LOCUS       sequence-224195-        7754 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of DENV2 NS2B3 protein with a C-terminal V5 
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7754)
  AUTHORS   Udawatte DJ, Lang DM, Currier JR, Medin CL, Rothman AL
  TITLE     Dengue virus downregulates TNFR1- and TLR3-stimulated NF-kappaB 
            activation by targeting RIPK1.
  JOURNAL   Front Cell Infect Microbiol. 2022 Oct 14;12:926036. doi: 
            10.3389/fcimb.2022.926036. eCollection 2022.
  PUBMED    36310878
REFERENCE   2  (bases 1 to 7754)
  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: "Front Cell 
            Infect Microbiol."; date: "2022-10-14"; volume: "12:926036. doi"; 
            pages: " 10.3389/fcimb.2022.926036. eCollection 2022"
COMMENT     Sequence Label: pcDNA_DENV2-NS2B3_V5
FEATURES             Location/Qualifiers
     source          1..7754
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        136..515
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        516..719
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     670..690
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     764..783
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        764..782
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    812..836
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3095..3119)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3145..3186
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3226..3245)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3270..3318
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3520..3948
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3607..3626)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3817..3838
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3957..3977)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3962..4291
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4142..4277
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4204..4223
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4358..5152
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4412..4431)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5022..5041
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5328..5449
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5365..5384)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5419..5438
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5498..5514)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5498..5514)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5511..5533)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5522..5538
                     /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)."
     promoter        complement(join(5546..5552,5553..5570,5571..5576))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5591..5612
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5729..5746)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5900..6488)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5980..5999)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6659..7450,7451..7519))
                     /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     7282..7301
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7520..7624)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7699..7718)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
       61 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
      121 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
      181 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
      241 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
      301 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
      361 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
      421 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
      481 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
      541 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
      601 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
      661 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc
      721 tctggctaac tagagaaccc actgcttact ggcttatcga aattaatacg actcactata
      781 gggagaccca agctggctag ttaagctatc aacaagtttg tacaaaaaag caggcttcga
      841 accatggcaa gctggccatt aaatgaggct atcatggcag tcgggatggt gagcattttg
      901 gccagttcac tcctaaagaa tgacattccc atgacaggac cattagtggc tggagggctc
      961 ctcactgtgt gctacgtgct cactggacga tcggccgatt tggaactgga gagagccgcc
     1021 gatgtcaaat gggaagatca ggcagagata tcaggaagca gtccaatcct gtcaataaca
     1081 atatcagaag atggtagcat gtcgataaaa aacgaagagg aagaacaaac actgaccata
     1141 ctcattagaa caggattgct ggtgatctca ggactttttc ctgtatcaat accaatcacg
     1201 gcagcagcat ggtacctgtg ggaagtgaag aaacaacggg ctggagtatt gtgggatgtc
     1261 ccttcacccc cacccgtggg aaaggctgaa ctggaagatg gagcctatag aatcaagcaa
     1321 aaagggattc ttggatattc ccagatcgga gccggagttt acaaagaagg aacattccat
     1381 acaatgtggc atgtcacacg cggcgctgtt ctaatgcata aaggaaagag gattgaacca
     1441 tcatgggcgg acgttaagaa agacctaata tcatatggag gaggctggaa gctagaagga
     1501 gaatggaagg aaggagaaga agtccaggtc ttggcattgg agcctggaaa aaatccaaga
     1561 gccgtccaaa caaaacctgg tcttttcaaa accaacgccg gaaccatagg tgccgtatct
     1621 ctggactttt ctcctggaac ctcaggatct ccaatcatcg acaaaaaagg aaaagttgtg
     1681 ggtctttatg gtaatggtgt tgttacaagg agtggggcat atgtgagtgc tatagcccag
     1741 actgaaaaaa gtattgaaga caatccagag atcgaagatg acatttttcg aaagagaaaa
     1801 ttgaccatca tggacctcca cccaggagcg ggaaagacga agagatacct tccggccata
     1861 gtcagagagg ctataaaacg gggcctgagg acattaatcc tggcccccac tagagtcgtg
     1921 gcagctgaaa tggaggaagc cctaagagga cttccaataa gataccaaac cccagccatc
     1981 agagctgagc acaccgggcg ggagattgtg gacctaatgt gtcatgccac attcactatg
     2041 aggctgctat caccagttag agtgccaaat tacaacctga tcatcatgga cgaagcccat
     2101 ttcacagacc cagcaagtat agcggctaga ggatacatct caactcgagt agagatgggt
     2161 gaggtagctg ggattttcat gacagccact cctccgggaa gcagagaccc attccctcag
     2221 agcaatgcac caatcatgga tgaagaaaga gaaatccctg aacgttcgtg gagttctgga
     2281 catgagtggg tcacggattt taaagggaag actgtttggt tcgttccaag tataaaagca
     2341 ggaaatgata tagcagcttg cctgagaaaa aatggaaaga aagtgataca actcagtagg
     2401 aagacctttg attctgagta tgtcaagact agaaccaatg attgggactt cgtggtcaca
     2461 actgacattt cagaaatggg tgccaacttc aaggctgaga gggttataga ccccagacgc
     2521 tgcatgaaac cagttatact aacagatggt gaagagcggg tgatcctggc aggacctatg
     2581 ccagtgaccc actctagtgc agcacaaaga agagggagaa taggaagaaa tccaaaaaat
     2641 gaaaatgacc agtacatata catgggggaa cctctggaaa atgatgaaga ctgtgcacac
     2701 tggaaagaag ctaaaatgct cctaaataac atcaacacac ctgaaggaat cattcctagc
     2761 atgttcgaac cagggcgtga aaaggtggat gccattgatg gtgaataccg cttgagagga
     2821 gaagcaagga aaacctttgt ggacctaatg agaagaggag acctaccagt ctggttggcc
     2881 tacagagtgg cagctgaagg catcaactac gcagacagaa ggtggtgttt tgatggaatt
     2941 aagaacaacc aaatcttgga agaaaatgtg gaggtggaaa tctggacaaa agaaggggaa
     3001 aggaagaaat taaaacccag atggttggat gccaggatct actctgaccc actggcgcta
     3061 aaggaattca aggaatttgc agccggaaga aaggacccag ctttcttgta caaagtggtt
     3121 gatctagagg gcccgcggtt cgaaggtaag cctatcccta accctctcct cggtctcgat
     3181 tctacgcgta ccggttagta atgagtttaa acgggggagg ctaactgaaa cacggaagga
     3241 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     3301 gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata
     3361 ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc
     3421 ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca
     3481 tagcagatct gcgcagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat
     3541 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     3601 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     3661 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     3721 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt
     3781 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa
     3841 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     3901 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa
     3961 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     4021 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     4081 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     4141 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     4201 ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg
     4261 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt
     4321 cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     4381 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     4441 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     4501 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
     4561 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     4621 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     4681 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     4741 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     4801 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     4861 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
     4921 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     4981 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     5041 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     5101 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
     5161 ctggggttcg cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     5221 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     5281 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     5341 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     5401 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     5461 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5521 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5581 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5641 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5701 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5761 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5821 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5881 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5941 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6001 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6061 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6121 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6181 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6241 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6301 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6361 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6421 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6481 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6541 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6601 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6661 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6721 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6781 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6841 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6901 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6961 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     7021 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7081 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7141 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7201 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7261 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7321 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7381 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7441 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7501 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7561 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7621 cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc
     7681 cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct
     7741 ccctgcttgt gtgt
//
