LOCUS       sequence-150307-        6101 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of I3-01.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6101)
  AUTHORS   Votteler J, Ogohara C, Yi S, Hsia Y, Nattermann U, Belnap DM, King 
            NP, Sundquist WI
  TITLE     Designed proteins induce the formation of nanocage-containing 
            extracellular vesicles.
  JOURNAL   Nature. 2016 Dec 8;540(7632):292-295. doi: 10.1038/nature20607. Epub
            2016 Nov 30.
  PUBMED    27919066
REFERENCE   2  (bases 1 to 6101)
  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"; doi: 
            "10.1038/nature20607"; journalName: "Nature"; date: "2016-12-8- 8"; 
            volume: "540"; issue: "7632"; pages: "292-295"
COMMENT     Sequence Label: pCMV-I3-01-Myc
FEATURES             Location/Qualifiers
     source          1..6101
                     /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             1564..1593
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1618..1637)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     primer_bind     complement(1692..1709)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1698..1922
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1968..2396
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2055..2074)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2265..2286
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2405..2425)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2410..2739
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2590..2725
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2652..2671
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2806..3600
                     /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(2860..2879)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3470..3489
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3774..3895
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3811..3830)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3865..3884
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3944..3960)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3944..3960)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3957..3979)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3968..3984
                     /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(3992..3998,3999..4016,4017..4022))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4037..4058
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4175..4192)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4346..4934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4426..4445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5105..5896,5897..5965))
                     /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     5728..5747
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5966..6070)
                     /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 gtttaaacgg gccctctaga attatcgatc cggaggtacc cgggcggccg catgaaaatg
      961 gaagagctgt tcaagaaaca caagatcgtt gccgtgctgc gtgccaatag tgtggaagaa
     1021 gcgaaaaaga aagcgctggc ggttttcctg ggcggcgttc atctgattga aattaccttt
     1081 accgtgccgg atgcggatac cgtgattaag gaactgagct ttctgaagga aatgggcgcg
     1141 attattggtg cgggcaccgt gaccagcgtg gagcagtgcc gtaaagcggt ggaaagtggc
     1201 gccgaattca ttgtgagtcc gcacctggac gaggaaatta gccaattttg caaggagaag
     1261 ggtgtgttct atatgccagg cgttatgacc ccgaccgaac tggtgaaagc catgaaactg
     1321 ggccatacca tcttaaaact gtttccgggt gaggtggtgg gtccgcagtt tgttaaagcg
     1381 atgaaaggtc cgtttccgaa tgtgaaattt gtgccaaccg gcggtgttaa tctggacaat
     1441 gtgtgcgaat ggttcaaagc gggcgtgctg gccgtgggcg tgggcagcgc gttagtgaaa
     1501 ggcaccccgg tggaagtggc ggaaaaggcc aaggcgttcg ttgagaagat tcgtggctgc
     1561 accgaacaga aactgattag cgaagaagat ctgtaactcg agaagcttgc tagcagatct
     1621 ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcact taagtttaaa
     1681 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     1741 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     1801 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     1861 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     1921 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     1981 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2041 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     2101 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     2161 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     2221 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     2281 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     2341 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     2401 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     2461 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     2521 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     2581 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     2641 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     2701 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     2761 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     2821 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     2881 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     2941 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     3001 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     3061 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     3121 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     3181 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     3241 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     3301 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     3361 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     3421 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     3481 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     3541 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     3601 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     3661 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     3721 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     3781 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     3841 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     3901 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     3961 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     4021 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     4081 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4141 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4201 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4261 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4321 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4381 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4441 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4501 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4561 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4621 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4681 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4741 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     4801 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4861 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4921 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4981 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5041 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5101 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5161 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5221 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5281 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5341 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5401 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5461 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5521 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5581 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5641 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5701 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5761 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5821 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5881 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5941 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6001 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6061 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
