LOCUS       sequence-443384-        7609 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of CHMP2A fused to the hepatitis C virus 
            protease NS3 through a short linker containing the NS3 cleavage 
            site, along with a blue fluorescent protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7609)
  AUTHORS   Wang H, Gallet B, Moriscot C, Pezet M, Chatellard C, Kleman JP, 
            Gottlinger H, Weissenhorn W, Boscheron C
  TITLE     An Inducible ESCRT-III Inhibition Tool to Control HIV-1 Budding.
  JOURNAL   Viruses. 2023 Nov 22;15(12):2289. doi: 10.3390/v15122289.
  PUBMED    38140530
REFERENCE   2  (bases 1 to 7609)
  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: "Viruses."; 
            date: "2023-11-22"; volume: "15(12):2289. doi"; pages: " 
            10.3390/v15122289"
COMMENT     Sequence Label: pCHMP2A-NS3-blue
FEATURES             Location/Qualifiers
     source          1..7609
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1601..1624
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             2315..2338
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    2359..2424
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      2432..2441
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2438..2440,2441..2443,2444..3157)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2483..2504)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2744..2763)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3091..3112
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3200..3217)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3206..3430
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3476..3904
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3563..3582)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3773..3794
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3913..3933)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3918..4247
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4098..4233
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4160..4179
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4314..5108
                     /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(4368..4387)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4978..4997
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5282..5403
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5319..5338)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5373..5392
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5452..5468)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5452..5468)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5465..5487)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5476..5492
                     /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(5500..5506,5507..5524,5525..5530))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5545..5566
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5683..5700)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5854..6442)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5934..5953)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6613..7404,7405..7473))
                     /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     7236..7255
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7474..7578)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttatgg acctgctgtt cggcagaaga aagacccctg aggaactgct
      961 gcggcagaac cagagagccc tgaacagagc catgagagag ctggacagag agcggcagaa
     1021 gctggaaacc caagagaaga agatcattgc cgacatcaag aagatggcca agcagggcca
     1081 gatggacgcc gtcagaatca tggccaaaga cctcgtgcgg actcggagat acgtgcggaa
     1141 gttcgtgctg atgcgggcca atatccaggc cgtgtctctg aagatccaga cactgaagtc
     1201 caacaacagc atggcccagg ccatgaaggg cgtgacaaag gccatgggca ccatgaacag
     1261 acagctgaag ctgccccaga tccagaaaat catgatggaa ttcgagcggc aggccgagat
     1321 catggacatg aaggaagaga tgatgaacga cgccatcgac gatgccatgg gcgacgaaga
     1381 ggatgaagag gaatccgacg ccgtggtgtc ccaggtgctg gatgaactgg gactgagcct
     1441 gaccgacgag ctgagcaatc tgcctagcac aggcggatct ctgtctgtgg ctgccggcgg
     1501 aaagaaagcc gaagctgctg cttctgccct ggccgatgct gatgccgatc tggaagagag
     1561 actgaagaac ctgcggcggg atggcggctc tacaggcaat tggagccatc ctcagttcga
     1621 gaaaggcagc ggcagctgtc aagagtttga tgagatggaa gagtgcagcc agcatctgcc
     1681 tggcgctgga tcttctggcg gatctcacat ggccagcatg aagaaaaagg gcagcgtggt
     1741 catcgtgggc agaatcaacc tgagcggcga taccgcctac gctcagcaga caagaggcga
     1801 ggaaggctgc caagagacaa gccagaccgg cagagacaag aaccaggtgg aaggcgaggt
     1861 gcagatcgtg tctacagcta cccagacctt cctggccacc agcatcaatg gcgtgctgtg
     1921 gacagtgtat cacggcgctg gcaccaggac aatcgcctct ccaaaaggcc ccgtgacaca
     1981 gatgtacacc aatgtggaca aagatctggt gggttggcag gcaccgcaag gcagccgtag
     2041 cctgacgccg tgcacctgtg gcagttccga tctgtatctg gttacccgcc atgcggacgt
     2101 gattccggtt cgtcgccgtg gcgatagtcg tggttccctg ctgtcaccgc gcccgatctc
     2161 gtacctgaaa ggttcatcgg gtggtccgct gctgtgcccg gcaggtcacg cagttggtat
     2221 ttttcgtgca gcagtcagca cccgcggcgt ggctaaagcg gttgatttca tcccggtgga
     2281 atccctggaa accacgatgc gttctggcag cggcgactac aaagacgatg atgataaatc
     2341 ctccggctcc ggcagcgctc cggactcaga tctcgagctc aagcttcgaa ttctgcagtc
     2401 gacggtaccg cgggcccggg atccaccggt cgccaccatg gtgagcaagg gcgaggagct
     2461 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     2521 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat
     2581 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgtcctgggg
     2641 cgtgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     2701 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     2761 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     2821 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actactttag
     2881 cgacaacgtc tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat
     2941 ccgccacaac atcgaggacg gcggcgtgca gctcgccgac cactaccagc agaacacccc
     3001 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccaagct
     3061 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
     3121 cgggatcact ctcggcatgg acgagctgta caagtaagcg gccgcgactc tagagggccc
     3181 gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     3241 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     3301 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     3361 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     3421 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg
     3481 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     3541 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     3601 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     3661 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     3721 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     3781 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     3841 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     3901 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     3961 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     4021 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     4081 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     4141 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     4201 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     4261 cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg
     4321 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     4381 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     4441 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg
     4501 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     4561 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     4621 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     4681 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     4741 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     4801 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg
     4861 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     4921 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     4981 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     5041 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     5101 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     5161 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     5221 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     5281 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     5341 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     5401 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     5461 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     5521 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5581 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5641 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5701 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5761 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5821 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5881 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5941 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     6001 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6061 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6121 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6181 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6241 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     6301 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6361 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6421 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6481 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6541 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6601 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6661 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6721 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6781 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6841 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6901 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6961 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     7021 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7081 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7141 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7201 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7261 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7321 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7381 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7441 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7501 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7561 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
