LOCUS       sequence-198005-        6870 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Point mutation in APPL1 that diminishes its interaction with Rab5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6870)
  TITLE     Webb Lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6870)
  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: 
            "Unpublished"
COMMENT     Sequence Label: APPL1-N308D-GFP
FEATURES             Location/Qualifiers
     source          1..6870
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     56..75
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(164..521)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      178..313
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        330..521
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     478..498
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(523..627)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      654..1109
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(741..760)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     951..972
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    1116..1237
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1127..1146)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1181..1200
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(3566..4276,4277..4279,4280..4282))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(3566..4276,4277..4279,4280..4282))
                     /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(3608..3629)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3957..3976
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4216..4237
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      4279..4288
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4327..4530)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4356..4376)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4531..4834
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(5009..5597)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5089..5108)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    5926..5973
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     5998..6017
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(join(6205..6870,1..129))
                     /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             complement(join(6205..6870,1..129))
                     /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(6316..6335)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 cccctgcgct gacagccgga acacggcggc atcagagcag ccgattgtct gttgtgccca
       61 gtcatagccg aatagcctct ccacccaagc ggccggagaa cctgcgtgca atccatcttg
      121 ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatcg atctttgcaa aagcctaggc
      181 ctccaaaaaa gcctcctcac tacttctgga atagctcaga ggccgaggcg gcctcggcct
      241 ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa ctgggcggag
      301 ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa ttgagatgca
      361 tgctttgcat acttctgcct gctggggagc ctggggactt tccacacctg gttgctgact
      421 aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac tttccacacc
      481 ctaactgaca cacattccac agctggttct ttccgcctca ggactcttcc tttttcaata
      541 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
      601 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgcgcc
      661 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
      721 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
      781 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
      841 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
      901 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
      961 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     1021 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     1081 ttttaacaaa atattaacgc ttacaattta cgcgttaaga tacattgatg agtttggaca
     1141 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc
     1201 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     1261 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa
     1321 atgtggtatg gctgattatg atcagttatc tagatccggt ggatcccggg cccgcggtac
     1381 cgtcgactgc agaattcggc tttgcttctg attctctctt ctttcctcct tctcccaaat
     1441 cactctcttc tgactggcta ctgctaagga caacaaactg cccctcacta ctggcttggt
     1501 ttggtctact ggaagcagct atcaaccgac tttgttcttc caagtccttt tcaatctgtt
     1561 tttgtttatt gagttctttc tgttgcttct cttttactct ctctatttct ttttgttttt
     1621 ctgatgccct acgatccagt tcagcatgca aagctatctg ttttgccagt ccaacagaat
     1681 cacatatctt ttccccctca ttgtttgact caaatatata gcagactgat gacagattac
     1741 tttcacttct cccgcttgat gtccgaagaa caaatccaaa aaggcgctta ttttcctggt
     1801 gtgtagcata caaaactaca caaggtaatg gaaacgtgag ccttgtaact tgtgtctgtg
     1861 gatcaattaa ctttaaacag tcacaagtga ctaataaatg cgattctgtc atacgaaaga
     1921 tgttatggat ggcccgggca gctaagattt ggcgcatagt ttcataaaca acatctggat
     1981 ggtcatctga tttcacctcc attgaaccaa ggaatcggac aataaataac tgatgaagaa
     2041 tagaatcttc agtttcagat tttgtacttc ctccagattc tccaaatgga tttgtacgcc
     2101 tgcctccctg gccaaaggct tttgcctggc caggctgatc ttcacacaca ggagaaatta
     2161 tgtcaaactg tattggggtg tctggggcaa caagagaatc tagagagagg gcagccaaat
     2221 ttgtagactc agatcctaag gatcctgaac tgctggttcg agctgtcggt ggccgagatt
     2281 gtcctgctgc tggccgcagg ctctcgtgcc tctgctggaa agatggggaa ggagtgacag
     2341 cttccagagc tgattgattt actcgtgcag cagtttcctc tggattttca cttaagtata
