LOCUS       sequence-261484-        7594 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  lentiviral expression vector with Ef1a promoter, N-terminal V5 tag, 
            EGFP gene, SV40 promoter, and ZeoResistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7594)
  TITLE     Barrett lab unpublished plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7594)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pSMPUW_6.17_Ex_Ef1a-v5-EGFP-SV40-Zeo
FEATURES             Location/Qualifiers
     source          1..7594
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             522..702
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    749..873
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    908..1024
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1158..2336
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1389..2327
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2284..2304
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      2391..2400
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2400..2441
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     protein_bind    2442..2466
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(2469..2471,2472..2474,2475..3188)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2514..2535)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2775..2794)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3122..3143
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(3195..3219)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        3312..3522
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3334..3530
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3381..3516
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      3567..3576
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3573..3947
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     misc_feature    4020..4608
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4073..4093)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4491..4502)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4798..4978
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        5047..5257
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5116..5251
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5367..5488
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     5458..5477
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5540..5557)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5711..6299)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5791..5810)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(6551..6590)
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     CDS             complement(6600..7415)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(6600..7415)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     7323..7342
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     7407..7426
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgggtctctc tggttagacc
      541 agatttgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
      601 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
      661 gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc cgaacaggga
      721 cctgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc ttgctgaagc
      781 gcgcacggca agaggcgagg ggcggcgact gcagagtacg ccaaaatttt gactagcgga
      841 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggaa aatagcggcc
      901 gccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag
      961 acataatagc aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa
     1021 ttttcggggg atccgcggaa ttcgtccacg gtccaatctc ggtccgaacg agatatctgc
     1081 aaagatggat aaagttttaa acagagagga atctttgcag ctaatggacc ttctaggtct
     1141 tgaaaggagt gggaattggc tccggtgccc gtcagtgggc agagcgcaca tcgcccacag
     1201 tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg
     1261 gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag
     1321 aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg tttgccgcca
     1381 gaacacaggt aagtgccgtg tgtggttccc gcgggcctgg cctctttacg ggttatggcc
     1441 cttgcgtgcc ttgaattact tccacctggc tgcagtacgt gattcttgat cccgagcttc
     1501 gggttggaag tgggtgggag agttcgaggc cttgcgctta aggagcccct tcgcctcgtg
     1561 cttgagttga ggcctggcct gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc
     1621 gcgcctgtct cgctgctttc gataagtctc tagccattta aaatttttga tgacctgctg
     1681 cgacgctttt tttctggcaa gatagtcttg taaatgcggg ccaagatctg cacactggta
     1741 tttcggtttt tggggccgcg ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg
     1801 cgaggcgggg cctgcgagcg cggccaccga gaatcggacg ggggtagtct caagctggcc
     1861 ggcctgctct ggtgcctggc ctcgcgccgc cgtgtatcgc cccgccctgg gcggcaaggc
     1921 tggcccggtc ggcaccagtt gcgtgagcgg aaagatggcc gcttcccggc cctgctgcag
     1981 ggagctcaaa atggaggacg cggcgctcgg gagagcgggc gggtgagtca cccacacaaa
     2041 ggaaaagggc ctttccgtcc tcagccgtcg cttcatgtga ctccacggag taccgggcgc
     2101 cgtccaggca cctcgattag ttctcgagct tttggagtac gtcgtcttta ggttgggggg
     2161 aggggtttta tgcgatggag tttccccaca ctgagtgggt ggagactgaa gttaggccag
     2221 cttggcactt gatgtaattc tccttggaat ttgccctttt tgagtttgga tcttggttca
     2281 ttctcaagcc tcagacagtg gttcaaagtt tttttcttcc atttcaggtg tcgtgaggcg
     2341 cgcctatcga aattcactat agggagaccc aagctggcta gttaagcttc gccaccatgg
     2401 gtaagcctat ccctaaccct ctcctcggtc tcgattctac gacaagtttg tacaaaaaag
     2461 caggctccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     2521 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     2581 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     2641 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     2701 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     2761 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     2821 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     2881 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     2941 agaagaacgg catcaaggcc aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     3001 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     3061 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     3121 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     3181 acaagtagta atgaacccag ctttcttgta caaagtggta gtaatgacct taggagccaa
     3241 cagctaagca cacacttcga atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3301 caggctcccc agcaggcaga agtatgcaaa gcatgcgtct caattagtca gcaaccatag
     3361 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3421 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     3481 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctttcta
     3541 gaggtaccac cggagcttat ttaaatgcca ccatggccaa gttgaccagt gccgttccgg
     3601 tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg ctcgggttct
     3661 cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg accctgttca
     3721 tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg
     3781 gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct
     3841 ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc gccctgcgcg
     3901 acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactaatta attaacacaa
     3961 atacgcgtat ctgcagtcga ctttgccaga ccggagtccg aacacagctc ttccagcata
     4021 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     4081 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     4141 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     4201 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     4261 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     4321 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     4381 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     4441 cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     4501 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     4561 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgctt agtactggta
     4621 cctttaagac caatgactta caaggcagct gtagatctta gccacttttt aaaagaaaag
     4681 gggggactgg aagggctaat tcactcccaa cgaagacaag attccggaat ttatttgtga
     4741 aatttgtgat gctattgctt tatttgtaaa ccggtgcagc tgctttttgc ctgtactggg
     4801 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
     4861 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
     4921 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcatc
     4981 tagagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     5041 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     5101 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     5161 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     5221 cagaagtagt gaggaggctt ttttggaggc ctaggctaga gatcataatc agccatacca
     5281 catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac
     5341 ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat
     5401 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     5461 gtttgtccaa actcatcaat gtatcttatc atgtctgcta gctgagcggg cttttttttc
     5521 ttaggccttc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5581 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5641 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5701 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5761 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5821 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5881 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5941 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6001 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6061 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6121 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     6181 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6241 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6301 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6361 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6421 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6481 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6541 tcctgcgcag tccaaaaaaa aaggctccaa aaggagcctt taattgtatc ggtgggccct
     6601 tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata
     6661 ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat
     6721 aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct
     6781 attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact
     6841 gaatccggtg agaatggcaa aagcttatgc atttctttcc agacttgttc aacaggccag
     6901 ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc
     6961 gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa
     7021 tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat
     7081 tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca
     7141 tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt
     7201 agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac
     7261 aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca
     7321 ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc
     7381 ctcgagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg
     7441 taagcagaca gttttattgt tcatgatgat atatttttat cttgtgcaat gtaacatcag
     7501 agattttgag acacaacgtg gtttaaacaa atagtcaaaa gcctccgggc gactagtcgg
     7561 ggtcattagt tcatagccca tatatggagt tccg
//
