LOCUS       sequence-274323-        8083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression, Lentiviral. Expression of human 
            phosphofructokinase, liver isoform PFKL-GFP fusion protein .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8083)
  AUTHORS   Park JS, Burckhardt CJ, Lazcano R, Solis LM, Isogai T, Li L, Chen 
            CS, Gao B, Minna JD, Bachoo R, DeBerardinis RJ, Danuser G
  TITLE     Mechanical regulation of glycolysis via cytoskeleton architecture.
  JOURNAL   Nature. 2020 Feb 12. pii: 10.1038/s41586-020-1998-1. doi: 
            10.1038/s41586-020-1998-1.
  PUBMED    32051585
REFERENCE   2  (bases 1 to 8083)
  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: "Nature. 
            2020 Feb 12. pii: 10.1038/s41586-020-1998-1. doi: 
            10.1038/s41586-020-1998-1."
COMMENT     Sequence Label: pLVX neo PFKL-GFP
FEATURES             Location/Qualifiers
     source          1..8083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1900..1909
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2647..2656
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2653..2655,2656..2658,2659..3372)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2698..2719)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2959..2978)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3306..3327
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    join(3390..3964,3965..3967,3968..3976)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3557..3574)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3784..3803
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3977..4771
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MAEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4031..4050)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4641..4660
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     misc_feature    4785..5373
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4838..4858)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5256..5267)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5580..6213
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    6260..6385
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6882..7115
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7300..7344
                     /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    7607..7722
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     enhancer        7780..8083
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg
      241 actcagatct cgagtttcga gatggccgcg gtggacctgg agaagctgcg ggcgtcgggc
      301 gcgggcaagg ccatcggcgt cctgaccagc ggcggcgacg cgcaaggcat gaacgctgct
      361 gtccgggctg tgacgcgcat gggcatttat gtgggtgcca aagtcttcct catctacgag
      421 ggctatgagg gcctcgtgga gggaggtgag aacatcaagc aggccaactg gctgagcgtc
      481 tccaacatca tccagctggg cggcactatc attggcagcg ctcgctgcaa ggcctttacc
      541 accagggagg ggcgccgggc agcggcctac aacctggtcc agcacggcat caccaacctg
      601 tgcgtcatcg gcggggatgg cagccttaca ggtgccaaca tcttccgcag cgagtggggc
      661 agcctgctgg aggagctggt ggcggaaggt aagatctcag agactacagc ccggacctac
      721 tcgcacctga acatcgcggg cctagtgggc tccatcgata acgacttctg cggcaccgac
      781 atgaccatcg gcacggactc ggccctccac cgcatcatgg aggtcatcga tgccatcacc
      841 accactgccc agagccacca gaggaccttc gtgctggaag tgatgggccg gcactgcggg
      901 tacctggcgc tggtatctgc actggcctca ggggccgact ggctgttcat ccccgaggct
      961 ccacccgagg acggctggga gaacttcatg tgtgagaggc tgggtgagac tcggagccgt
     1021 gggtcccgac tgaacatcat catcatcgct gagggtgcca ttgaccgcaa cgggaagccc
     1081 atctcgtcca gctacgtgaa ggacctggtg gttcagaggc tgggcttcga cacccgtgta
     1141 actgtgctgg gccacgtgca gcggggaggg acgccctctg ccttcgaccg gatcctgagc
     1201 agcaagatgg gcatggaggc ggtgatggcg ctgctggaag ccacgcctga cacgccggcc
     1261 tgcgtggtca ccctctcggg gaaccagtca gtgcggctgc ccctcatgga gtgcgtgcag
     1321 atgaccaagg aagtgcagaa agccatggat gacaagaggt ttgacgaggc cacccagctc
     1381 cgtggtggga gcttcgagaa caactggaac atttacaagc tcctcgccca ccagaagccc
     1441 cccaaggaga agtctaactt ctccctggcc atcctgaatg tgggggcccc ggcggctggc
     1501 atgaatgcgg ccgtgcgctc ggcggtgcgg accggcatct cccatggaca cacagtatac
     1561 gtggtgcacg atggcttcga aggcctagcc aagggtcagg tgcaagaagt aggctggcac
     1621 gacgtggccg gctggttggg gcgtggtggc tccatgctgg ggaccaagag gaccctgccc
     1681 aagggccagc tggagtccat tgtggagaac atccgcatct atggtattca cgccctgctg
     1741 gtggtcggtg ggtttgaggc ctatgaaggg gtgctgcagc tggtggaggc tcgcgggcgc
     1801 tacgaggagc tctgcatcgt catgtgtgtc atcccagcca ccatcagcaa caacgtccct
     1861 ggcaccgact tcagcctggg ctccgacact gctgtaaatg ccgccatgga gagctgtgac
     1921 cgcatcaaac agtctgcctc ggggaccaag cgccgtgtgt tcatcgtgga gaccatgggg
     1981 ggttactgtg gctacctggc caccgtgact ggcattgctg tgggggccga cgccgcctac
     2041 gtcttcgagg accctttcaa catccacgac ttaaaggtca acgtggagca catgacggag
     2101 aagatgaaga cagacattca gaggggcctg gtgctgcgga acgagaagtg ccatgactac
     2161 tacaccacgg agttcctgta caacctgtac tcatcagagg gcaagggcgt cttcgactgc
     2221 aggaccaatg tcctgggcca cctgcagcag ggtggcgctc caaccccctt tgaccggaac
     2281 tatgggacca agctgggggt gaaggccatg ctgtggttgt cggagaagct gcgcgaggtt
     2341 taccgcaagg gacgggtgtt cgccaatgcc ccagactcgg cctgcgtgat cggcctgaag
     2401 aagaaggcgg tggccttcag ccccgtcact gagctcaaga aagacactga tttcgagcac
     2461 cgcatgccac gggagcagtg gtggctgagc ctgcggctca tgctgaagat gctggcacaa
     2521 taccgcatca gtatggccgc ctacgtgtca ggggagctgg agcacgtgac ccgccgcacc
     2581 ctgagcatgg acaagggctt cagaattctg cagtcgacgg taccgcgggc ccgggatcca
     2641 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     2701 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     2761 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     2821 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     2881 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     2941 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     3001 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     3061 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     3121 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     3181 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     3241 cccgacaacc actacctgag cacccagtcc aagctgagca aagaccccaa cgagaagcgc
