LOCUS       sequence-256207-        7579 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  lentiviral expression vector with Ef1a promoter, N-terminal HA tag, 
            EGFP gene, SV40 promoter, and Zeo Resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7579)
  TITLE     Barrett lab unpublished plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7579)
  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            "Unpublished"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_6.18_Ex_Ef1a-HA-EGFP-SV40-Zeo
FEATURES             Location/Qualifiers
     source          1..7579
                     /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"
     CDS             2400..2426
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    2427..2451
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(2454..2456,2457..2459,2460..3173)
                     /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(2499..2520)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2760..2779)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3107..3128
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(3180..3204)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        3297..3507
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3319..3515
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3366..3501
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      3552..3561
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3558..3932
                     /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    4005..4593
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4058..4078)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4476..4487)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4783..4963
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        5032..5242
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5101..5236
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5352..5473
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     5443..5462
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5525..5542)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5696..6284)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5776..5795)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(6536..6575)
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     CDS             complement(6585..7400)
                     /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(6585..7400)
                     /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     7308..7327
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     7392..7411
                     /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 gccaccatgt
     2401 acccatacga tgttccagat tacgctacaa gtttgtacaa aaaagcaggc tccatggtga
     2461 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     2521 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     2581 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     2641 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     2701 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     2761 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     2821 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     2881 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
     2941 aggccaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
     3001 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     3061 gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     3121 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag tagtaatgaa
     3181 cccagctttc ttgtacaaag tggtagtaat gaccttagga gccaacagct aagcacacac
     3241 ttcgaatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3301 gcagaagtat gcaaagcatg cgtctcaatt agtcagcaac catagtcccg cccctaactc
     3361 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3421 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3481 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct ttctagaggt accaccggag
     3541 cttatttaaa tgccaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg
     3601 acgtcgccgg agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg
     3661 aggacgactt cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg
     3721 accaggtggt gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt
     3781 acgccgagtg gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga
     3841 ccgagatcgg cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact
     3901 gcgtgcactt cgtggccgag gagcaggact aattaattaa cacaaatacg cgtatctgca
     3961 gtcgactttg ccagaccgga gtccgaacac agctcttcca gcataatcaa cctctggatt
     4021 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     4081 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     4141 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     4201 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     4261 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     4321 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     4381 ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct
     4441 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     4501 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     4561 cgagtcggat ctccctttgg gccgcctccc cgcttagtac tggtaccttt aagaccaatg
     4621 acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg
     4681 ctaattcact cccaacgaag acaagattcc ggaatttatt tgtgaaattt gtgatgctat
     4741 tgctttattt gtaaaccggt gcagctgctt tttgcctgta ctgggtctct ctggttagac
     4801 cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa
     4861 agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag
     4921 agatccctca gaccctttta gtcagtgtgg aaaatctcta gcatctagag tatgcaaagc
     4981 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     5041 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     5101 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     5161 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     5221 ggcttttttg gaggcctagg ctagagatca taatcagcca taccacattt gtagaggttt
     5281 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     5341 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     5401 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     5461 tcaatgtatc ttatcatgtc tgctagctga gcgggctttt ttttcttagg ccttcttccg
     5521 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     5581 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5641 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5701 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5761 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5821 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5881 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgcaccaagc
     5941 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     6001 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     6061 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     6121 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     6181 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     6241 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     6301 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     6361 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     6421 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6481 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcctg cgcagtccaa
     6541 aaaaaaaggc tccaaaagga gcctttaatt gtatcggtgg gcccttagaa aaactcatcg
     6601 agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa
     6661 agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc
     6721 tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg
     6781 tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat
     6841 ggcaaaagct tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca
     6901 tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga
     6961 aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg
     7021 aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg
     7081 aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata
     7141 aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca
     7201 tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg
     7261 ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat
     7321 ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctcga gcaagacgtt
     7381 tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt
     7441 attgttcatg atgatatatt tttatcttgt gcaatgtaac atcagagatt ttgagacaca
     7501 acgtggttta aacaaatagt caaaagcctc cgggcgacta gtcggggtca ttagttcata
     7561 gcccatatat ggagttccg
//
