LOCUS       sequence-256731-        8123 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8123)
  AUTHORS   Itakura E, Mizushima N
  TITLE     Characterization of autophagosome formation site by a hierarchical 
            analysis of mammalian Atg proteins.
  JOURNAL   Autophagy. 2010 Aug;6(6):764-76.
  PUBMED    20639694
REFERENCE   2  (bases 1 to 8123)
  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: 
            "Autophagy."; date: "2010-08"; volume: "6(6)"; pages: "764-76"
COMMENT     Sequence Label: pMXs-IP spGFP-ERGIC53
FEATURES             Location/Qualifiers
     source          1..8123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..594
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    657..1014
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1073..1489
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1419..1441
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1457..1473
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1499..1873
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1811..1827
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             join(1962..1964,1965..2675)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1962..1964,1965..2675)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2004..2025)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2265..2284)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2612..2633
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    join(4247..4821,4822..4824,4825..4833)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4414..4431)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4641..4660
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4834..5436
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MVTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     5333..5353
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5607..6076
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(6206..6223)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6377..6965)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6457..6476)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7129..7920,7921..7989))
                     /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     7752..7771
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7990..8094)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaaat acataactga gaatagaaaa gttcagatca aggtcaggaa cagatggaac
      121 agctgaatat gggccaaaca ggatatctgt ggtaagcagt tcctgccccg gctcagggcc
      181 aagaacagat ggaacagctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg
      241 ccccggctca gggccaagaa cagatggtcc ccagatgcgg tccagccctc agcagtttct
      301 agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt
      361 tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca
      421 ataaaagagc ccacaacccc tcactcggcg cgccagtcct ccgattgact gagtcgcccg
      481 ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt ctcgctgttc
      541 cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt catttggggg
      601 ctcgtccggg atcgggagac ccctgcccag ggaccaccga cccaccaccg ggaggtaagc
      661 tggccagcaa cttatctgtg tctgtccgat tgtctagtgt ctatgactga ttttatgcgc
      721 ctgcgtcggt actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac
      781 gagttcggaa cacccggccg caaccctggg agacgtccca gggacttcgg gggccgtttt
      841 tgtggcccga cctgagtcca aaaatcccga tcgttttgga ctctttggtg caccccccta
      901 ataggaggga tatgtggttc tggtaggaga cgagaaccta aaacagttcc cgcctccgtc
      961 tgaatttttg ctttcggttt gggaccgaag ccgcgccgcg cgtcttgtct gctgcagcat
     1021 cgttctgtgt tgtctctgtc tgactgtgtt tctgtatttg tctgaaaatt agggccagac
     1081 tgttaccact cccttaagtt tgaccttagg tcactggaaa gatgtcgagc ggatcgctca
     1141 caaccagtcg gtagatgtca agaagagacg ttgggttacc ttctgctctg cagaatggcc
     1201 aacctttaac gtcggatggc cgcgagacgg cacctttaac cgagacctca tcacccaggt
     1261 taagatcaag gtcttttcac ctggcccgca tggacaccca gaccaggtcc cctacatcgt
     1321 gacctgggaa gccttggctt ttgacccccc tccctgggtc aagccctttg tacaccctaa
     1381 gcctccgcct cctcttcctc catccgcccc gtctctcccc cttgaacctc ctcgttcgac
     1441 cccgcctcga tcctcccttt atccagccct cactccttct ctaggcgccc ccatatggcc
     1501 atatgagatc ttatatgggg cacccccgcc ccttgtaaac ttccctgacc ctgacatgac
     1561 aagagttact aacagcccct ctctccaagc tcacttacag gctctctact tagtccagca
     1621 cgaagtctgg agacctctgg cggcagccta ccaagaacaa ctggaccgac cggtggtacc
     1681 tcacccttac cgagtcggcg acacagtgtg ggtccgccga caccagacta agaacctaga
     1741 acctcgctgg aaaggacctt acacagtcct gctgaccacc cccaccgccc tcaaagtaga
     1801 cggcatcgca gcttggatac acgccgccca cgtgaaggct gccgaccccg ggggtggacc
     1861 atcctctaga ctgccggatc tagctagtta attaaggatc taccgccatg ctcctttcgg
     1921 tgccgctcct gcttggcctc ctcggcctgg ccgccgcaga cgtgagcaag ggcgaggagc
     1981 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     2041 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     2101 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     2161 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     2221 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     2281 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     2341 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     2401 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     2461 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     2521 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     2581 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     2641 ccgggatcac tctcggcatg gacgagctgt acaagggtgg cggtggctcg ggcggtggtg
     2701 ggtcgggtgg cggcggatct atcgatgaat tgaattcacg cgttcgcgag ttaacggatc
     2761 cagacggcgt gggaggagac cccgcggtcg cgttgccaca tcgccgtttc gagtacaaat
     2821 acagcttcaa ggggccgcac ctggtgcaga gcgacgggac cgtgcccttc tgggcccacg
     2881 cggggaatgc tattccaagt tcagatcaaa ttcgagtagc accatcttta aaaagccaaa
     2941 gaggctcagt gtggacaaag acaaaagcgg cctttgagaa ctgggaagtt gaggtgacat