     3301 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     3361 ctgtacaagt aaagcggccg cggatcccgc ccctctccct cccccccccc taacgttact
     3421 ggccgaagcc gcttggaata aggccggtgt gcgtttgtct atatgttatt ttccaccata
     3481 ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc ctgtcttctt gacgagcatt
     3541 cctaggggtc tttcccctct cgccaaagga atgcaaggtc tgttgaatgt cgtgaaggaa
     3601 gcagttcctc tggaagcttc ttgaagacaa acaacgtctg tagcgaccct ttgcaggcag
     3661 cggaaccccc cacctggcga caggtgcctc tgcggccaaa agccacgtgt ataagataca
     3721 cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt ggatagttgt ggaaagagtc
     3781 aaatggctct cctcaagcgt attcaacaag gggctgaagg atgcccagaa ggtaccccat
     3841 tgtatgggat ctgatctggg gcctcggtgc acatgcttta catgtgttta gtcgaggtta
     3901 aaaaaacgtc taggcccccc gaaccacggg gacgtggttt tcctttgaaa aacacgatga
     3961 taagcttgcc acaaccatgg ctgaacaaga tggattgcac gcaggttctc cggccgcttg
     4021 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     4081 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     4141 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4201 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4261 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4321 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     4381 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     4441 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     4501 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4561 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4621 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4681 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4741 cttctatcgc cttcttgacg agttcttctg aacgcgtctg gaacaatcaa cctctggatt
     4801 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     4861 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     4921 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     4981 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     5041 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     5101 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     5161 ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct gctcgcctgt gttgccacct
     5221 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     5281 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     5341 cgagtcggat ctccctttgg gccgcctccc cgcctggaat taattctgca gtcgagacct
     5401 agaaaaacat ggagcaatca caagtagcaa tacagcagct accaatgctg attgtgcctg
     5461 gctagaagca caagaggagg aggaggtggg ttttccagtc acacctcagg tacctttaag
     5521 accaatgact tacaaggcag ctgtagatct tagccacttt ttaaaagaaa agaggggact
     5581 ggaagggcta attcactccc aacgaagaca agatatcctt gatctgtgga tctaccacac
     5641 acaaggctac ttccctgatt agcagaacta cacaccaggg ccaggggtca gatatccact
     5701 gacctttgga tggtgctaca agctagtacc agttgagcca gataaggtag aagaggccaa
     5761 taaaggagag aacaccagct tgttacaccc tgtgagcctg catgggatgg atgacccgga
     5821 gagagaagtg ttagagtgga ggtttgacag ccgcctagca tttcatcacg tggcccgaga
     5881 gctgcatccg gagtacttca agaactgctg atatcgagct tgctacaagg gactttccgc
     5941 tggggacttt ccagggaggc gtggcctggg cgggactggg gagtggcgag ccctcagatc
     6001 ctgcatataa gcagctgctt tttgcctgta ctgggtctct ctggttagac cagatctgag
     6061 cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt
     6121 gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca
     6181 gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc
     6241 gaaagggaaa ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc
     6301 aagaggcgag gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa
     6361 ggagagagat gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga
     6421 aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc
     6481 aagcagggag ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg
     6541 tagacaaata ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc
     6601 attatataat acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac
     6661 caaggaagct ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca
     6721 agcggccggc cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt
     6781 gaattatata aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca
     6841 aagagaagag tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg
     6901 ttcttgggag cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc
     6961 agacaattat tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg
     7021 caacagcatc tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg
     7081 gctgtggaaa gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa
     7141 ctcatttgca ccactgctgt gccttggaat gctagttgga gtaataaatc tctggaacag
     7201 atttggaatc acacgacctg gatggagtgg gacagagaaa ttaacaatta cacaagctta
     7261 atacactcct taattgaaga atcgcaaaac cagcaagaaa agaatgaaca agaattattg
     7321 gaattagata aatgggcaag tttgtggaat tggtttaaca taacaaattg gctgtggtat
     7381 ataaaattat tcataatgat agtaggaggc ttggtaggtt taagaatagt ttttgctgta
     7441 ctttctatag tgaatagagt taggcaggga tattcaccat tatcgtttca gacccacctc
     7501 ccaaccccga ggggacccga caggcccgaa ggaatagaag aagaaggtgg agagagagac
     7561 agagacagat ccattcgatt agtgaacgga tctcgacggt atcgccttta aaagaaaagg
     7621 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     7681 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     7741 cagcagagat ccagtttatc gataagcttg ggagttccgc gttacataac ttacggtaaa
     7801 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     7861 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     7921 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     7981 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     8041 tacttggcag tacatctacg tattagtcat cgctattacc atg
//
