LOCUS       Exported                7553 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pMXs-IP-EGFP-mAtg5
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7553)
  AUTHORS   Hara T, Takamura A, Kishi C, Iemura S, Natsume T, Guan JL, Mizushima
            N
  TITLE     FIP200, a ULK-interacting protein, is required for autophagosome 
            formation in mammalian cells.
  JOURNAL   J Cell Biol. 2008 May 5. 181(3):497-510.
  PUBMED    18443221
REFERENCE   2  (bases 1 to 7553)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7553
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     LTR             22..615
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    678..1035
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1094..1510
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1440..1462
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1478..1494
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1520..1894
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1832..1848
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      1980..1989
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1986..2702
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             1986..2702
                     /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(2031..2052)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2292..2311)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2639..2660
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3584..3600)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3585..3601)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     misc_feature    3699..4284
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3865..3882)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4092..4111
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4285..4887
                     /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     4784..4804
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5058..5527
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5657..5674)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5828..6416)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5908..5927)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6580..7440)
                     /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     7203..7222
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7441..7545)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cccgaaaagt gccacctgca taatgaaaga ccccacctgt aggtttggca agctagctta
       61 agtaacgcca ttttgcaagg catggaaaaa tacataactg agaatagaaa agttcagatc
      121 aaggtcagga acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag
      181 ttcctgcccc ggctcagggc caagaacaga tggaacagct gaatatgggc caaacaggat
      241 atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc cccagatgcg
      301 gtccagccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc ccaaggacct
      361 gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct tctgttcgcg
      421 cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggc gcgccagtcc
      481 tccgattgac tgagtcgccc gggtacccgt gtatccaata aaccctcttg cagttgcatc
      541 cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac tacccgtcag
      601 cgggggtctt tcatttgggg gctcgtccgg gatcgggaga cccctgccca gggaccaccg
      661 acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga ttgtctagtg
      721 tctatgactg attttatgcg cctgcgtcgg tactagttag ctaactagct ctgtatctgg
      781 cggacccgtg gtggaactga cgagttcgga acacccggcc gcaaccctgg gagacgtccc
      841 agggacttcg ggggccgttt ttgtggcccg acctgagtcc aaaaatcccg atcgttttgg
      901 actctttggt gcacccccct aataggaggg atatgtggtt ctggtaggag acgagaacct
      961 aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tgggaccgaa gccgcgccgc
     1021 gcgtcttgtc tgctgcagca tcgttctgtg ttgtctctgt ctgactgtgt ttctgtattt
     1081 gtctgaaaat tagggccaga ctgttaccac tcccttaagt ttgaccttag gtcactggaa
     1141 agatgtcgag cggatcgctc acaaccagtc ggtagatgtc aagaagagac gttgggttac
     1201 cttctgctct gcagaatggc caacctttaa cgtcggatgg ccgcgagacg gcacctttaa
     1261 ccgagacctc atcacccagg ttaagatcaa ggtcttttca cctggcccgc atggacaccc
     1321 agaccaggtc ccctacatcg tgacctggga agccttggct tttgaccccc ctccctgggt
     1381 caagcccttt gtacacccta agcctccgcc tcctcttcct ccatccgccc cgtctctccc
     1441 ccttgaacct cctcgttcga ccccgcctcg atcctccctt tatccagccc tcactccttc
     1501 tctaggcgcc cccatatggc catatgagat cttatatggg gcacccccgc cccttgtaaa
     1561 cttccctgac cctgacatga caagagttac taacagcccc tctctccaag ctcacttaca
     1621 ggctctctac ttagtccagc acgaagtctg gagacctctg gcggcagcct accaagaaca
     1681 actggaccga ccggtggtac ctcaccctta ccgagtcggc gacacagtgt gggtccgccg
     1741 acaccagact aagaacctag aacctcgctg gaaaggacct tacacagtcc tgctgaccac
     1801 ccccaccgcc ctcaaagtag acggcatcgc agcttggata cacgccgccc acgtgaaggc
     1861 tgccgacccc gggggtggac catcctctag actgccggat ctagctagtt aattaaggat
     1921 cccagtgtgg tggtacggga attcctgcag gcctcgaggg ccggcgcgcc gcggccgccg
     1981 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2041 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2101 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2161 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2221 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     2281 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     2341 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     2401 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     2461 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     2521 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2581 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     2641 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     2701 agtccggact cagatccccc gggatgacag atgacaaaga tgtgcttcga gatgtgtggt
