LOCUS       sequence-273897-        7810 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentivirus reporter assay plasmid that contains a I-SceI site and a 
            EGFP reporter gene..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7810)
  AUTHORS   Gordon MG, Inoue F, Martin B, Schubach M, Agarwal V, Whalen S, Feng 
            S, Zhao J, Ashuach T, Ziffra R, Kreimer A, Georgakopoulous-Soares I,
            Yosef N, Ye CJ, Pollard KS, Shendure J, Kircher M, Ahituv N
  TITLE     lentiMPRA and MPRAflow for high-throughput functional 
            characterization of gene regulatory elements.
  JOURNAL   Nat Protoc. 2020 Aug;15(8):2387-2412. doi: 
            10.1038/s41596-020-0333-5. Epub 2020 Jul 8.
  PUBMED    32641802
REFERENCE   2  (bases 1 to 7810)
  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/s41596-020-0333-5"; journalName: "Nat Protoc"; date: 
            "2020-08"; volume: "15"; issue: "8"; pages: "2387-2412"
COMMENT     Sequence Label: pLS-SceI
FEATURES             Location/Qualifiers
     source          1..7810
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1447..1680
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1865..1909
                     /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    2207..2324
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     regulatory      2828..2837
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2834..2836,2837..2839,2840..3553)
                     /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(2834..2836,2837..2839,2840..3550)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2879..2900)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3140..3159)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3487..3508
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(3573..3606)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3662..4250
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3715..3735)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4133..4144)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4253..4269)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4254..4270)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5579..5759
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5821..6409)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5901..5920)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6580..7371,7372..7440))
                     /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     7203..7222
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7441..7545)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7620..7639)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg
     1321 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1381 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1441 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1501 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1561 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1621 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1681 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     1741 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     1801 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     1861 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     1921 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     1981 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2041 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     2101 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc
     2161 actgcgtgcg ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag
     2221 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2281 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2341 acagcagaga tccagtttgg ttagtaccgg gcccggtgct ttgctctgag ccagcccacc
     2401 agtttggaat gactcctttt tatgacttga attttcaagt ataaagtcta gtgctaaatt
     2461 taatttgaac aactgtatag tttttgctgg ttgggggaag gaaaaaaaat ggtggcagtg
     2521 tttttttcag aattagaagt gaaatgaaaa cttgttgtgt gtgaggattt ctaatgacat
     2581 gtggtggttg catactgagt gaagccggtg agcattctgc catgtcaccc cctcgtgctc
     2641 agtaatgtac tttacagaaa tcctaaactc aaaagattga tataaaccat gcttcttgtg
     2701 tatatccggt ctcttctctg ggtagtctca ctcagcctgc atttctgcca gggcccgctc
     2761 tagacctgca gggcatctaa tgccacttag ggataacagg gtaatgctat ccaggttaac
     2821 accggtcgcc accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     2881 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     2941 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     3001 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     3061 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     3121 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     3181 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     3241 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     3301 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     3361 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     3421 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     3481 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     3541 gctgtacaag taggaattcg tcgagggacc taataacttc gtatagcata cattatacga
     3601 agttatacat gtttaagggt tccggttcca ctaggtacaa ttcgatatca agcttatcga
     3661 taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     3721 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     3781 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     3841 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     3901 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     3961 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     4021 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct
     4081 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     4141 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     4201 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg
     4261 tcgacctcga tcgagaccta gaaaaacatg gagcaatcac aagtagcaat acagcagcta
     4321 ccaatgctga ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt tttccagtca
     4381 cacctcaggt accaagcatg gggtaaagta ctgttctcat cacatcatat caaggttata
     4441 taccatcaat attgccacag atgttactta gccttttaat atttctctaa tttagtgtat
     4501 atgcaatgat agttctctga tttctgagat tgagtttctc atgtgtaatg attatttaga
     4561 gtttctcttt catctgttca aatttttgtc tagttttatt ttttactgat ttgtaagact
     4621 tctttttata atctgcatat tacaattctc tttactgggg tgttgcaaat attttctgtc
     4681 attctatggc ctgacttttc ttaatggttt tttaatttta aaaataagtc ttaatattca
     4741 tgcaatctaa ttaacaatct tttctttgtg gttaggactt tgagtcataa gaaatttttc
     4801 tctacactga agtcatgatg gcatgcttct atattatttt ctaaaagatt taaagttttg
     4861 ccttctccat ttagacttat aattcactgg aatttttttg tgtgtatggt atgacatatg
     4921 ggttcccttt tattttttac atataaatat atttccctgt ttttctaaaa aagaaaaaga
     4981 tcatcatttt cccattgtaa aatgccatat ttttttcata ggtcacttac atatatcaat
     5041 gggtctgttt ctgagctcta ctctatttta tcagcctcac tgtctatccc cacacatctc
     5101 atgctttgct ctaaatcttg atatttagtg gaacattctt tcccattttg ttctacaaga
     5161 atatttttgt tattgtcttt tgggcttcta tatacatttt agaatgaggt tggcaagttc
     5221 tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca
     5281 acgaagacaa gatatccttg atctgtggat ctaccacaca caaggctact tccctgattg
     5341 gcagaactac acaccagggc cagggatcag atatccactg acctttggat ggtgctacaa
     5401 gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga acacccgctt
     5461 gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat tagagtggag
     5521 gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg actgtactgg
     5581 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
     5641 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
     5701 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
     5761 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5821 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5881 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5941 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     6001 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6061 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6121 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6181 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6241 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6301 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6361 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6421 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6481 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6541 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6601 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6661 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6721 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6781 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6841 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6901 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6961 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     7021 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7081 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7141 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7201 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7261 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7321 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7381 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7441 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7501 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7561 agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt
     7621 acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag
     7681 gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat
     7741 tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga
     7801 tatacgcgtt
//
