LOCUS       sequence-202163-        5668 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Rluc-LC3wt. Can be used for measuring 
            autophagic flux..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5668)
  AUTHORS   Farkas T, Jaattela M
  TITLE     Renilla Luciferase-LC3 Based Reporter Assay for Measuring Autophagic
            Flux.
  JOURNAL   Methods Enzymol. 2017;588:1-13. doi: 10.1016/bs.mie.2016.09.072. 
            Epub 2016 Nov 12.
  PUBMED    28237095
REFERENCE   2  (bases 1 to 5668)
  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: "Methods 
            Enzymol."; date: "2017"; volume: "588:1-13. doi"; pages: " 
            10.1016/bs.mie.2016.09.072. Epub 2016 Nov 12"
COMMENT     Sequence Label: pRluc-LC3wt
FEATURES             Location/Qualifiers
     source          1..5668
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             553..1485
                     /codon_start=1
                     /product="luciferase from the anthozoan coelenterate 
                     Renilla reniformis (sea pansy)"
                     /label=Rluc
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGAALAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             1504..1926
                     /codon_start=1
                     /gene="Map1lc3b"
                     /product="rat LC3 protein"
                     /label=LC3
                     /note="Atg8 family member found on autophagosomes"
                     /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV
                     LDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE
                     DGFLYMVYASQETFGTALAVTYMSALKATATGREPCL"
     polyA_signal    2396..2517
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2433..2452)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2487..2506
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2524..2979)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2661..2682)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2873..2892
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3006..3110
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3112..3469
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3112..3303
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3135..3155)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3320..3455
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3504..4298
                     /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             3504..4298
                     /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(3558..3577)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4168..4187
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4486..4505)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4530..4577
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4906..5494
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5395..5414
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggtcgcca ccatgacttc gaaagtttat gatccagaac aaaggaaacg gatgataact
      601 ggtccgcagt ggtgggccag atgtaaacaa atgaatgttc ttgattcatt tattaattat
      661 tatgattcag aaaaacatgc agaaaatgct gttatttttt tacatggtaa cgcggcctct
      721 tcttatttat ggcgacatgt tgtgccacat attgagccag tagcgcggtg tattatacca
      781 gaccttattg gtatgggcaa atcaggcaaa tctggtaatg gttcttatag gttacttgat
      841 cattacaaat atcttactgc atggtttgaa cttcttaatt taccaaagaa gatcattttt
      901 gtcggccatg attggggtgc tgctttggca tttcattata gctatgagca tcaagataag
      961 atcaaagcaa tagttcacgc tgaaagtgta gtagatgtga ttgaatcatg ggatgaatgg
     1021 cctgatattg aagaagatat tgcgttgatc aaatctgaag aaggagaaaa aatggttttg
     1081 gagaataact tcttcgtgga aaccatgttg ccatcaaaaa tcatgagaaa gttagaacca
     1141 gaagaatttg cagcatatct tgaaccattc aaagagaaag gtgaagttcg tcgtccaaca
     1201 ttatcatggc ctcgtgaaat cccgttagta aaaggtggta aacctgacgt tgtacaaatt
     1261 gttaggaatt ataatgctta tctacgtgca agtgatgatt taccaaaaat gtttattgaa
     1321 tcggacccag gattcttttc caatgctatt gttgaaggtg ccaagaagtt tcctaatact
     1381 gaatttgtca aagtaaaagg tcttcatttt tcgcaagaag atgcacctga tgaaatggga
     1441 aaatatatca aatcgttcgt tgagcgagtt ctcaaaaatg aacaatccgg actcagatct
     1501 agaccgtccg agaagacctt caaacagcgc cggagcttcg aacaaagagt ggaagatgtc
     1561 cggctcatcc gggagcagca ccccaccaag atcccagtga ttatagagcg atacaagggt
     1621 gagaagcagc tgcccgtcct ggacaagacc aagttccttg tacctgatca cgtgaatatg
     1681 agcgaactca tcaagataat tagaaggcgc ctgcagctca atgctaacca agccttcttc
     1741 ctcctggtga atgggcacag catggtgagt gtgtccacac ccatctctga agtgtacgag
     1801 agcgagagag atgaagacgg cttcctgtac atggtctatg cctcccagga gacgttcggg
     1861 acagcactgg ctgttacata catgtcagct ctgaaggcaa cagcaacagg aagagagcca
     1921 tgcttgtgac agacatacag ccacttccaa ctaaagcaag cctctgctcc ctgctacctg
     1981 catggagccc actgtgacac tcagaccatc cccggtcact cactcactcg tgtctgagaa
     2041 tctcagtgag agctgcctct gtcacggacg gaagccaaca cagccacctc tcgacctggc
     2101 tccccacagc acccaccctc cctgcatgca gctgtccctg ctaaccccca atgttatgtt
     2161 acactgtgta agtccccact gctgccgtgt gtgggttgtg tacgtcgtca tgtccctggt
     2221 ttataactat ggtgcgaatt ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc
     2281 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     2341 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     2401 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     2461 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg
     2521 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     2581 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     2641 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     2701 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     2761 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     2821 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     2881 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     2941 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa
     3001 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     3061 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg
     3121 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     3181 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     3241 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     3301 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     3361 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     3421 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag
     3481 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     3541 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     3601 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     3661 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     3721 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     3781 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     3841 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     3901 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     3961 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     4021 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     4081 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     4141 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     4201 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     4261 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     4321 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     4381 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     4441 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga
     4501 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     4561 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     4621 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     4681 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     4741 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     4801 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     4861 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4921 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4981 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5041 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     5101 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     5161 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     5221 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     5281 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     5341 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     5401 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     5461 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     5521 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     5581 tgtggataac cgtattaccg ccatgcatta gttattaata gtaatcaatt acggggtcat
     5641 tagttcatag cccatatatg gagttccg
//
