LOCUS       sequence-220257-        7480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing GFP-YKT6 for mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7480)
  AUTHORS   Matsui T, Jiang P, Nakano S, Sakamaki Y, Yamamoto H, Mizushima N
  TITLE     Autophagosomal YKT6 is required for fusion with lysosomes 
            independently of syntaxin 17.
  JOURNAL   J Cell Biol. 2018 Aug 6;217(8):2633-2645. doi: 
            10.1083/jcb.201712058. Epub 2018 May 22.
  PUBMED    29789439
REFERENCE   2  (bases 1 to 7480)
  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: 
            "10.1083/jcb.201712058"; journalName: "J Cell Biol"; date: 
            "2018-08-6- 6"; volume: "217"; issue: "8"; pages: "2633-2645"
COMMENT     Sequence Label: pMRXIP-GFP-YKT6
FEATURES             Location/Qualifiers
     source          1..7480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    667..1024
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1089..1505
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1435..1457
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1473..1489
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1515..1889
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1827..1843
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             join(1901..1903,1904..1906,1907..2617)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1901..1903,1904..1906,1907..2617)
                     /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(1946..1967)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2207..2226)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2554..2575
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3331..3897
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3347..3899
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3491..3508)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3718..3737
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3911..4510
                     /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(3911..3930)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4407..4427
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4734..5203
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5333..5350)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5504..6092)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5584..5603)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6256..7047,7048..7116))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6879..6898
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7117..7221)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7289..7307
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(7409..7480,1..308)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
       61 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      121 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      181 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
      241 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      301 ttaccatgct agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct
      361 gtgccttatt tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc
      421 cccgagctca ataaaagagc ccacaacccc tcactcggcg cgccagtcct ccgattgact
      481 gagtcgcccg ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt
      541 ctcgctgttc cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt
      601 catttggggg ctcgtccggg atcgggagac ccctgcccag ggaccaccga cccaccaccg
      661 ggaggtaagc tggccagcaa cttatctgtg tctgtccgat tgtctagtgt ctatgactga
      721 ttttatgcgc ctgcgtcggt actagttagc taactagctc tgtatctggc ggacccgtgg
      781 tggaactgac gagttcggaa cacccggccg caaccctggg agacgtccca gggacttcgg
      841 gggccgtttt tgtggcccga cctgagtcca aaaatcccga tcgttttgga ctctttggtg
      901 cacccccctt agaggaggga tatgtggttc tggtaggaga cgagaaccta aaacagttcc
      961 cgcctccgtc tgaatttttg ctttcggttt gggaccgaag ccgcgccgcg cgtcttgtct
     1021 gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt tctgtatttg tctgaaaata
     1081 tgggcccggg ccagactgtt accactccct taagtttgac cttaggtcac tggaaagatg
     1141 tcgagcggat cgctcacaac cagtcggtag atgtcaagaa gagacgttgg gttaccttct
     1201 gctctgcaga atggccaacc tttaacgtcg gatggccgcg agacggcacc tttaaccgag
     1261 acctcatcac ccaggttaag atcaaggtct tttcacctgg cccgcatgga cacccagacc
     1321 aggtccccta catcgtgacc tgggaagcct tggcttttga cccccctccc tgggtcaagc
     1381 cctttgtaca ccctaagcct ccgcctcctc ttcctccatc cgccccgtct ctcccccttg
     1441 aacctcctcg ttcgaccccg cctcgatcct ccctttatcc agccctcact ccttctctag
     1501 gcgcccccat atggccatat gagatcttat atggggcacc cccgcccctt gtaaacttcc
     1561 ctgaccctga catgacaaga gttactaaca gcccctctct ccaagctcac ttacaggctc
     1621 tctacttagt ccagcacgaa gtctggagac ctctggcggc agcctaccaa gaacaactgg
     1681 accgaccggt ggtacctcac ccttaccgag tcggcgacac agtgtgggtc cgccgacacc
     1741 agactaagaa cctagaacct cgctggaaag gaccttacac agtcctgctg accaccccca
     1801 ccgccctcaa agtagacggc atcgcagctt ggatacacgc cgcccacgtg aaggctgccg
     1861 accccggggg tggaccatcc tctagactgc cggatctgcc atggtgagca agggcgagga
     1921 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa
     1981 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt
     2041 catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta
     2101 cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc
     2161 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta
     2221 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa
     2281 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa
     2341 cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa
     2401 gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac
