LOCUS       sequence-426578-        8706 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Produces the Chikungunya virus nonstructural protein 2 with a 
            c-terminal 3xFLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8706)
  AUTHORS   Reyes Ballista JM, Hoover AJ, Noble JT, Acciani MD, Miazgowicz KL, 
            Harrison SA, Tabscott GAL, Duncan A, Barnes DN, Jimenez AR, Brindley
            MA
  TITLE     Chikungunya virus release is reduced by TIM-1 receptors through 
            binding of envelope phosphatidylserine.
  JOURNAL   J Virol. 2024 Jul 15:e0077524. doi: 10.1128/jvi.00775-24.
  PUBMED    39007616
REFERENCE   2  (bases 1 to 8706)
  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: "J Virol. 
            2024 Jul 15:e0077524. doi: 10.1128/jvi.00775-24."
COMMENT     Sequence Label: pcDNAintron-CHIKV-nsp2-c3xFLAG
FEATURES             Location/Qualifiers
     source          1..8706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     325..344
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        325..343
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      511..520
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2914..2979
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3056..3073)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3062..3286
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3332..3760
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3419..3438)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3629..3650
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3769..3789)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3774..4103
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3954..4089
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4016..4035
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4170..4964
                     /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(4224..4243)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4834..4853
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5138..5259
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5175..5194)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5229..5248
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5308..5324)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5308..5324)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5321..5343)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5332..5348
                     /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(5356..5362,5363..5380,5381..5386))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5401..5422
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5539..5556)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5710..6298)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5790..5809)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6469..7260,7261..7329))
                     /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     7092..7111
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7330..7434)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7509..7528)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        7700..8079
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8080..8283
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8234..8254
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     8280..8304
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          join(8404..8706,1..270)
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
ORIGIN
        1 caaggcaatc agggtatatt atattgtact tcagcacagt tttagagaac aattgttata
       61 attaaatgat aaggtagaat atttctgcat ataaattctg gctggcgtgg aaatattctt
      121 attggtagaa acaactacat cctggtcatc atcctgcctt tctctttatg gttacaatga
      181 tatacactgt ttgagatgag gataaaatac tctgagtcca aaccgggccc ctctgctaac
      241 catgttcatg ccttcttctt tttcctacag ctcctgggca acgtgctggt tattgtgctg
      301 tctcatcatt ttggcaaaga attgtaatac gactcactat agggcgaatt cggatccact
      361 agtccagtgt ggtggaattc tgcagatcta gagcctctgc taaccatgtt catgccttct
      421 tctttttcct acagctcctg ggcaacgtgc tggttattgt gctgtctcat cattttggca
      481 aagaattcga gctcatcgat gcatggtacc gccaccatgg gaataataga gactccgaga
      541 ggcgctatca aagttactgc ccaactaaca gaccacgtcg tgggggagta cctggtactt
      601 tccccgcaga ccgtactacg cagccagaag ctcagcctga tccacgcttt agcggagcaa
      661 gtgaagacgt gtacgcacag cggacgagca gggaggtatg cggtcgaagc gtacgatggc
      721 cgagtcctag tgccctcagg ctatgcaatt tcgcctgaag acttccagag tctaagcgaa
      781 agcgcaacga tggtgtacaa cgaaagagag ttcgtaaaca gaaagttaca ccacattgcg
      841 atgcacggac cagccctgaa cactgacgaa gagtcgtatg agctggtgag ggcagagagg
      901 acagaacacg agtacgtcta cgacgtggac cagagaagat gctgtaagaa ggaagaagct
      961 gcaggactgg tactggtggg cgacttgact aatccgccct accacgaatt cgcatacgaa
     1021 gggctaaaaa ttcgccccgc ctgcccatac aaaattgcag tcataggagt cttcggggta
     1081 ccaggatctg gcaagtcagc cattatcaag aacctagtta ccaggcaaga cctggtgact
     1141 agcggaaaga aagaaaactg ccaagaaatc agcaccgacg tgatgagaca gagaggtcta
     1201 gagatatctg cacgtacggt agattcgctg ctcttgaatg gatgcaacag accagtcgac
     1261 gtgttgtacg tagacgaggc gtttgcgtgc cactctggaa cgttacttgc tttgatcgcc
     1321 ttggtgagac caagacagaa agttgtactt tgtggtgacc cgaagcagtg cggcttcttc
     1381 aatatgatgc agatgaaagt caactacaat cataacatct gcacccaagt gtaccacaaa
     1441 agtatctcca ggcggtgtac actgcctgtg actgccattg tgtcatcgtt gcattacgaa
     1501 ggcaaaatgc gcactacgaa tgagtacaac atgccgattg tagtggacac tacaggctca
     1561 acgaaacctg accctggaga cctcgtgtta acgtgcttca gagggtgggt taaacaactg
     1621 caaattgact atcgtggaca cgaggtcatg acagcagccg catcccaagg gttaactaga
     1681 aaaggagttt acgcagttag gcaaaaagtt aacgaaaacc cactctatgc atcaacatca
     1741 gagcacgtca acgtactcct aacgcgtacg gaaggtaaac tggtatggaa gacactctct
     1801 ggtgacccgt ggataaagac gctgcagaac ccaccgaaag gaaacttcaa agcaactatt
     1861 aaggagtggg aggtggagca cgcatcgata atggcgggca tctgcagtca ccaagtgacc
     1921 tttgacacat tccaaaacaa agccaacgtt tgctgggcta agagcttggt ccctatcctc
     1981 gaaacagcgg ggataaaact aaatgatagg cagtggtccc agataattca agccttcaaa
     2041 gaagacaaag catactcacc cgaagtagcc ctgaatgaaa tatgcacgcg catgtatggg
     2101 gtggatctag acagtgggct attctctaaa ccgttggtat ctgtgtatta cgcggataac
     2161 cattgggata ataggccggg aggaaagatg ttcggattca accctgaggc agcgtccatt
     2221 ctagaaagaa agtacccatt tacaaaagga aagtggaaca tcaacaagca gatctgcgtg
     2281 actaccagga ggatagaaga cttcaaccct accaccaaca ttataccggt caacaggaga
     2341 ctaccacact cattagtggc cgaacaccgc ccagtaaaag gggaaagaat ggaatggctg
     2401 gttaacaaga taaacggaca ccacgtactc ctggttagcg gctataacct tgcactgcct
     2461 actaagagag tcacctgggt agcgccacta ggtgtccgcg gagcggacta tacatacaac
