LOCUS       sequence-371293-        7587 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for autophagy.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7587)
  AUTHORS   Minami Y, Hoshino A, Higuchi Y, Hamaguchi M, Kaneko Y, Kirita Y, 
            Taminishi S, Nishiji T, Taruno A, Fukui M, Arany Z, Matoba S
  TITLE     Liver lipophagy ameliorates nonalcoholic steatohepatitis through 
            extracellular lipid secretion.
  JOURNAL   Nat Commun. 2023 Jul 13;14(1):4084. doi: 10.1038/s41467-023-39404-6.
  PUBMED    37443159
REFERENCE   2  (bases 1 to 7587)
  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: "Nat 
            Commun."; date: "2023-07-13"; volume: "14(1):4084. doi"; pages: " 
            10.1038/s41467-023-39404-6"
COMMENT     Sequence Label: pMSCV mKeima-LC3
FEATURES             Location/Qualifiers
     source          1..7587
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1422..1431
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1428..1430,1431..1433,1434..2093)
                     /codon_start=1
                     /product="monomeric Keima-Red fluorescent protein, also 
                     known as mKeima"
                     /label=mKeima-Red
                     /translation="MVSVIAKQMTYKVYMSGTVNGHYFEVEGDGKGKPYEGEQTVKLTV
                     TKGGPLPFAWDILSPQLQYGSIPFTKYPEDIPDYFKQSFPEGYTWERSMNFEDGAVCTV
                     SNDSSIQGNCFIYNVKISGENFPPNGPVMQKKTQGWEPSTERLFARDGMLIGNDYMALK
                     LEGGGHYLCEFKSTYKAKKPVRMPGRHEIDRKLDVTSHNRDYTSVEQCEIAIARHSLLG
                     "
     CDS             2115..2537
                     /codon_start=1
                     /gene="Map1lc3b"
                     /product="rat LC3 protein"
                     /label=LC3
                     /note="Atg8 family member found on autophagosomes"
                     /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV
                     LDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE
                     DGFLYMVYASQETFGTALAVTYMSALKATATGREPCL"
     promoter        2551..3050
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2586..2609)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2983..3002
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3071..3874
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3134..3153)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3744..3763
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             3980..4494
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4656..4672
                     /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)."
     promoter        complement(join(4680..4686,4687..4704,4705..4710))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4725..4746
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4863..4880)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5034..5622)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5114..5133)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5793..6584,6585..6653))
                     /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     6416..6435
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6654..6758)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6826..6844
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6882..6904)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7004..7023
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7217..7239
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7367..7386
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7586..7587,1..515)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaatta gatctctcga ggccaccatg gtgagtgtga
     1441 tcgctaaaca aatgacctac aaggtttata tgtcaggcac ggtcaatgga cactactttg
     1501 aggtcgaagg cgatggaaaa ggaaagcctt acgagggaga gcagacagta aagctcactg
     1561 tcaccaaggg tggacctctg ccatttgctt gggatatttt atcaccacag cttcagtacg
     1621 gaagcatacc attcaccaag taccctgaag acatccctga ttatttcaag cagtcattcc
     1681 ctgagggata tacatgggag aggagcatga actttgaaga tggtgcagtg tgtactgtca
     1741 gcaatgattc cagcatccaa ggcaactgtt tcatctacaa tgtcaaaatc tctggtgaga
     1801 actttcctcc caatggacct gttatgcaga agaagacaca gggctgggaa cccagcactg
     1861 agcgtctctt tgcacgagat ggaatgctga taggaaacga ttatatggct ctgaagttgg
     1921 aaggaggtgg tcactatttg tgtgaattta aatctactta caaggcaaag aagcctgtga
     1981 ggatgccagg gcgccacgag attgaccgca aactggatgt aaccagtcac aacagggatt
     2041 acacatctgt tgagcagtgt gaaatagcca ttgcacgcca ctctttgctc ggtgttaacg
     2101 cggccgcggg gacaccgtcc gagaagacct tcaaacagcg ccggagcttc gaacaaagag
     2161 tggaagatgt ccggctcatc cgggagcagc accccaccaa gatcccagtg attatagagc
     2221 gatacaaggg tgagaagcag ctgcccgtcc tggacaagac caagttcctt gtacctgatc
     2281 acgtgaatat gagcgaactc atcaagataa ttagaaggcg cctgcagctc aatgctaacc
     2341 aagccttctt cctcctggtg aatgggcaca gcatggtgag tgtgtccaca cccatctctg
     2401 aagtgtacga gagcgagaga gatgaagacg gcttcctgta catggtctat gcctcccagg
     2461 agacgttcgg gacagcactg gctgttacat acatgtcagc tctgaaggca acagcaacag
     2521 gaagagagcc atgcttgtga gaattctacc gggtagggga ggcgcttttc ccaaggcagt
     2581 ctggagcatg cgctttagca gccccgctgg gcacttggcg ctacacaagt ggcctctggc
     2641 ctcgcacaca ttccacatcc accggtaggc gccaaccggc tccgttcttt ggtggcccct
     2701 tcgcgccacc ttctactcct cccctagtca ggaagttccc ccccgccccg cagctcgcgt
     2761 cgtgcaggac gtgacaaatg gaagtagcac gtctcactag tctcgtgcag atggacagca