     2401 tttgtttaga tatgttgttt attgtacaga tccactcttc atgatctttt ttactctctg
     2461 cttgcaaaat tgaagatttt tttccatcga aagaggtgat ctgaaaacaa tatcgtctgt
     2521 cttcacagtc cacagccatc actgaacagt tgtctatgtc catggccagg cctcctgcta
     2581 catccccacg ggcctgactc attaaatctc caccctgcgt gaagtaaaac tgtctgtccc
     2641 aggtagatga caccaagcct gttttattcc tagcattaag gtatccagcc tttcgggtta
     2701 aatttcgatt aacaggaaat ttggtggggt ctgggtcagg cacatataag ggatcactgg
     2761 ctacttccaa atcctctatt gtctgttgca tggtctctat atcactgtcc atttccctgc
     2821 gaacattctg aacgcttgtt ccaatattag ctaaaaattc ttccagttgt tcattaagat
     2881 tttcagaacc catcttaaag aaacttatct gagcttgcat gtacccaagt agaggttcta
     2941 acaatgctat tttcttcttg tactgaagag tatttaatgc acaaaaataa tgcatcatgg
     3001 tctggtgttg tttctttctg gatgtgtaca catcttctgt tacttcatac ttcaccttgt
     3061 cattttctct tttttttgat aaacggctat atctattaat cgcagcatca tgatcattac
     3121 ttgcaatctg aaatacttcc tttaatgtta gtatttcttt cagatctctt tctttaaact
     3181 gggtaatggg gaacatcatg gcatcagcaa gttgagttga aagcactgca tgacaagagc
     3241 taagctcatc tataactttt gaaaactgtt gcaatgtaga gctcataact tcatcatcac
     3301 ctcccaatgg aaaacgctgt ttttcatatt cttttaaaag ttttgaggtc aggtgtgttg
     3361 ctgcacttaa ttcattctgt gcatcataaa tccgatgcat agcttgatac aactggttca
     3421 tatagttgga aatagctgtg gcatcttctt caaatacacc tagtaaagac cttgtctgcg
     3481 ggctgtcctc cagcgtctcc tcgatgggca gcttgtcgat ccccggcata agccgaattc
     3541 gaagcttgag ctcgagatct gagtacttgt acagctcgtc catgccgaga gtgatcccgg
     3601 cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca
     3661 gggcggactg ggtgctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg
     3721 gggtgttctg ctggtagtgg tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga
     3781 tcttgaagtt caccttgatg ccgttcttct gcttgtcggc catgatatag acgttgtggc
     3841 tgttgtagtt gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc
     3901 ccttcagctc gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct
     3961 tgtagttgcc gtcgtccttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg
     4021 cggacttgaa gaagtcgtgc tgcttcatgt ggtcggggta gcggctgaag cactgcacgc
     4081 cgtaggtcag ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga
     4141 tgaacttcag ggtcagcttg ccgtaggtgg catcgccctc gccctcgccg gacacgctga
     4201 acttgtggcc gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca
     4261 gctcctcgcc cttgctcacc atggtggcga ccggtagcgc tagcggatct gacggttcac
     4321 taaaccagct ctgcttatat agacctccca ccgtacacgc ctaccgccca tttgcgtcaa
     4381 tggggcggag ttgttacgac attttggaaa gtcccgttga ttttggtgcc aaaacaaact
     4441 cccattgacg tcaatggggt ggagacttgg aaatccccgt gagtcaaacc gctatccacg
     4501 cccattgatg tactgccaaa accgcatcac catggtaata gcgatgacta atacgtagat
     4561 gtactgccaa gtaggaaagt cccataaggt catgtactgg gcataatgcc aggcgggcca
     4621 tttaccgtca ttgacgtcaa tagggggcgt acttggcata tgatacactt gatgtactgc
     4681 caagtgggca gtttaccgta aatactccac ccattgacgt caatggaaag tccctattgg
     4741 cgttactatg ggaacatacg tcattattga cgtcaatggg cgggggtcgt tgggcggtca
     4801 gccaggcggg ccatttaccg taagttatgt aacgcggaac tccatatatg ggctatgaac
     4861 taatgacccc gtaattgatt actattaata actaatgcat ggcggtaata cggttatcca
     4921 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     4981 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     5041 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5101 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5161 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5221 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5281 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5341 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5401 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5461 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5521 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5581 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5641 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5701 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taacctgagg
     5761 ctatggcagg gcctgccgcc ccgacgttgg ctgcgagccc tgggccttca cccgaacttg
     5821 gggggtgggg tggggaaaag gaagaaacgc gggcgtattg gccccaatgg ggtctcggtg
     5881 gggtatcgac agagtgccag ccctgggacc gaaccccgcg tttatgaaca aacgacccaa
     5941 caccgtgcgt tttattctgt ctttttattg ccgtcatagc gcgggttcct tccggtattg
     6001 tctccttccg tgtttcagtt agcctccccc tagggtgggc gaagaactcc agcatgagat
     6061 ccccgcgctg gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt
     6121 catagaaggc ggcggtggaa tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg
     6181 tcatttcgaa ccccagagtc ccgctcagaa gaactcgtca agaaggcgat agaaggcgat
     6241 gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc
     6301 gccaagctct tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac
     6361 acccagccgg ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg
     6421 caagcaggca tcgccatggg tcacgacgag atcctcgccg tcgggcatgc tcgccttgag
     6481 cctggcgaac agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc
     6541 gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc
     6601 gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga
     6661 tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa
     6721 tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc
     6781 cgtcgtggcc agccacgata gccgcgctgc ctcgtcttgc agttcattca gggcaccgga
     6841 caggtcggtc ttgacaaaaa gaaccgggcg
//