     3001 ttcgagtgac tggaagaggt cgaattggag ctgatggcct agcaatttgg tatgcagaaa
     3061 atcaaggctt ggagggccct gtgtttggat cagctgatct gtggaatggt gttggaatat
     3121 tttttgattc ttttgacaat gatggaaaga aaaataatcc tgctatagta attataggca
     3181 acaatggaca aatccattat gaccatcaaa atgacggggc tagtcaagct ttggcaagtt
     3241 gccagaggga cttccgcaac aaaccctatc ctgtccgagc aaagattacc tattaccaga
     3301 acacactgac agtaatgatc aataatggct ttacaccaga taaaaatgat tatgaatttt
     3361 gtgccaaagt ggaaaatatg attatccctg cacaagggca ttttggaata tctgctgcaa
     3421 ctggaggtct tgcagatgac catgatgtcc tttcttttct gactttccag ttgactgaac
     3481 ctggaaaaga gccgcccaca ccagataaag aaatttcgga aaaggaaaaa gaaaagtatc
     3541 aggaggaatt tgagcacttt caacaagaat tggataaaaa aaaagaggaa ttccagaagg
     3601 gccaccccga cctccaaggg cagcctgcgg aggaaatatt tgagagtgta ggagatcgag
     3661 agctaagaca agtctttgaa ggacagaatc gtattcatct tgaaatcaag cagctgaacc
     3721 ggcagttaga tatgattctt gatgaacaga gaagatatgt ctcttcctta acagaggaaa
     3781 tctctaaaag aggagcagga atgcctgggc agcatgggca gattactcaa caagaactgg
     3841 atactgttgt gaaaactcag catgagattc tgagacaagt aaatgaaatg aaaaattcct
     3901 tgagtgaaac cgtcagactg gtcagtggaa tgcagcaccc tggctctgct ggaggcgtct
     3961 atgagacaac acagcacttc attgacatca aagagcacct gcacatagta aagagggaca
     4021 tagataactt agtgcagcga aatatgccat caaatgaaaa gccgaaatgc ccagaactac
     4081 caccatttcc atcatgtttg tctacggtcc acttcattat atttgttgtg gtgcaaactg
     4141 tattattcat tggttatatc atgtataggt ctcagcaaga agcagctgcc aaaaaattct
     4201 tttgaggatc cctcgagcgg ccgctacgta aattccgccc cccccccccc tctccctccc
     4261 ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata
     4321 tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg
     4381 tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg caaggtctgt
     4441 tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca acgtctgtag
     4501 cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc ggccaaaagc
     4561 cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga
     4621 tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg ctgaaggatg
     4681 cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca tgctttacat
     4741 gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc
     4801 tttgaaaaac acgatgataa tatggccaca accatggtta ccgagtacaa gcccacggtg
     4861 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     4921 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     4981 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     5041 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     5101 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     5161 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     5221 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     5281 gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     5341 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     5401 cgcacctggt gcatgacccg caagcccggt gcctgagtcg acgataaaat aaaagatttt
     5461 atttagtctc cagaaaaagg ggggaatgaa agaccccacc tgtaggtttg gcaagctagc
     5521 ttaagtaacg ccattttgca aggcatggaa aaatacataa ctgagaatag agaagttcag
     5581 atcaaggtca ggaacagatg gaacagctga atatgggcca aacaggatat ctgtggtaag
     5641 cagttcctgc cccggctcag ggccaagaac agatggaaca gctgaatatg ggccaaacag
     5701 gatatctgtg gtaagcagtt cctgccccgg ctcagggcca agaacagatg gtccccagat
     5761 gcggtccagc cctcagcagt ttctagagaa ccatcagatg tttccagggt gccccaagga
     5821 cctgaaatga ccctgtgcct tatttgaact aaccaatcag ttcgcttctc gcttctgttc
     5881 gcgcgcttct gctccccgag ctcaataaaa gagcccacaa cccctcactc ggggcgccag
     5941 tcctccgatt gactgagtcg cccgggtacc cgtgtatcca ataaaccctc ttgcagttgc
     6001 atccgacttg tggtctcgct gttccttggg agggtctcct ctgagtgatt gactacccgt
     6061 cagcgggggt ctttcacatg cagcatgtat caaaattaat ttggtttttt ttcttaagta
     6121 tttacattaa atggccatag ttgcattaat gaatcggcca acgcgcgggg agaggcggtt
     6181 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     6241 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     6301 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     6361 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     6421 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     6481 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     6541 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     6601 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     6661 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6721 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6781 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     6841 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6901 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6961 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     7021 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttgcggc
     7081 cggccgcaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     7141 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     7201 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     7261 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     7321 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     7381 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     7441 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     7501 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     7561 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     7621 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     7681 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     7741 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     7801 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     7861 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     7921 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     7981 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     8041 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     8101 tccccgaaaa gtgccacctg cat
//
