LOCUS       sequence-363200-        7588 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for mammalian cells: Expression of mScarlet 
            with SARS-CoV-2 5'UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7588)
  AUTHORS   Vora SM, Fontana P, Mao T, Leger V, Zhang Y, Fu TM, Lieberman J, 
            Gehrke L, Shi M, Wang L, Iwasaki A, Wu H
  TITLE     Targeting stem-loop 1 of the SARS-CoV-2 5' UTR to suppress viral 
            translation and Nsp1 evasion.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 Mar 1;119(9). pii: 2117198119. doi: 
            10.1073/pnas.2117198119.
  PUBMED    35149555
REFERENCE   2  (bases 1 to 7588)
  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: "2022-03-1"; volume: "119(9). pii: 
            2117198119. doi"; pages: " 10.1073/pnas.2117198119"
COMMENT     Sequence Label: pLV-mScarlet-SARS-CoV-2-5'UTR
FEATURES             Location/Qualifiers
     source          1..7588
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             787..1482
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(920..938)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1188..1207
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    1492..2080
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1545..1565)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1963..1974)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2167..2400
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2472..2593
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2509..2528)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2563..2582
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2633..2768
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2788..2807)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2789..2807)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2817..2834)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2817..2833)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2826..2848)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2975..3430
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3062..3081)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3272..3293
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3456..3560
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3561..3629,3630..4421)
                     /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(3779..3798)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4592..5180
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5081..5100
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5334..5351
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5468..5489
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5504..5509,5510..5527,5528..5534)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5542..5558
                     /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     5547..5569
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5566..5582
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5566..5582
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5601..5621
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5603..5621
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5649..5875
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5876..6056
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6103..6228
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6721..6954
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7139..7183
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    7450..7566
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctcg tttagtgaac cattaaaggt ttataccttc
      541 ccaggtaaca aaccaaccaa ctttcgatct cttgtagatc tgttctctaa acgaacttta
      601 aaatctgtgt ggctgtcact cggctgcatg cttagtgcac tcacgcagta taattaataa
      661 ctaattactg tcgttgacag gacacgagta actcgtctat cttctgcagg ctgcttacgg
      721 tttcgtccgt gttgcagccg atcatcagca catctaggtt tcgtccgggt gtgaccgaaa
      781 ggtaagatgg tgagcaaggg cgaggcagtg atcaaggagt tcatgcggtt caaggtgcac
      841 atggagggct ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc
      901 tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttctcc
      961 tgggacatcc tgtcccctca gttcatgtac ggctccaggg ccttcaccaa gcaccccgcc
     1021 gacatccccg actactataa gcagtccttc cccgagggct tcaagtggga gcgcgtgatg
     1081 aacttcgagg acggcggcgc cgtgaccgtg acccaggaca cctccctgga ggacggcacc
     1141 ctgatctaca aggtgaagct ccgcggcacc aacttccctc ctgacggccc cgtaatgcag
     1201 aagaagacaa tgggctggga agcgtccacc gagcggttgt accccgagga cggcgtgctg
     1261 aagggcgaca ttaagatggc cctgcgcctg aaggacggcg gccgctacct ggcggacttc
     1321 aagaccacct acaaggccaa gaagcccgtg cagatgcccg gcgcctacaa cgtcgaccgc
     1381 aagttggaca tcacctccca caacgaggac tacaccgtgg tggaacagta cgaacgctcc
     1441 gagggccgcc actccaccgg cggcatggac gagctgtaca agtgagtcga caatcaacct
     1501 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     1561 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     1621 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     1681 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     1741 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     1801 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     1861 gacaattccg tggtgttgtc ggggaagctg acgtcctttc catggctgct cgcctgtgtt
     1921 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     1981 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     2041 cctcagacga gtcggatctc cctttgggcc gcctccccgc ctggaattcg agctcggtac
     2101 ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg
     2161 ggggactgga agggctaatt cactcccaac gaagacaaga tctgcttttt gcttgtactg
     2221 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac
     2281 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt
     2341 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca
     2401 gtagtagttc atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag
     2461 agagtgagag gaacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     2521 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     2581 tcaatgtatc ttatcatgtc tggctctagc tatcccgccc ctaactccgc ccatcccgcc
     2641 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     2701 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     2761 tggaggccta gggacgtacc caattcgccc tatagtgagt cgtattacgc gcgctcactg
     2821 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     2881 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     2941 tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     3001 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     3061 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3121 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3181 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     3241 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3301 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3361 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     3421 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     3481 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     3541 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     3601 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     3661 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     3721 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     3781 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     3841 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     3901 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     3961 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4021 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4081 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     4141 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4201 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4261 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4321 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4381 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     4441 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4501 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4561 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4621 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4681 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     4741 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4801 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4861 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     4921 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     4981 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5041 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5101 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5161 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5221 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5281 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5341 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     5401 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     5461 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttccac tttatgcttc
     5521 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     5581 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctgc
     5641 aagcttaatg tagtcttatg caatactctt gtagtcttgc aacatggtaa cgatgagtta
     5701 gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc cgattggtgg aagtaaggtg
     5761 gtacgatcgt gccttattag gaaggcaaca gacgggtctg acatggattg gacgaaccac
     5821 tgaattgccg cattgcagag atattgtatt taagtgccta gctcgataca taaacgggtc
     5881 tctctggtta gaccagatct gagcctggga gctctctggc taactaggga acccactgct
     5941 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga
     6001 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg
     6061 cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg agctctctcg acgcaggact
     6121 cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc gactggtgag tacgccaaaa
     6181 attttgacta gcggaggcta gaaggagaga gatgggtgcg agagcgtcag tattaagcgg
     6241 gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat
     6301 aaattaaaac atatagtatg ggcaagcagg gagctagaac gattcgcagt taatcctggc
     6361 ctgttagaaa catcagaagg ctgtagacaa atactgggac agctacaacc atcccttcag
     6421 acaggatcag aagaacttag atcattatat aatacagtag caaccctcta ttgtgtgcat
     6481 caaaggatag agataaaaga caccaaggaa gctttagaca agatagagga agagcaaaac
     6541 aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc agacctggag gaggagatat
     6601 gagggacaat tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg
     6661 agtagcaccc accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat
     6721 aggagctttg ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat
     6781 gacgctgacg gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt
     6841 gctgagggct attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca
     6901 gctccaggca agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat
     6961 ttggggttgc tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag
     7021 taataaatct ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat
     7081 taacaattac acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa
     7141 gaatgaacaa gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat
     7201 aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt
     7261 aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt
     7321 atcgtttcag acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga
     7381 agaaggtgga gagagagaca gagacagatc cattcgatta gtgaacggat ctcgacggta
     7441 tcggttaact tttaaaagaa aaggggggat tggggggtat agtgcagggg aaagaatagt
     7501 agacataata gcaacagaca tacaaactaa agaattacaa aaacaaatta caaaattcaa
     7561 aattttatcg ataagcttgg gagttccg
//
