LOCUS       sequence-291815-        7486 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentivector that expresses LifeAct-GFP from internal human PGK 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7486)
  TITLE     Lentivirus
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7486)
  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            "Unpublished"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: pLenti.PGK.LifeAct-GFP.W
FEATURES             Location/Qualifiers
     source          1..7486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        8..234
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             235..415
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    462..587
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1080..1313
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1498..1542
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    1809..1926
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1972..2482
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     2401..2419
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             2533..2583
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     regulatory      2599..2608
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2605..2607,2608..2610,2611..3324)
                     /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(2605..2607,2608..2610,2611..3321)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2650..2671)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2911..2930)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3258..3279
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3337..3925
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3390..3410)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3808..3819)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4012..4245
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4317..4438
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4354..4373)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4408..4427
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4478..4613
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4633..4652)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4634..4652)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4662..4679)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4662..4678)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4671..4693)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4820..5275
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4907..4926)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5117..5138
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5301..5405
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5406..5474,5475..6266)
                     /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     complement(5624..5643)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6437..7025
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6926..6945
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7179..7196
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7313..7334
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7349..7354,7355..7372,7373..7379)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7387..7403
                     /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)."
     primer_bind     7392..7414
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7411..7427
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7411..7427
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7446..7466
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7448..7466
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 agcttaatgt agtcttatgc aatactcttg tagtcttgca acatggtaac gatgagttag
       61 caacatgcct tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg
      121 tacgatcgtg ccttattagg aaggcaacag acgggtctga catggattgg acgaaccact
      181 gaattgccgc attgcagaga tattgtattt aagtgcctag ctcgatacat aaacgggtct
      241 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      301 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      361 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
      421 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
      481 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
      541 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
      601 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
      661 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
      721 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
      781 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
      841 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
      901 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
      961 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1021 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1081 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1141 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1201 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1261 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1321 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1381 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     1441 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     1501 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     1561 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     1621 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     1681 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     1741 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat
     1801 cggttaactt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga aagaatagta
     1861 gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac aaaaattcaa
     1921 aattttatcg atcacgagac tagcctcgag aagcttgata tcgaattcca cggggttggg
     1981 gttgcgcctt ttccaaggca gccctgggtt tgcgcaggga cgcggctgct ctgggcgtgg
     2041 ttccgggaaa cgcagcggcg ccgaccctgg gtctcgcaca ttcttcacgt ccgttcgcag
     2101 cgtcacccgg atcttcgccg ctacccttgt gggccccccg gcgacgcttc ctgctccgcc
     2161 cctaagtcgg gaaggttcct tgcggttcgc ggcgtgccgg acgtgacaaa cggaagccgc
     2221 acgtctcact agtaccctcg cagacggaca gcgccaggga gcaatggcag cgcgccgacc
     2281 gcgatgggct gtggccaata gcggctgctc agcagggcgc gccgagagca gcggccggga
     2341 aggggcggtg cgggaggcgg ggtgtggggc ggtagtgtgg gccctgttcc tgcccgcgcg
     2401 gtgttccgca ttctgcaagc ctccggagcg cacgtcggca gtcggctccc tcgttgaccg
     2461 aatcaccgac ctctctcccc agggggatcc tagcgctacc ggactcagat ctcgagctca
     2521 agcttcgaat tcatgggtgt cgcagatttg atcaagaaat tcgaaagcat ctcaaaggaa
     2581 gaaggggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg
     2641 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     2701 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     2761 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
     2821 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     2881 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     2941 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     3001 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     3061 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     3121 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     3181 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc
     3241 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     3301 ggcatggacg agctgtacaa gtaaagcggc ctcgacaatc aacctctgga ttacaaaatt
     3361 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     3421 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     3481 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     3541 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     3601 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     3661 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     3721 ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg
     3781 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     3841 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
     3901 atctcccttt gggccgcctc cccgcctgga attcgagctc ggtaccttta agaccaatga
     3961 cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc
     4021 taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag
     4081 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
     4141 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
     4201 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc
     4261 atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact
     4321 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4381 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4441 atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     4501 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     4561 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac
     4621 gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa
     4681 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     4741 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     4801 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4861 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4921 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4981 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     5041 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     5101 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     5161 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5221 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     5281 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5341 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5401 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5461 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5521 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5581 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5641 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5701 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5761 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5821 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5881 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5941 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     6001 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     6061 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     6121 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6181 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6241 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     6301 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     6361 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     6421 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     6481 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     6541 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6601 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6661 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6721 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6781 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6841 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6901 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6961 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     7021 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     7081 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     7141 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     7201 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     7261 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     7321 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     7381 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     7441 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgca
//