     2521 ctagagctgg gtctaccagc aacacttggt aggtatgacc tagtggtcat aaacatccac
     2581 acaccttttc gcatacacca ttaccaacag tgcgtagatc acgcaatgaa actgcaaatg
     2641 ctagggggtg actcactgag actgctcaaa ccgggtggct ctctattgat cagagcatac
     2701 ggttacgcag atagaaccag tgaacgagtc atctgcgtac tgggacgcaa gtttagatcg
     2761 tctagagcat tgaaaccacc atgtgtcacc agtaatactg agatgttttt cctatttagc
     2821 aattttgaca atggcagaag gaattttaca acgcatgtca tgaacaatca actgaatgca
     2881 gcctttgtag gacaggccac ccgagcagga tgtgactaca aagaccatga cggtgattat
     2941 aaagatcatg atatcgacta caaggatgac gatgacaagt gactcgagct agcagatcat
     3001 ccagcacagt ggcggccgct cgagtctaga gggcccgttt aaacccgctg atcagcctcg
     3061 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     3121 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     3181 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     3241 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa
     3301 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg
     3361 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     3421 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     3481 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     3541 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     3601 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     3661 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     3721 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     3781 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3841 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3901 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3961 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4021 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     4081 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     4141 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     4201 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     4261 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     4321 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg
     4381 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     4441 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     4501 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     4561 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     4621 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     4681 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     4741 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     4801 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     4861 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     4921 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     4981 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     5041 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     5101 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     5161 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     5221 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     5281 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     5341 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     5401 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     5461 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     5521 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     5581 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     5641 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     5701 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     5761 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     5821 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     5881 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5941 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     6001 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     6061 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     6121 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     6181 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     6241 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     6301 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     6361 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     6421 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     6481 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     6541 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     6601 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     6661 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     6721 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     6781 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     6841 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     6901 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     6961 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     7021 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     7081 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     7141 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     7201 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     7261 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     7321 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     7381 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     7441 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc
     7501 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt
     7561 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg
     7621 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta
     7681 cgggccagat atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg
     7741 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     7801 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     7861 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     7921 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     7981 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     8041 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
     8101 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     8161 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     8221 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     8281 gctcgtttag tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat
     8341 agaagacacc gggaccgatc cagcctccgg gggatcgatc cccccgatcc tgagaacttc
     8401 agggtgagtt tggggaccct tgattgttct ttctttttcg ctattgtaaa attcatgtta
     8461 tatggagggg gcaaagtttt cagggtgttg tttagaatgg gaagatgtcc cttgtatcac
     8521 catggaccct catgataatt ttgtttcttt cactttctac tctgttgaca accattgtct
     8581 cctcttattt tcttttcatt ttctgtaact ttttcgttaa actttagctt gcatttgtaa
     8641 cgaattttta aattcacttt tgtttatttg tcagattgta agtactttct ctaatcactt
     8701 tttttt
//
