LOCUS       sequence-9854-ex        8482 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral EOS reporter with Oct3/4 enhancer (x3), expresses EGFP 
            and Puro resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8482)
  AUTHORS   Hotta A, Cheung AY, Farra N, Vijayaragavan K, Seguin CA, Draper JS, 
            Pasceri P, Maksakova IA, Mager DL, Rossant J, Bhatia M, Ellis J
  TITLE     Isolation of human iPS cells using EOS lentiviral vectors to select 
            for pluripotency.
  JOURNAL   Nat Methods. 2009 May . 6(5):370-6.
  PUBMED    19404254
REFERENCE   2  (bases 1 to 8482)
  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: "Nat 
            Methods. 2009 May . 6(5):370-6."
COMMENT     Sequence Label: PL-SIN-EOS-C(3+)-EiP
FEATURES             Location/Qualifiers
     source          1..8482
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             785..1042
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1693..1810
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    2200..2433
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     primer_bind     2762..2778
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      3675..3684
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3681..3683,3684..3686,3687..4400)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3681..3683,3684..3686,3687..4397)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3726..3747)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3987..4006)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4334..4355
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    join(4441..5015,5016..5018,5019..5027)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4608..4625)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4835..4854
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5028..5627
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5028..5047)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5524..5544
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5769..6002
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(6039..6061)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6050..6066
                     /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(6074..6080,6081..6098,6099..6104))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6119..6140
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6257..6274)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6428..7016)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6508..6527)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7485..7862)
                     /label=ISS
                     /note="immunostimulatory sequence from the AmpR gene; 
                     contains unmethylated CpG dinucleotides in the context of 
                     5'-AACGTT-3' (Sato et al., 1996)"
     primer_bind     7809..7828
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8047..8151)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        complement(8188..8384)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      8202..8337
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     8455..8473
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gcggccgctg gaagggctaa
      421 tttggtccca aaaaagacaa gagatccttg atctgtggat ctaccacaca caaggctact
      481 tccctgattg gcagaactac acaccagggc cagggatcag atatccactg acctttggat
      541 ggtgcttcaa gttagtacca gttgaaccag agcaagtaga agaggccaaa taaggagaga
      601 agaacagctt gttacaccct atgagccagc atgggatgga ggacccggag ggagaagtat
      661 tagtgtggaa gtttgacagc ctcctagcat ttcgtcacat ggcccgagag ctgcatccgg
      721 agtactacaa agactgctga catcgagctt tctacaaggg actttccgct ggggactttc
      781 cagggaggtg tggcctgggc gggactgggg agtggcgagc cctcagatgc tacatataag
      841 cagctgcttt ttgcctgtac tgggtctctc tggttagacc agatctgagc ctgggagctc
      901 tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgctcaaa
      961 gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag
     1021 tcagtgtgga aaatctctag cagtggcgcc cgaacaggga cttgaaagcg aaagtaaagc
     1081 cagaggagat ctctcgacgc aggactcggc ttgctgaagc gcgcacggca agaggcgagg
     1141 ggcggcgact ggtgagtacg ccaaaaattt tgactagcgg aggctagaag gagagagatg
     1201 ggtgcgagag cgtcggtatt aagcggggga gaattagata aatgggaaaa aattcggtta
     1261 aggccagggg gaaagaaaca atataaacta aaacatatag tatgggcaag cagggagcta
     1321 gaacgattcg cagttaatcc tggcctttta gagacatcag aaggctgtag acaaatactg
     1381 ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca
     1441 atagcagtcc tctattgtgt gcatcaaagg atagatgtaa aagacaccaa ggaagcctta
     1501 gataagatag aggaagagca aaacaaaagt aagaaaaagg cacagcaagc agcagctgac
     1561 acaggaaaca acagccaggt cagccaaaat taccctatag tgcagaacct ccaggggcaa
     1621 atggtacatc aggccatatc acctagaact ttaaattaag acagcagtac aaatggcagt
     1681 attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat
     1741 agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat
     1801 tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggaaag gaccagcaaa
     1861 gctcctctgg aaaggtgaag gggcagtagt aatacaagat aatagtgaca taaaagtagt
     1921 gccaagaaga aaagcaaaga tcatcaggga ttatggaaaa cagatggcag gtgatgattg
     1981 tgtggcaagt agacaggatg aggattaaca catggaaaag attagtaaaa caccatagct
     2041 ctagagcgat ccgatcttca gacctggagg aggcgatatg agggacaatt ggagaagtga
     2101 attatataaa tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa
     2161 gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt
     2221 cttgggagca gcaggaagca ctatgggctg cacgtcaatg acgctgacgg tacaggccag
     2281 acaattattg tctgatatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca
     2341 acagcatctg ttgcaactca cagtctgggg catcaaacag ctccaggcaa gaatcctggc
     2401 tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact
     2461 catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat
     2521 ttggaataac atgacctgga tggagtggga cagagaaatt aacaattaca caagcttggt
     2581 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2641 accattatcg tttcagaccc acctcccaat cccgagggga cccgacaggc ccgaaggaat
     2701 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcctc
     2761 cctttatcca gccctcactc cttctctagg cgccggaatt cgggtgtggg gaggttgtag
     2821 cccgaccctg cccctccccc cagggaggtt gagagttctg ggcagacggc agatgcataa
     2881 caaaggtgca tgatagctct gccctggggg cagagaagat ggttggggag gggtccctct
