LOCUS       sequence-350185-        6288 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of tetraspanin CD63 fused to a C-terminal HA 
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6288)
  AUTHORS   Hung ME, Leonard JN
  TITLE     A platform for actively loading cargo RNA to elucidate limiting 
            steps in EV-mediated delivery.
  JOURNAL   J Extracell Vesicles. 2016 May 13;5:31027. doi: 
            10.3402/jev.v5.31027. eCollection 2016.
  PUBMED    27189348
REFERENCE   2  (bases 1 to 6288)
  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: "2016-05-13"; volume: "5:31027. doi"; pages: " 
            10.3402/jev.v5.31027. eCollection 2016"
COMMENT     Sequence Label: pMEH371 CD63 HA
FEATURES             Location/Qualifiers
     source          1..6288
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        85..464
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        465..668
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     619..639
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     713..732
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        713..731
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             751..1464
                     /codon_start=1
                     /product="human CD63 tetraspanin"
                     /label=CD63
                     /note="enriched in intraluminal vesicles of late endosomes 
                     (multivesicular bodies) and in exosomes"
                     /translation="MAVEGGMKCVKFLLYVLLLAFCACAVGLIAVGVGAQLVLSQTIIQ
                     GATPGSLLPVVIIAVGVFLFLVAFVGCCGACKENYCLMITFAIFLSLIMLVEVAAAIAG
                     YVFRDKVMSEFNNNFRQQMENYPKNNHTASILDRMQADFKCCGAANYTDWEKIPSMSKN
                     RVPDSCCINVTVGCGINFNEKAIHKEGCVEKIGGWLRKNVLVVAAAALGIAFVEVLGIV
                     FACCLVKSIRSGYEVM"
     CDS             1486..1512
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1560..1577)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1566..1790
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1836..2264
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1923..1942)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2133..2154
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2273..2293)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2278..2607
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2458..2593
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2520..2539
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2656..3681
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             2656..3681
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    3811..3932
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3848..3867)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3902..3921
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3981..3997)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3981..3997)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3994..4016)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4005..4021
                     /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(4029..4035,4036..4053,4054..4059))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4074..4095
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4212..4229)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4383..4971)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4463..4482)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5142..5933,5934..6002))
                     /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     5765..5784
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6003..6107)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6182..6201)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg
       61 atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa
      121 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      181 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      241 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      301 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      361 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
      421 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
      481 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
      541 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
      601 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
      661 gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga
      721 ctcactatag ggagacccaa gctgggtacc atggcggtgg aaggaggaat gaaatgtgtg
      781 aagttcttgc tctacgtcct cctgctggcc ttttgcgcct gtgcagtggg actgattgcc
      841 gtgggtgtcg gggcacagct tgtcctgagt cagaccataa tccagggggc tacccctggc
      901 tctctgttgc cagtggtcat catcgcagtg ggtgtcttcc tcttcctggt ggcttttgtg
      961 ggctgctgcg gggcctgcaa ggagaactat tgtcttatga tcacgtttgc catctttctg
     1021 tctcttatca tgttggtgga ggtggccgca gccattgctg gctatgtgtt tagagataag
     1081 gtgatgtcag agtttaataa caacttccgg cagcagatgg agaattaccc gaaaaacaac
     1141 cacactgctt cgatcctgga caggatgcag gcagatttta agtgctgtgg ggctgctaac
     1201 tacacagatt gggagaaaat cccttccatg tcgaagaacc gagtccccga ctcctgctgc
     1261 attaatgtta ctgtgggctg tgggattaat ttcaacgaga aggcgatcca taaggagggc
     1321 tgtgtggaga agattggggg ctggctgagg aaaaatgtgc tggtggtagc tgcagcagcc
     1381 cttggaattg cttttgtcga ggttttggga attgtctttg cctgctgcct cgtgaagagt
     1441 atcagaagtg gctacgaggt gatggctagc agtggctctg gtagctaccc atacgatgtg
     1501 ccagattacg cttaagcggc cgctcgagtc tagagggccc gtttaaaccc gctgatcagc
     1561 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     1621 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     1681 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     1741 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     1801 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     1861 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1921 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1981 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     2041 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     2101 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     2161 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     2221 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     2281 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2341 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     2401 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     2461 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     2521 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     2581 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     2641 tctgatcagc acgtgatgaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg
     2701 atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt
     2761 gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat
     2821 ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg
     2881 gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca
     2941 cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc
     3001 gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca
     3061 ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct
     3121 gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg
     3181 caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg
     3241 cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt
     3301 gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg
     3361 aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag
     3421 cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat
     3481 cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca
     3541 atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc
     3601 gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact
     3661 cgtccgaggg caaaggaata gcacgtgcta cgagatttcg attccaccgc cgccttctat
     3721 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     3781 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     3841 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3901 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     3961 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     4021 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     4081 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     4141 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4201 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4261 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4321 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4381 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4441 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4501 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4561 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4621 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4681 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4741 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4801 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     4861 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4921 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4981 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5041 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5101 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5161 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5221 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5281 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5341 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5401 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5461 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5521 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5581 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5641 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5701 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5761 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5821 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5881 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5941 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6001 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     6061 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6121 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
     6181 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
     6241 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaag
//
