LOCUS       sequence-319369-        7139 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  human CD9-TEV cleavage site-tTA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7139)
  AUTHORS   Somiya M, Kuroda S
  TITLE     Reporter gene assay for membrane fusion of extracellular vesicles.
  JOURNAL   J Extracell Vesicles. 2021 Nov;10(13):e12171. doi: 
            10.1002/jev2.12171.
  PUBMED    34807503
REFERENCE   2  (bases 1 to 7139)
  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 Extracell
            Vesicles."; date: "2021-11"; volume: "10(13):e12171. doi"; pages: " 
            10.1002/jev2.12171"
COMMENT     Sequence Label: pcDNA3.1-CD9-(TEV)-tTA
FEATURES             Location/Qualifiers
     source          1..7139
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1442..1462
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             join(1460..2077,2078..2464)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIE
                     GLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHAD
                     ALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(1530..1549)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(2496..2513)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2502..2726
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2772..3200
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2859..2878)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3069..3090
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3209..3229)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3214..3543
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3394..3529
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3456..3475
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3610..4404
                     /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(3664..3683)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4274..4293
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4578..4699
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4615..4634)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4669..4688
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4748..4764)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4748..4764)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4761..4783)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4772..4788
                     /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(4796..4802,4803..4820,4821..4826))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4841..4862
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4979..4996)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5150..5738)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5230..5249)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5909..6700,6701..6769))
                     /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     6532..6551
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6770..6874)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6949..6968)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacttaagc ttggtaccga gctcggatcc actagtccag tgtggtggaa
      721 ttctgcagat atccagcaca gtggcggccg cgccaccatg ccggtcaaag gaggcaccaa
      781 gtgcatcaaa tacctgctgt tcggatttaa cttcatcttc tggcttgccg ggattgctgt
      841 ccttgccatt ggactatggc tccgattcga ctctcagacc aagagcatct tcgagcaaga
      901 aactaataat aataattcca gcttctacac aggagtctat attctgatcg gagccggcgc
      961 cctcatgatg ctggtgggct tcctgggctg ctgcggggct gtgcaggagt cccagtgcat
     1021 gctgggactg ttcttcggct tcctcttggt gatattcgcc attgaaatag ctgcggccat
     1081 ctggggatat tcccacaagg atgaggtgat taaggaagtc caggagtttt acaaggacac
     1141 ctacaacaag ctgaaaacca aggatgagcc ccagcgggaa acgctgaaag ccatccacta
     1201 tgcgttgaac tgctgtggtt tggctggggg cgtggaacag tttatctcag acatctgccc
     1261 caagaaggac gtactcgaaa ccttcaccgt gaagtcctgt cctgatgcca tcaaagaggt
     1321 cttcgacaat aaattccaca tcatcggcgc agtgggcatc ggcattgccg tggtcatgat
     1381 atttggcatg atcttcagta tgatcttgtg ctgtgctatc cgcaggaacc gcgagatggt
     1441 cgagaacctg tacttccagt ctagattaga taaaagtaaa gtgattaaca gcgcattaga
     1501 gctgcttaat gaggtcggaa tcgaaggttt aacaacccgt aaactcgccc agaagctagg
     1561 tgtagagcag cctacattgt attggcatgt aaaaaataag cgggctttgc tcgacgcctt
     1621 agccattgag atgttagata ggcaccatac tcacttttgc cctttagaag gggaaagctg
     1681 gcaagatttt ttacgtaata acgctaaaag ttttagatgt gctttactaa gtcatcgcga
     1741 tggagcaaaa gtacatttag gtacacggcc tacagaaaaa cagtatgaaa ctctcgaaaa
     1801 tcaattagcc tttttatgcc aacaaggttt ttcactagag aatgcattat atgcactcag
     1861 cgctgtgggg cattttactt taggttgcgt attggaagat caagagcatc aagtcgctaa
     1921 agaagaaagg gaaacaccta ctactgatag tatgccgcca ttattacgac aagctatcga
     1981 attatttgat caccaaggtg cagagccagc cttcttattc ggccttgaat tgatcatatg
     2041 cggattagaa aaacaactta aatgtgaaag tgggtccgcg tacagccgcg cgcgtacgaa
     2101 aaacaattac gggtctacca tcgagggcct gctcgatctc ccggacgacg acgcccccga
     2161 agaggcgggg ctggcggctc cgcgcctgtc ctttctcccc gcgggacaca cgcgcagact
     2221 gtcgacggcc cccccgaccg atgtcagcct gggggacgag ctccacttag acggcgagga
     2281 cgtggcgatg gcgcatgccg acgcgctaga cgatttcgat ctggacatgt tgggggacgg
     2341 ggattccccg ggtccgggat ttacccccca cgactccgcc ccctacggcg ctctggatat
     2401 ggccgacttc gagtttgagc agatgtttac cgatgccctt ggaattgacg agtacggtgg
     2461 gtagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     2521 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2581 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2641 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     2701 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     2761 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2821 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2881 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2941 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     3001 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     3061 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     3121 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     3181 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     3241 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     3301 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3361 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3421 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3481 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3541 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga
     3601 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3661 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3721 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3781 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3841 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3901 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3961 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4021 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4081 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     4141 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4201 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4261 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4321 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4381 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     4441 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4501 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4561 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     4621 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     4681 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     4741 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     4801 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     4861 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     4921 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4981 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5041 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5101 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5161 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5221 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5281 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5341 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5401 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5461 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5521 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5581 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5641 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5701 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5761 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5821 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     5881 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5941 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6001 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6061 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6121 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6181 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6241 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6301 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6361 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6421 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6481 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6541 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6601 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6661 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6721 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6781 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6841 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     6901 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     6961 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     7021 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
     7081 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