     2821 ccgctgagca atggaagcgg gtaggccttt ggggcagcgg ccaatagcag ctttgctcct
     2881 tcgctttctg ggctcagagg ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc
     2941 tcaggggcgg ggcgggcgcc cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa
     3001 agcgcacgtc tgccgcgctg ttctcctctt cctcatctcc gggcctttcg acctgcagcc
     3061 caagcttacc atgggatcgg ccattgaaca agatggattg cacgcaggtt ctccggccgc
     3121 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     3181 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     3241 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg
     3301 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     3361 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     3421 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     3481 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     3541 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     3601 caaggcgcgc atgcccgacg gcgatgatct cgtcgtgacc catggcgatg cctgcttgcc
     3661 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     3721 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     3781 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     3841 cgccttctat cgccttcttg acgagttctt ctgaggatcc cgcccgcccc acgacccgca
     3901 gcgcccgacc gaaaggagcg cacgacccca tgcatcgata aaataaaaga ttttatttag
     3961 tctccagaaa aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt
     4021 aacgccattt tgcaaggcat ggaaaataca taactgagaa tagagaagtt cagatcaagg
     4081 ttaggaacag agagacagca gaatatgggc caaacaggat atctgtggta agcagttcct
     4141 gccccggctc agggccaaga acagatggtc cccagatgcg gtcccgccct cagcagtttc
     4201 tagagaacca tcagatgttt ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat
     4261 ttgaactaac caatcagttc gcttctcgct tctgttcgcg cgcttctgct ccccgagctc
     4321 aataaaagag cccacaaccc ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc
     4381 gggtacccgt gtatccaata aaccctcttg cagttgcatc cgacttgtgg tctcgctgtt
     4441 ccttgggagg gtctcctctg agtgattgac tacccgtcag cgggggtctt tcatgggtaa
     4501 cagtttcttg aagttggaga acaacattct gagggtagga gtcgaatatt aagtaatcct
     4561 gactcaatta gccactgttt tgaatccaca tactccaata ctcctgaaat agttcattat
     4621 ggacagcgca gaaagagctg gggagaattg tgaaattgtt atccgctcac aattccacac
     4681 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     4741 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4801 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     4861 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     4921 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4981 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     5041 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     5101 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     5161 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     5221 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     5281 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     5341 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     5401 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     5461 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     5521 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     5581 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5641 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     5701 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     5761 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     5821 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     5881 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     5941 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     6001 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     6061 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     6121 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     6181 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     6241 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     6301 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     6361 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     6421 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6481 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     6541 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     6601 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6661 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     6721 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     6781 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     6841 aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc
     6901 ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc
     6961 gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt
     7021 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     7081 cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
     7141 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
     7201 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggcgcaagg aatggtgcat
     7261 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga
     7321 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     7381 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     7441 agaggcgatt agtccaattt gttaaagaca ggatatcagt ggtccaggct ctagttttga
     7501 ctcaacaata tcaccagctg aagcctatag agtacgagcc atagataaaa taaaagattt
     7561 tatttagtct ccagaaaaag gggggaa
//
