LOCUS       sequence-215602-        6952 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  retroviral expression of mCherry, EGFP-tagged LC3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6952)
  AUTHORS   N'Diaye EN, Kajihara KK, Hsieh I, Morisaki H, Debnath J, Brown EJ
  TITLE     PLIC proteins or ubiquilins regulate autophagy-dependent cell 
            survival during nutrient starvation.
  JOURNAL   EMBO Rep. 2009 Feb;10(2):173-9. doi: 10.1038/embor.2008.238. Epub 
            2009 Jan 16.
  PUBMED    19148225
REFERENCE   2  (bases 1 to 6952)
  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"; doi: 
            "10.1038/embor.2008.238"; journalName: "EMBO Rep"; date: "2009-02"; 
            volume: "10"; issue: "2"; pages: "173-9"
COMMENT     Sequence Label: pBABE-puro mCherry-EGFP-LC3B
FEATURES             Location/Qualifiers
     source          1..6952
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..470
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    535..734
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             935..1351
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1281..1303
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1319..1335
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1362..1371
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1368..2075
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1513..1531)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1781..1800
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    2103..2127
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      2136..2145
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2142..2144,2145..2147,2148..2858)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2142..2144,2145..2147,2148..2858)
                     /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(2187..2208)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2448..2467)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2795..2816
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2886..3260
                     /codon_start=1
                     /gene="MAP1LC3B"
                     /product="human LC3 protein"
                     /label=LC3B
                     /note="Atg8 family member found on autophagosomes"
                     /translation="MPSEKTFKQRRTFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLP
                     VLDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESEKD
                     EDGFLYMVYASQETFGMKLSV"
     primer_bind     complement(3275..3295)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3280..3609
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3460..3595
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3619..4218
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3619..3638)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4115..4135
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4439..4908
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5038..5055)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5209..5797)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5289..5308)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5957..6748,6749..6817))
                     /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     6580..6599
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6818..6922)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
       61 gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc agttcctgcc
      121 ccggctcagg gccaagaaca gatggtcccc agatgcggtc cagccctcag cagtttctag
      181 agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt gccttatttg
      241 aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc cgagctcaat
      301 aaaagagccc acaacccctc actcggggcg ccagtcctcc gattgactga gtcgcccggg
      361 tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct cgctgttcct
      421 tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca tttgggggct
      481 cgtccgggat cgggagaccc ctgcccaggg accaccgacc caccaccggg aggcaagctg
      541 gccagcaact tatctgtgtc tgtccgattg tctagtgtct atgactgatt ttatgcgcct
      601 gcgtcggtac tagttagcta actagctctg tatctggcgg acccgtggtg gaactgacga
      661 gttctgaaca cccggccgca accctgggag acgtcccagg gactttgggg gccgtttttg
      721 tggcccgacc tgaggaaggg agtcgatgtg gaatccgacc ccgtcaggat atgtggttct
      781 ggtaggagac gagaacctaa aacagttccc gcctccgtct gaatttttgc tttcggtttg
      841 gaaccgaagc cgcgcgtctt gtctgctgca gcgctgcagc atcgttctgt gttgtctctg
      901 tctgactgtg tttctgtatt tgtctgaaaa ttagggccag actgttacca ctcccttaag
      961 tttgacctta ggtcactgga aagatgtcga gcggatcgct cacaaccagt cggtagatgt
     1021 caagaagaga cgttgggtta ccttctgctc tgcagaatgg ccaaccttta acgtcggatg
     1081 gccgcgagac ggcaccttta accgagacct catcacccag gttaagatca aggtcttttc
     1141 acctggcccg catggacacc cagaccaggt cccctacatc gtgacctggg aagccttggc
     1201 ttttgacccc cctccctggg tcaagccctt tgtacaccct aagcctccgc ctcctcttcc
     1261 tccatccgcc ccgtctctcc cccttgaacc tcctcgttcg accccgcctc gatcctccct
     1321 ttatccagcc ctcactcctt ctctaggcgc cggccggatc tgccaccatg gtgagcaagg
     1381 gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg
     1441 gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg
     1501 gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca
     1561 tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc
     1621 ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg
     1681 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct
     1741 acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga
     1801 ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg
     1861 agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca
     1921 cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg
     1981 acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc
     2041 gccactccac cggcggcatg gacgagctgt acaagtccgg actcagatcc gctggcccag
     2101 gaacaagttt gtacaaaaaa gcaggctttc cggtcgccac catggtgagc aagggcgagg
     2161 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     2221 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     2281 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     2341 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     2401 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     2461 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     2521 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     2581 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     2641 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     2701 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     2761 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     2821 ccgccgggat cactctcggc atggacgagc tgtacaagtc cggactcaga tccggtaccg
     2881 aattcatgcc gtcggagaag accttcaagc agcgccgcac cttcgaacaa agagtagaag
     2941 atgtccgact tattcgagag cagcatccaa ccaaaatccc ggtgataata gaacgataca
     3001 agggtgagaa gcagcttcct gttctggata aaacaaagtt ccttgtacct gaccatgtca
     3061 acatgagtga gctcatcaag ataattagaa ggcgcttaca gctcaatgct aatcaggcct
     3121 tcttcctgtt ggtgaacgga cacagcatgg tcagcgtctc cacaccaatc tcagaggtgt
     3181 atgagagtga gaaagatgaa gatggattcc tgtacatggt ctatgcctcc caggagacgt
     3241 tcgggatgaa attgtcagtg taagtcgacc ctgtggaatg tgtgtcagtt agggtgtgga
     3301 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3361 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     3421 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     3481 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     3541 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     3601 ttttgcaaaa agcttaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac
     3661 gacgtcccca gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc
     3721 cacaccgtcg atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc
     3781 acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg
     3841 gtctggacca cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc
     3901 atggccgagt tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg
     3961 ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac
     4021 cagggcaagg gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc
     4081 ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc
     4141 ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc
     4201 cgcaagcccg gtgcctgacg cccgccccac gacccgcagc gcccgaccga aaggagcgca
     4261 cgaccccatg catcgataaa ataaaagatt ttatttagtc tccagaaaaa ggggggaatg
     4321 aaagacccca cctgtaggtt tggcaagcta gcttaagtaa cgccattttg caaggcatgg
     4381 aaaaatacat aactgagaat agagaagttc agatcaaggt caggaacaga tggaacagct
     4441 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
     4501 acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag ttcctgcccc
     4561 ggctcagggc caagaacaga tggtccccag atgcggtcca gccctcagca gtttctagag
     4621 aaccatcaga tgtttccagg gtgccccaag gacctgaaat gaccctgtgc cttatttgaa
     4681 ctaaccaatc agttcgcttc tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa
     4741 aagagcccac aacccctcac tcggggcgcc agtcctccga ttgactgagt cgcccgggta
     4801 cccgtgtatc caataaaccc tcttgcagtt gcatccgact tgtggtctcg ctgttccttg
     4861 ggagggtctc ctctgagtga ttgactaccc gtcagcgggg gtctttcaca tgcagcatgt
     4921 atcaaaatta atttggtttt ttttcttaag tatttacatt aaatggccat agttgcatta
     4981 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5041 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5101 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5161 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5221 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5281 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5341 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5401 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5461 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5521 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5581 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5641 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5701 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5761 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5821 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5881 aaaaaggatc ttcacctaga tccttttgcg gccgcaaatc aatctaaagt atatatgagt
     5941 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6181 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6241 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6481 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6541 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6601 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6661 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6781 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6841 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6901 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgca gc
//
