LOCUS       sequence-220582-        5452 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of rat LC3 fused to EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5452)
  AUTHORS   Kabeya Y, Mizushima N, Ueno T, Yamamoto A, Kirisako T, Noda T, 
            Kominami E, Ohsumi Y, Yoshimori T
  TITLE     LC3, a mammalian homologue of yeast Apg8p, is localized in 
            autophagosome membranes after processing.
  JOURNAL   EMBO J. 2000 Nov 1. 19(21):5720-8.
  PUBMED    11060023
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: "EMBO J. 
            2000 Nov 1. 19(21):5720-8."
COMMENT     Sequence Label: pEGFP-LC3
FEATURES             Location/Qualifiers
     source          1..5452
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        121..424
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        425..628
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     579..599
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      667..676
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(673..675,676..678,679..1389)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(673..675,676..678,679..1389)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(718..739)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(979..998)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1326..1347
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1408..1830
                     /codon_start=1
                     /gene="Map1lc3b"
                     /product="rat LC3 protein"
                     /label=LC3
                     /note="Atg8 family member found on autophagosomes"
                     /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV
                     LDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE
                     DGFLYMVYASQETFGTALAVTYMSALKATATGREPCL"
     polyA_signal    2300..2421
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2337..2356)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2391..2410
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2428..2883)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2565..2586)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2777..2796
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2910..3014
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3016..3373
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3016..3207
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3039..3059)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3224..3359
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3408..4202
                     /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             3408..4202
                     /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(3462..3481)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4072..4091
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4390..4409)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4434..4481
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4810..5398
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5299..5318
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgccatgcat
       61 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      121 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      181 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      241 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      301 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      361 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      421 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      481 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      541 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      601 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      661 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      721 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      781 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      841 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      901 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      961 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1021 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1081 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1141 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1201 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1261 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1321 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1381 ctgtacaagt ccggactcag atctagaccg tccgagaaga ccttcaaaca gcgccggagc
     1441 ttcgaacaaa gagtggaaga tgtccggctc atccgggagc agcaccccac caagatccca
     1501 gtgattatag agcgatacaa gggtgagaag cagctgcccg tcctggacaa gaccaagttc
     1561 cttgtacctg atcacgtgaa tatgagcgaa ctcatcaaga taattagaag gcgcctgcag
     1621 ctcaatgcta accaagcctt cttcctcctg gtgaatgggc acagcatggt gagtgtgtcc
     1681 acacccatct ctgaagtgta cgagagcgag agagatgaag acggcttcct gtacatggtc
     1741 tatgcctccc aggagacgtt cgggacagca ctggctgtta catacatgtc agctctgaag
     1801 gcaacagcaa caggaagaga gccatgcttg tgacagacat acagccactt ccaactaaag
     1861 caagcctctg ctccctgcta cctgcatgga gcccactgtg acactcagac catccccggt
     1921 cactcactca ctcgtgtctg agaatctcag tgagagctgc ctctgtcacg gacggaagcc
     1981 aacacagcca cctctcgacc tggctcccca cagcacccac cctccctgca tgcagctgtc
     2041 cctgctaacc cccaatgtta tgttacactg tgtaagtccc cactgctgcc gtgtgtgggt
     2101 tgtgtacgtc gtcatgtccc tggtttataa ctatggtgcg aattctgcag tcgacggtac
     2161 cgcgggcccg ggatccaccg gatctagata actgatcata atcagccata ccacatttgt
     2221 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     2281 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     2341 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     2401 caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg
     2461 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     2521 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     2581 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     2641 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     2701 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     2761 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     2821 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     2881 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     2941 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     3001 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     3061 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3121 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3181 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     3241 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3301 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3361 aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     3421 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     3481 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     3541 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag
     3601 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     3661 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     3721 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     3781 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3841 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3901 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg
     3961 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     4021 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     4081 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     4141 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     4201 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     4261 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     4321 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg
     4381 gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat
     4441 aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
     4501 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
     4561 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
     4621 cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt
     4681 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     4741 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     4801 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4861 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     4921 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     4981 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     5041 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     5101 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     5161 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     5221 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     5281 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     5341 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     5401 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ct
//