     2941 cgtcctagct cgagcttaat cactagtgaa ttcgggtgtg gggaggttgt agcccgaccc
     3001 tgcccctccc cccagggagg ttgagagttc tgggcagacg gcagatgcat aacaaaggtg
     3061 catgatagct ctgccctggg ggcagagaag atggttgggg aggggtccct ctcgtcctag
     3121 ctcgagctta atcactagtg aattcgggtg tggggaggtt gtagcccgac cctgcccctc
     3181 cccccaggga ggttgagagt tctgggcaga cggcagatgc ataacaaagg tgcatgatag
     3241 ctctgccctg ggggcagaga agatggttgg ggaggggtcc ctctcgtcct agctcgagct
     3301 taatcactag tgaattctcc cctcccccag cctgaaacct gcttgctcag gggtggagct
     3361 tcctgctcat tcgttctgcc acgcccactg ctggaacctg cggagccaca cccgtgcacc
     3421 tttctactgg accagagatt attcggcggg aatcgggtcc cctccccctt ccttcataac
     3481 tagtgtcgca actataaaat ttgagccttg atcagagtaa ctgtcttggc tacattcttt
     3541 cttccgcccc gtctagattc ctctcttaaa gctcgagcgg ccttctcagt cgaaccgttc
     3601 acgttgcgag ctgctggcgg ccgcaacaag atctgcgatc taagtaagct tggcattccg
     3661 gtactgttgg taaagccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     3721 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     3781 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     3841 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     3901 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     3961 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     4021 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     4081 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     4141 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     4201 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     4261 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     4321 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     4381 tggacgagct gtacaagtaa agcggccaac ctcgacgtta ttcccttcga tcgaattccg
     4441 cccctctccc tccccccccc ctaacgttac tggccgaagc cgcttggaat aaggccggtg
     4501 tgcgtttgtc tatatgttat tttccaccat attgccgtct tttggcaatg tgagggcccg
     4561 gaaacctggc cctgtcttct tgacgagcat tcctaggggt ctttcccctc tcgccaaagg
     4621 aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct ctggaagctt cttgaagaca
     4681 aacaacgtct gtagcgaccc tttgcaggca gcggaacccc ccacctggcg acaggtgcct
     4741 ctgcggccaa aagccacgtg tataagatac acctgcaaag gcggcacaac cccagtgcca
     4801 cgttgtgagt tggatagttg tggaaagagt caaatggctc tcctcaagcg tattcaacaa
     4861 ggggctgaag gatgcccaga aggtacccca ttgtatggga tctgatctgg ggcctcggtg
     4921 cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc cgaaccacgg
     4981 ggacgtggtt ttcctttgaa aaacacgatg ataatatggc cacaaccatg accgagtaca
     5041 agcccacggt gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg
     5101 ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc
     5161 gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt
     5221 gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac gccggagagc gtcgaagcgg
     5281 gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg
     5341 cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc
     5401 tggccaccgt cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc
     5461 tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acctccgcgc
     5521 cccgcaacct ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc
     5581 ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg tgcctgacgc ccgccccacg
     5641 acccgcagcg cccgaccgaa aggagcgcac gaccccatgc atcgatttcg aacccggggt
     5701 acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa
     5761 ggggggactg gaagggctaa ttcactccca aagaagacaa gatctgcttt ttgcctgtac
     5821 tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc
     5881 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt
     5941 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag
     6001 cagaattcta gcttggcgta atcatggtca tagctgttcc tgtgtgaaat tgttatccgc
     6061 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     6121 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     6181 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     6241 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     6301 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     6361 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     6421 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     6481 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     6541 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6601 cgggaagcgt ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg
     6661 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6721 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     6781 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6841 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     6901 cacttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6961 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     7021 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     7081 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     7141 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     7201 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     7261 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     7321 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     7381 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     7441 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gtgccattgc
     7501 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     7561 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     7621 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     7681 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     7741 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     7801 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     7861 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     7921 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     7981 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     8041 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     8101 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     8161 ccgaaaagtg ccacctggga ctagcttttt gcaaaagcct aggcctccaa aaaagcctcc
     8221 tcactacttc tggaatagct cagaggccga ggcggcctcg gcctctgcat aaataaaaaa
     8281 aattagtcag ccatggggcg gagaatgggc ggaactgggc ggagttaggg gcgggatggg
     8341 cggagttagg ggcgggacta tggttgctga ctaattgaga tgtagcttgc atgccgacat
     8401 ggattattga ctagtcccta agaaaccatt attatcatga cattaaccta taaaaatagg
     8461 cgtatcacga ggccctttcg tc
//
