LOCUS       sequence-388296-        7250 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of ATG9A-GFP fusion protein in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7250)
  AUTHORS   Mattera R, Park SY, De Pace R, Guardia CM, Bonifacino JS
  TITLE     AP-4 mediates export of ATG9A from the trans-Golgi network to 
            promote autophagosome formation.
  JOURNAL   Proc Natl Acad Sci U S A. 2017 Dec 12;114(50):E10697-E10706. doi: 
            10.1073/pnas.1717327114. Epub 2017 Nov 27.
  PUBMED    29180427
REFERENCE   2  (bases 1 to 7250)
  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: 
            "10.1073/pnas.1717327114"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2017-12-12- 12"; volume: "114"; issue: "50"; pages: 
            "E10697-E10706"
COMMENT     Sequence Label: pEGFPN1-ATG9A
FEATURES             Location/Qualifiers
     source          1..7250
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        111..414
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        415..618
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     569..589
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      3240..3249
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3246..3248,3249..3251,3252..3965)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3291..3312)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3552..3571)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3899..3920
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4088..4209
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4125..4144)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4179..4198
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4216..4671)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4353..4374)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4565..4584
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4698..4802
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4804..5161
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4804..4995
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4827..4847)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5012..5147
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5196..5990
                     /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             5196..5990
                     /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(5250..5269)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5860..5879
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6178..6197)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6222..6269
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6598..7186
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7087..7106
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttcctgcgtt atcccctgat tctgtggata accgtattac cgccatgcat tagttattaa
       61 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
      121 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
      181 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
      241 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
      301 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
      361 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
      421 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
      481 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
      541 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
      601 gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta ccggactcag
      661 atctcgagct caagcttcga attctgatgg cgcagttcga cactgaatac cagcgcctag
      721 aggcctccta tagtgattca cccccagggg aggaggacct gttggtgcac gtcgccgagg
      781 ggagcaagtc accttggcac catattgaaa accttgacct cttcttctct cgagtttata
      841 atctgcacca gaagaatggc ttcacatgta tgctcatcgg ggagatcttt gagctcatgc
      901 agttcctctt tgtggttgcc ttcactacct tcctggtcag ctgcgtggac tatgacatcc
      961 tatttgccaa caagatggtg aaccacagtc ttcaccctac tgaacccgtc aaggtcactc
     1021 tgccagacgc ctttttgcct gctcaagtct gtagtgccag gattcaggaa aatggctccc
     1081 ttatcaccat cctggtcatt gctggtgtct tctggatcca ccggcttatc aagttcatct
     1141 ataacatttg ctgctactgg gagatccact ccttctacct gcacgctctg cgcatcccta
     1201 tgtctgccct tccgtattgc acgtggcaag aagtgcaggc ccggatcgtg cagacgcaga
     1261 aggagcacca gatctgcatc cacaaacgtg agctgacaga actggacatc taccaccgca
     1321 tcctccgttt ccagaactac atggtggcac tggttaacaa atccctcctg cctctgcgct
     1381 tccgcctgcc tggcctcggg gaagctgtct tcttcacccg tggtctcaag tacaactttg
     1441 agctgatcct cttctgggga cctggctctc tgtttctcaa tgaatggagc ctcaaggccg
     1501 agtacaaacg tggggggcaa cggctagagc tggcccagcg cctcagcaac cgcatcctgt
     1561 ggattggcat cgctaacttc ctgctgtgcc ccctcatcct catatggcaa atcctctatg
     1621 ccttcttcag ctatgctgag gtgctgaagc gggagccggg ggccctggga gcacgctgct
     1681 ggtcactcta tggccgctgc tacctccgcc acttcaacga gctggagcac gagctgcagt
     1741 cccgcctcaa ccgtggctac aagcccgcct ccaagtacat gaattgcttc ttgtcacctc
     1801 ttttgacact gctggccaag aatggagcct tcttcgctgg ctccatcctg gctgtgctta
     1861 ttgccctcac catttatgac gaagatgtgt tggctgtgga acatgtgctg accaccgtca
     1921 cactcctggg ggtcaccgtg accgtgtgca ggtcctttat cccggaccag cacatggtgt
     1981 tctgccctga gcagctgctc cgcgtgatcc tcgctcacat ccactacatg cctgaccact
     2041 ggcagggtaa tgcccaccgc tcgcagaccc gggacgagtt tgcccagctc ttccagtaca
     2101 aggcagtgtt cattttggaa gagttgctga gccccattgt cacacccctc atcctcatct
     2161 tctgcctgcg cccacgggcc ctggagatta tagacttctt ccgaaacttc accgtggagg
     2221 tcgttggtgt gggagatacc tgctcctttg ctcagatgga tgttcgccag catggtcatc
     2281 cccagtggct atctgctggg cagacagagg cctcagtgta ccagcaagct gaggatggaa
     2341 agacagagtt gtcactcatg cactttgcca tcaccaaccc tggctggcag ccaccacgtg
     2401 agagcacagc cttcctaggc ttcctcaagg agcaggttca gcgggatgga gcagctgcta
     2461 gcctcgccca agggggtctg ctccctgaaa atgccctctt tacgtctatc cagtccttac
     2521 aatctgagtc tgagcccctg agccttatcg caaatgtggt agctggctca tcctgccggg
     2581 gccctccact gcccagagac ctgcagggct ccaggcacag ggctgaagtc gcctctgccc
     2641 tgcgctcctt ctccccgctg caacccgggc aggcgcccac aggccgggct cacagcacca
     2701 tgacaggctc tggggtggat gccaggacag ccagctccgg gagcagcgtg tgggaaggac
     2761 agctgcagag cctggtgctg tcagaatatg catccacaga gatgagcctg catgccctct
     2821 atatgcacca gctccacaag cagcaggccc aggctgaacc tgagcggcat gtatggcacc
     2881 gccgggagag tgatgagagt ggagaaagcg cccctgatga agggggagag ggcgcccggg
     2941 ccccccagtc tatccctcgc tctgctagct atccctgtgc agcaccccgg cctggagctc
     3001 ctgagaccac cgccctgcat gggggcttcc agaggcgcta cggtggcatc acagatcctg
     3061 gcacagtgcc cagggttccc tctcatttct ctcggctgcc tcttggaggg tgggcagaag
     3121 atgggcagtc ggcatcaagg caccctgagc ccgtgcccga agagggctcg gaggatgagc
     3181 taccccctca ggtgcacaag gtaacgtcga cggtaccgcg ggcccgggat ccaccggtcg
     3241 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     3301 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     3361 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     3421 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     3481 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     3541 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     3601 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     3661 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     3721 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     3781 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3841 accactacct gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca
     3901 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3961 agtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag aggttttact
     4021 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     4081 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     4141 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4201 tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     4261 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     4321 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     4381 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     4441 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     4501 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     4561 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     4621 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     4681 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4741 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4801 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     4861 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     4921 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4981 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     5041 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     5101 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     5161 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     5221 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     5281 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     5341 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     5401 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     5461 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     5521 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     5581 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     5641 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     5701 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     5761 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     5821 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     5881 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     5941 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     6001 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     6061 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     6121 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     6181 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     6241 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     6301 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     6361 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     6421 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     6481 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6541 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     6601 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     6661 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     6721 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     6781 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     6841 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6901 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6961 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     7021 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     7081 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     7141 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     7201 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
//
