LOCUS       sequence-466789-        7716 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For production of Extracellular Vesicles (EVs), with a Strep-tag II 
            labeled Sodium-iodide symporter (NIS), and an HA epitope at its 
            N-terminus on their surface.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7716)
  AUTHORS   Gonzalez-Magaldi M, Gullapalli A, Papoulas O, Liu C, Leung AY, Guo 
            L, Brilot A, Marcotte EM, Ke Z, Leahy DJ
  TITLE     Structure and organization of full-length Epidermal Growth Factor 
            Receptor in extracellular vesicles by cryo-electron tomography.
  JOURNAL   bioRxiv [Preprint]. 2024 Nov 28:2024.11.25.625301. doi: 
            10.1101/2024.11.25.625301.
  PUBMED    39651119
REFERENCE   2  (bases 1 to 7716)
  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: "bioRxiv 
            [Preprint]. 2024 Nov 28:2024.11.25.625301. doi: 
            10.1101/2024.11.25.625301."
COMMENT     Sequence Label: pHA StreptagII 3C NIS EABR
FEATURES             Location/Qualifiers
     source          1..7716
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /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     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1211..1230)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1402..1781
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1782..1985
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1936..1956
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2030..2049
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2030..2048
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2097..2123
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2133..2156
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(4474..4491)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4480..4704
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4750..5178
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4837..4856)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5047..5068
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5187..5207)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5192..5521
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5372..5507
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5434..5453
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5588..6382
                     /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(5642..5661)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6252..6271
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6556..6677
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6593..6612)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6647..6666
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6726..6742)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6726..6742)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6739..6761)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6750..6766
                     /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(6774..6780,6781..6798,6799..6804))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6819..6840
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6957..6974)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7128..7716)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7208..7227)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     1201 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     1261 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
     1321 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
     1381 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
     1441 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
     1501 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
     1561 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
     1621 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
     1681 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
     1741 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
     1801 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
     1861 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
     1921 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
     1981 gagctctctg gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc
     2041 actataggga gacccaagct ggctagcgtt taaacgggcc ctctagagcc accatgtacc
     2101 catacgatgt tccagattac gcttccggat cttggagcca cccgcagttc gaaaagggca
     2161 gcgaggccgt ggagaccggg gaacggccca ccttcggagc ctgggactac ggggtctttg
     2221 ccctcatgct cctggtgtcc actggcatcg ggctgtgggt cgggctggct cggggcgggc
     2281 agcgcagcgc tgaggacttc ttcaccgggg gccggcgcct ggcggccctg cccgtgggcc
     2341 tgtcgctgtc tgccagcttc atgtcggccg tgcaggtgct gggcgtgccg tcggaggcct
     2401 atcgctatgg cctcaagttc ctctggatgt gcctgggcca gcttctgaac tcggtcctca
     2461 ccgccctgct cttcatgccc gtcttctacc gcctgggcct caccagcacc tacgagtacc
     2521 tggagatgcg cttcagccgc gcagtgcggc tctgcgggac tttgcagtac attgtagcca
     2581 cgatgctgta caccggcatc gtaatctacg caccggccct catcctgaac caagtgaccg
     2641 ggctggacat ctgggcgtcg ctcctgtcca ccggaattat ctgcaccttc tacacggctg
     2701 tgggcggcat gaaggctgtg gtctggactg atgtgttcca ggtcgtggtg atgctaagtg
     2761 gcttctgggt tgtcctggca cgcggtgtca tgcttgtggg cgggccccgc caggtgctca
     2821 cgctggccca gaaccactcc cggatcaacc tcatggactt taaccctgac ccgaggagcc
     2881 gctatacatt ctggactttt gtggtgggtg gcacgttggt gtggctctcc atgtatggcg
     2941 tgaaccaggc gcaggtgcag cgctacgtgg cttgccgcac agagaagcag gccaagctgg
     3001 ccctgctcat caaccaggtc ggcctgttcc tgatcgtgtc cagcgccgcc tgctgtggca
     3061 tcgtcatgtt tgtgttctac actgactgcg accctctcct cctggggcgc atctctgccc
     3121 cagaccagta catgcctctg ctggtgctgg acatcttcga agatctgcct ggagtccccg
     3181 ggcttttcct ggcctgtgct tacagtggca ccctcagcac agcatccacc agcatcaatg
     3241 ctatggctgc agtcactgta gaagacctca tcaaacctcg gctgcggagc ctggcaccca
     3301 ggaaactcgt gattatctcc aaggggctct cactcatcta cggatcggcc tgtctcaccg
     3361 tggcagccct gtcctcactg ctcggaggag gtgtccttca gggctccttc accgtcatgg
     3421 gagtcatcag cggccccctg ctgggagcct tcatcttggg aatgttcctg ccggcctgca
     3481 acacaccggg cgtcctcgcg ggactaggcg cgggcttggc gctgtcgctg tgggtggcct
     3541 tgggcgccac gctgtaccca cccagcgagc agaccatgag ggtcctgcca tcgtcggctg
     3601 cccgctgcgt ggctctctca gtcaacgcct ctggcctcct ggacccggct ctcctccctg
     3661 ctaacgactc cagcagggcc cccagctcag gaatggacgc cagccgaccc gccttagctg
     3721 acagcttcta tgccatctcc tatctctatt acggtgccct gggcacgctg accactgtgc
     3781 tgtgcggagc cctcatcagc tgcctgacag gccccaccaa gcgcagcacc ctggccccgg
     3841 gattgttgtg gtgggacctc gcacggcaga cagcatcagt ggcccccaag gaagaagtgg
     3901 ccatcctgga tgacaacttg gtcaagggtc ctgaagaact ccccactgga aacaagaagc
     3961 cccctggctt cctgcccacc aatgaggatc gtctgttttt cttggggcag aaggagctgg
     4021 agggggctgg ctcttggacc ccctgtgttg gacatgatgg tggtcgagac cagcaggaga
     4081 caaacctcgc tttgcctggc aatcccgatc acagggaaat gggggaaact ctcccagaag
     4141 aggttggaga atatcggcag ccatctggtg gcagtgtacc tgtgtctccg ggtccgccca
     4201 gtggtcttga acccactagc tcctcacctt acggcggcgg ctcctttaac agctcaatca
     4261 acaacatcca cgaaatggag attcagctga aggatgcact cgaaaagaat caacaatggc
     4321 tcgtttacga ccaacaacgc gaggtgtacg tgaaaggatt gcttgcaaaa attttcgaac
     4381 tggaaaagaa aacagagact gcagcacata gtcttccttg aggacagggc cagacctgca
     4441 ggcatgcaag cttaagttta aaccgctgat cagcctcgac tgtgccttct agttgccagc
     4501 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     4561 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     4621 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     4681 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     4741 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4801 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     4861 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4921 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     4981 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     5041 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     5101 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     5161 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     5221 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     5281 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     5341 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     5401 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     5461 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     5521 aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc
     5581 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     5641 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     5701 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     5761 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     5821 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     5881 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     5941 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     6001 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     6061 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     6121 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     6181 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     6241 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     6301 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     6361 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     6421 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     6481 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     6541 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     6601 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     6661 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     6721 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     6781 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     6841 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     6901 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     6961 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     7021 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     7081 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     7141 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     7201 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     7261 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     7321 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     7381 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     7441 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     7501 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     7561 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     7621 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     7681 aagcagcaga ttacgcgcag aaaaaaagga tctcaa
//