     2461 ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc
     2521 cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc
     2581 cgccgggatc actctcggca tggacgagct gtacaagggt ggcggtggct cgggcggtgg
     2641 tgggtcgggt ggcggcggat ctatcgatga attgaattca cgcgttcgcg agttaacgga
     2701 tccctcgagc ggccgcatga agctgtacag cctcagcgtc ctctacaaag gcgaggccaa
     2761 ggtggtgctg ctcaaagccg catacgatgt gtcttccttc agctttttcc agagatccag
     2821 cgttcaggaa ttcatgacct tcacgagtca actgattgtg gagcgctcat cgaaaggcac
     2881 tagagcttct gtcaaagaac aagactatct gtgccacgtc tacgtccgga atgatagtct
     2941 tgcaggtgtg gtcattgctg acaatgaata cccatcccgg gtggccttta ccttgctgga
     3001 gaaggtacta gatgaattct ccaagcaagt cgacaggata gactggccag taggatcccc
     3061 tgctacaatc cattacccag ccctggatgg tcacctcagt agataccaga acccacgaga
     3121 agctgatccc atgactaaag tgcaggccga actagatgag accaaaatca ttctgcacaa
     3181 caccatggag tctctgttag agcgaggtga gaagctagat gacttggtgt ccaaatccga
     3241 ggtgctggga acacagtcta aagccttcta taaaactgcc cggaaacaaa actcatgctg
     3301 tgccatcatg tgagcggccg ctacgtaaat tccgcccccc cccctaacgt tactggccga
     3361 agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg
     3421 tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg
     3481 ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt
     3541 cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac
     3601 cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca
     3661 aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg
     3721 ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg
     3781 ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag gttaaaaaaa
     3841 cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg atgataatat
     3901 ggccacaacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     3961 ccgggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     4021 cgacccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     4081 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac
     4141 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     4201 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     4261 gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa
     4321 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     4381 cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     4441 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     4501 cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg cacgacccca
     4561 tggctcgacg ataaaataaa agattttatt tagtctccag aaaaaggggg gaatgaaaga
     4621 ccccacctgt aggtttggca agctagctta agtaacgcca ttttgcaagg catggaaaaa
     4681 tacataactg agaatagaga agttcagatc aaggtcagga acagatggaa cagctgaata
     4741 tgggccaaac aggatatctg tggtaagcag ttcctgcccc ggctcagggc caagaacaga
     4801 tggaacagct gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc
     4861 agggccaaga acagatggtc cccagatgcg gtccagccct cagcagtttc tagagaacca
     4921 tcagatgttt ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac
     4981 caatcagttc gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag
     5041 cccacaaccc ctcactcggg gcgccagtcc tccgattgac tgagtcgccc gggtacccgt
     5101 gtatccaata aaccctcttg cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg
     5161 gtctcctctg agtgattgac tacccgtcag cgggggtctt tcacatgcag catgtatcaa
     5221 aattaatttg gttttttttc ttaagtattt acattaaatg gccatagttg cattaatgaa
     5281 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     5341 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     5401 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     5461 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     5521 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     5581 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     5641 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     5701 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     5761 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5821 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5881 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5941 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     6001 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     6061 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     6121 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     6181 ggatcttcac ctagatcctt ttgcggccgg ccgcaaatca atctaaagta tatatgagta
     6241 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6301 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6361 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6421 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6481 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6541 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6601 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6661 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6721 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6781 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6841 gcggcgaccg agttgctctt gcccggcgtc aacacgggat aataccgcgc cacatagcag
     6901 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6961 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7021 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7081 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7141 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7201 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     7261 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtcttca
     7321 ctcgaggagc ttggcccatt gcatacgttg tatccatatc ataatatgta catttatatt
     7381 ggctcatgtc caacattacc gccatgttga cattgattat tgactagtta ttaatagtaa
     7441 tcaattacgg ggtcattagt tcatagccca tatatggagt
//
