LOCUS       sequence-359658-        5452 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Autophagy Marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5452)
  AUTHORS   Dunsing V, Petrich A, Chiantia S
  TITLE     Multicolor fluorescence fluctuation spectroscopy in living cells via
            spectral detection.
  JOURNAL   Elife. 2021 Sep 8;10. pii: 69687. doi: 10.7554/eLife.69687.
  PUBMED    34494547
REFERENCE   2  (bases 1 to 5452)
  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: "Elife."; 
            date: "2021-09-8"; volume: "10. pii: 69687. doi"; pages: " 
            10.7554/eLife.69687"
COMMENT     Sequence Label: LC3-mEYFP
FEATURES             Location/Qualifiers
     source          1..5452
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        258..561
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        562..765
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     716..736
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      804..813
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(810..812,813..815,816..1526)
                     /codon_start=1
                     /product="enhanced YFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(855..876)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1116..1135)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1463..1484
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1545..1967
                     /codon_start=1
                     /gene="Map1lc3b"
                     /product="rat LC3 protein"
                     /label=LC3
                     /note="Atg8 family member found on autophagosomes"
                     /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV
                     LDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE
                     DGFLYMVYASQETFGTALAVTYMSALKATATGREPCL"
     polyA_signal    2437..2558
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2474..2493)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2528..2547
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2565..3020)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2702..2723)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2914..2933
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3047..3151
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3153..3510
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3153..3344
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3176..3196)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3361..3496
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3545..4339
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3545..4339
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3599..3618)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4209..4228
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4527..4546)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4571..4618
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      join(4947..5452,1..83)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     join(5436..5452,1..3)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
       61 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
      121 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
      181 gtattaccgc catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc
      241 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
      301 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      361 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      421 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      481 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      541 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
      601 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
      661 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
      721 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt
      781 cagatccgct agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg
      841 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
      901 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
      961 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cttcggctac ggcctgcagt
     1021 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     1081 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     1141 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     1201 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
     1261 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
     1321 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
     1381 acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccaaa ctgagcaaag
     1441 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
     1501 ctctcggcat ggacgagctg tacaagtccg gactcagatc tagaccgtcc gagaagacct
     1561 tcaaacagcg ccggagcttc gaacaaagag tggaagatgt ccggctcatc cgggagcagc
     1621 accccaccaa gatcccagtg attatagagc gatacaaggg tgagaagcag ctgcccgtcc
     1681 tggacaagac caagttcctt gtacctgatc acgtgaatat gagcgaactc atcaagataa
     1741 ttagaaggcg cctgcagctc aatgctaacc aagccttctt cctcctggtg aatgggcaca
     1801 gcatggtgag tgtgtccaca cccatctctg aagtgtacga gagcgagaga gatgaagacg
     1861 gcttcctgta catggtctat gcctcccagg agacgttcgg gacagcactg gctgttacat
     1921 acatgtcagc tctgaaggca acagcaacag gaagagagcc atgcttgtga cagacataca
     1981 gccacttcca actaaagcaa gcctctgctc cctgctacct gcatggagcc cactgtgaca
     2041 ctcagaccat ccccggtcac tcactcactc gtgtctgaga atctcagtga gagctgcctc
     2101 tgtcacggac ggaagccaac acagccacct ctcgacctgg ctccccacag cacccaccct
     2161 ccctgcatgc agctgtccct gctaaccccc aatgttatgt tacactgtgt aagtccccac
     2221 tgctgccgtg tgtgggttgt gtacgtcgtc atgtccctgg tttataacta tggtgcgaat
     2281 tctgcagtcg acggtaccgc gggcccggga tccaccggat ctagataact gatcataatc
     2341 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     2401 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     2461 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     2521 tctagttgtg gtttgtccaa actcatcaat gtatcttaac gcgtaaattg taagcgttaa
     2581 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     2641 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     2701 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     2761 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     2821 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     2881 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     2941 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     3001 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3061 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3121 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     3181 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3241 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3301 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3361 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3421 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     3481 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     3541 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     3601 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3661 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3721 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     3781 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     3841 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     3901 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3961 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     4021 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     4081 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     4141 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     4201 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     4261 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     4321 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     4381 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     4441 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     4501 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     4561 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     4621 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     4681 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     4741 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     4801 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     4861 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     4921 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     4981 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     5041 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     5101 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     5161 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     5221 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     5281 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     5341 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     5401 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tc
//