     2761 ttggacgaat tccaacttgc tttactctct atcaggatga gataactgaa agagaagcag
     2821 aaccatacta tttgcttttg ccaagagtca gctatttgac gttggtaact gacaaagtga
     2881 aaaagcactt tcagaaggtt atgagacaag aagatgttag tgagatatgg tttgaatatg
     2941 aaggcacacc cctgaaatgg cattatccaa ttggtttact atttgatctt cttgcatcaa
     3001 gttcagctct tccttggaac atcacagtac atttcaagag ttttccagaa aaggaccttc
     3061 tacactgtcc atccaaggat gcggttgagg ctcactttat gtcgtgtatg aaagaagctg
     3121 atgctttaaa gcataaaagt caagtgatca acgaaatgca gaaaaaagac cacaagcagc
     3181 tctggatggg actgcagaat gacagatttg accagttttg ggccatcaac cggaaactca
     3241 tggaatatcc tccagaagaa aatggatttc gttatatccc ctttagaata tatcagacca
     3301 cgacggagcg gcctttcatc cagaagctgt tccggcctgt ggccgcagat ggacagctgc
     3361 acacacttgg agatctcctc agagaagtct gtccttccgc agtcgcccct gaagatggag
     3421 agaagaggag ccaggtgatg attcacggga tagagccaat gctggaaacc cctctgcagt
     3481 ggctgagcga gcatctgagc tacccagata actttcttca tattagcatt gtcccccagc
     3541 caacagattg acccgggctg caggaattcg atatcaagct tatcgatacc gtcgacctcg
     3601 acggtaccgc gggcccggga tccaccggat ctagataact gatcataatc agccatacca
     3661 catttgtagg cggccgctac gtaaattccg cccccccccc cctctccctc ccccccccta
     3721 acgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat atgttatttt
     3781 ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct gtcttcttga
     3841 cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg
     3901 tgaaggaagc agttcctctg gaagcttctt gaagacaaac aacgtctgta gcgacccttt
     3961 gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat
     4021 aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg atagttgtgg
     4081 aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg
     4141 taccccattg tatgggatct gatctggggc ctcggtgcac atgctttaca tgtgtttagt
     4201 cgaggttaaa aaaacgtcta ggccccccga accacgggga cgtggttttc ctttgaaaaa
     4261 cacgatgata atatggccac aaccatggtt accgagtaca agcccacggt gcgcctcgcc
     4321 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     4381 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     4441 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     4501 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     4561 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     4621 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     4681 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     4741 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     4801 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     4861 tgcatgaccc gcaagcccgg tgcctgagtc gacgataaaa taaaagattt tatttagtct
     4921 ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag cttaagtaac
     4981 gccattttgc aaggcatgga aaaatacata actgagaata gagaagttca gatcaaggtc
     5041 aggaacagat ggaacagctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg
     5101 ccccggctca gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt
     5161 ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag
     5221 ccctcagcag tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg
     5281 accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc
     5341 tgctccccga gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat
     5401 tgactgagtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt
     5461 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg
     5521 tctttcacat gcagcatgta tcaaaattaa tttggttttt tttcttaagt atttacatta
     5581 aatggccata gttgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5641 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5701 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5761 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5821 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5881 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5941 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6001 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     6061 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6121 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     6181 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6241 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     6301 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6361 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6421 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6481 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttgcgg ccggccgcaa
     6541 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6601 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6661 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6721 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6781 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6841 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6901 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6961 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     7021 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7081 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7141 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7201 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7261 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7321 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7381 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7441 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7501 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttc
//
