggcgagaa aatggtgctt gagaataact
      661 tcttcgtcga gaccatgctc ccaagcaaga tcatgcggaa actggagcct gaggagttcg
      721 ctgcctacct ggagccattc aaggagaagg gcgaggttag acggcctacc ctctcctggc
      781 ctcgcgagat ccctctcgtt aagggaggca agcccgacgt cgtccagatt gtccgcaact
      841 acaacgccta ccttcgggcc agcgacgatc tgcctaagat gttcatcgag tccgaccctg
      901 ggttcttttc caacgctatt gtcgagggag ctaagaagtt ccctaacacc gagttcgtga
      961 aggtgaaggg cctccacttc agccaggagg acgctccaga tgaaatgggt aagtacatca
     1021 agagcttcgt ggagcgcgtg ctgaagaacg agcagaccgg tggtgggagc ggaggtggcg
     1081 gatcaggtgg cggaggctcc ggagggattg aacaagatgg attgcacgca ggttctccgg
     1141 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     1201 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     1261 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     1321 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     1381 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     1441 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     1501 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     1561 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     1621 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     1681 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     1741 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     1801 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     1861 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     1921 gaccgaccaa gcgacgccca acctgccatc acgatggccg caataaaata tctttatttt
     1981 cattacatct gtgtgttggt tttttgtgtg aatcgatagc gataaggatc ctctttgcgc
     2041 ttgcgttttc ccttgtccag atagcccagt agctgacatt catccggggt cagcaccgtt
     2101 tctgcggact ggctttctac gtaatggttt cttagacgtc aggtggcact tttcggggaa
     2161 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     2221 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     2281 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     2341 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     2401 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     2461 ttccaatgat gagcactttc aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     2521 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     2581 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     2641 ccataaccat gagtgataac actgcggcca acttacttct gacaactatc ggaggaccga
     2701 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     2761 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     2821 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     2881 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     2941 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     3001 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     3061 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     3121 agcattggta attcgaaatg accgaccaag cgacgcccaa ccggtatcag ctcactcaaa
     3181 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3241 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3301 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3361 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3421 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3481 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3541 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3601 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3661 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3721 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3781 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3841 caagcagcag attacgcgca gaaaaaaagg atttcaagaa gatcctttga tcttttctac
     3901 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     3961 aaaaaggatc ttcacctaga tccttttata gtccggaaat acaggaacgc acgctggatg
     4021 gcccttcgct gggatggtga aaccatgaaa aatggcagct tcagtggatt aagtgggggt
     4081 aatgtggcct gtaccctctg gttgcatagg tattcatacg gttaaaattt atcaggcgcg
     4141 attgcggcag tttttcgggt ggtttgttgc catttttacc tgtctgctgc cgtgatcgcg
     4201 ctgaacgcgt tttagcggtg cgtacaatta agggattatg gtaaatccac ttactgtctg
     4261 ccctcgtagc catcgagata aaccgcagta ctccggccac gatgcgtccg gcgtagagga
     4321 tcgagatcta ccgggtaggg gaggcgcttt tcccaaggca gtctggagca tgcgctttag
     4381 cagccccgct gggcacttgg cgctacacaa gtggcctctg gcctcgcaca cattccacat
     4441 ccaccggtag gcgccaaccg gctccgttct ttggtggccc cttcgcgcca ccttctactc
     4501 ctcccctagt caggaagttc ccccccgccc cgcagctcgc gtcgtgcagg acgtgacaaa
     4561 tggaagtagc acgtctcact agtctcgtgc agatggacag caccgctgag caatggaagc
     4621 gggtaggcct ttggggcagc ggccaatagc agctttgctc cttcgctttc tgggctcaga
     4681 ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc ggggcgggcg
     4741 cccgaaggtc ctccggaggc ccggcattct gcacgcttca aaagcgcacg tctgccgcgc
     4801 tgttctcctc ttcctcatct ccgggccttt cgacctgcag cccaagcttg gcaatccggt
     4861 actgttggta aagccaccat ggaagatgcc aaaaacatta agaagggccc agcgccattc
     4921 tacccactcg aagacgggac cgccggcgag cagctgcaca aagccatgaa gcgctacgcc
     4981 ctggtgcccg gcaccatcgc ctttaccgac gcacatatcg aggtggacat tacctacgcc
     5041 gagtacttcg agatgagcgt tcggctggca gaagctatga agcgctatgg gctgaataca
     5101 aaccatcgga tcgtggtgtg cagcgagaat agcttgcagt tcttcatgcc cgtgttgggt
     5161 gccctgttca tcggtgtggc tgtggcccca gctaacgaca tctacaacga gcgcgagctg
     5221 ctgaacagca tgggcatcag ccagcccacc gtcgtattcg tgagcaagaa agggctgcaa
     5281 aagatcctca acgtgcaaaa gaagctaccg atcatacaaa agatcatcat catggatagc
     5341 aagaccgact accagggctt ccaaagcatg tacaccttcg tgacttccca tttgccaccc
     5401 ggcttcaacg agtacgactt cgtgcccgag agcttcgacc gggacaaaac catcgccctg
     5461 atcatgaaca gtagtggcag taccggattg cccaagggcg tagccctacc gcaccgcacc
     5521 gcttgtgtcc gattcagtca tgcccgcgac cccatcttcg gcaaccagat catccccgac
     5581 accgctatcc tcagcgtggt gccatttcac cacggcttcg gcatgttcac cacgctgggc
     5641 tacttgatct gcggctttcg ggtcgtgctc atgtaccgct tcgaggagga gctattcttg
     5701 cgcagcttgc aagactataa gattcaatct gccctgctgg tgcccacact atttagcttc
     5761 ttcgctaaga gcactctcat cgacaagtac gacctaagca acttgcacga gatcgccagc
     5821 ggcggggcgc cgctcagcaa ggaggtaggt gaggccgtgg ccaaacgctt ccacctacca
     5881 ggcatccgcc agggctacgg cctgacagaa acaaccagcg ccattctgat cacccccgaa
     5941 ggggacgaca agcctggcgc agtaggcaag gtggtgccct tcttcgaggc taaggtggtg
     6001 gacttggaca ccggtaagac actgggtgtg aaccagcgcg gcgagctgtg cgtccgtggc
     6061 cccatgatca tgagcggcta cgttaacaac cccgaggcta caaacgctct catcgacaag
     6121 gacggctggc tgcacagcgg cgacatcgcc tactgggacg aggacgagca cttcttcatc
     6181 gtggaccggc tgaagagcct gatcaaatac aagggctacc aggtagcccc agccgaactg
     6241 gagagcatcc tgctgcaaca ccccaacatc ttcgacgccg gggtcgccgg cctgcccgac
     6301 gacgatgccg gcgagctgcc cgccgcagtc gtcgtgctgg aacacggtaa aaccatgacc
     6361 gagaaggaga tcgtggacta tgtggccagc caggttacaa ccgccaagaa gctgcgcggt
     6421 ggtgttgtgt tcgtggacga ggtgcctaaa ggactgaccg gcaagttgga cgcccgcaag
     6481 atccgcgaga ttctcatt
//
