LOCUS       sequence-489404-        4999 bp    DNA     circular SYN 15-OCT-2025
DEFINITION  Express RAB10 in mammalian cells with 2xmyc tag and T73A mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4999)
  AUTHORS   Bonet-Ponce L, Tegicho T, Fernandez-Martinez N, Rozenberg IA, 
            Ashriem M, Beilina A, Kluss JH, Li Y, Cookson MR
  TITLE     JIP4 and RILPL1 utilize opposing motor force to dynamically regulate
            lysosomal tubulation.
  JOURNAL   J Cell Biol. 2025 Nov 3;224(11):e202404018. doi: 
            10.1083/jcb.202404018. Epub 2025 Sep 24.
  PUBMED    40990465
REFERENCE   2  (bases 1 to 4999)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2025-11-3"; volume: "224(11):e202404018. doi"; pages:
            " 10.1083/jcb.202404018. Epub 2025 Sep 24"
COMMENT     Sequence Label: 2xmyc-RAB10 (T73A)
FEATURES             Location/Qualifiers
     source          1..4999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        67..370
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        371..574
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     525..545
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        620..638
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      670..679
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             679..708
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             724..753
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    772..796
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1427..1451)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(1500..1519)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1501..1519)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    1793..1914
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1830..1849)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1884..1903
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1921..2376)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2058..2079)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2270..2289
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2403..2507
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2509..2866
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2509..2700
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2532..2552)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2717..2852
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2901..3695
                     /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             2901..3695
                     /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(2955..2974)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3565..3584
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3883..3902)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3927..3974
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4303..4891
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4792..4811
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcattagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
       61 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
      121 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
      181 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      241 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
      301 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      361 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      421 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      481 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      541 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta
      601 gcgattacgc caagctcgaa attaaccctc actaaaggga acaaaagctg gagctccacc
      661 gcggtggcgg ccgccatgga gcagaaactc atctctgaag aggatctgag cccgggcgga
      721 tccgagcaga aactcatctc tgaagaggat ctgctgcagg aattcgatat cacaagtttg
      781 tacaaaaaag caggctccga attcgccctt atggcgaaga agacgtacga cctgcttttc
      841 aagctgctcc tgatcgggga ttccggagtg gggaagacct gcgtcctttt tcgtttttcg
      901 gatgatgcct tcaatactac ctttatttcc accataggaa tagacttcaa gatcaaaaca
      961 gttgaattac aaggaaagaa gatcaagcta cagatatggg atacagcagg ccaggagcga
     1021 tttcacgcca tcacaacctc ctactacaga ggcgcaatgg gtatcatgct agtatatgac
     1081 atcaccaatg gtaaaagttt tgaaaacatc agcaaatggc ttagaaacat agatgagcat
     1141 gccaatgaag atgtggaaag aatgttacta ggaaacaagt gtgatatgga cgacaaaaga
     1201 gttgtaccta aaggaaaagg agaacagatt gcaagggagc atggtattag gttttttgag
     1261 actagtgcaa aagcaaatat aaacatcgaa aaggcgttcc tcacgttagc tgaagatatc
     1321 cttcgaaaga cccctgtaaa agagcccaac agtgaaaatg tagatatcag cagtggagga
     1381 ggcgtgacag gctggaagag caaatgctgc tgaaagggcg aattcgaccc agctttcttg
     1441 tacaaagtgg tgatcggtac cttaattaat taaggtacca ggtaagtgta cccaattcgc
     1501 cctatagtga gtcgtattac aattcactcg atcgcccttc ccaacagttg cgcagcctga
     1561 atggcgaatg gagatccaat ttttaagtgt ataatgtgtt aaactactga ttctaattgt
     1621 ttgtgtattt tagattcaca gtcccaaggc tcatttcagg cccctcagtc ctcacagtct
     1681 gttcatgatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct
     1741 cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     1801 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1861 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaacgcgt
     1921 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     1981 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     2041 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     2101 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     2161 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     2221 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     2281 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     2341 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg
     2401 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     2461 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagaatcct gaggcggaaa
     2521 gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     2581 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     2641 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     2701 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     2761 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     2821 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga
     2881 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     2941 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     3001 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     3061 ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg
     3121 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     3181 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     3241 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     3301 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     3361 atcaggatga tctggacgaa gaacatcagg ggctcgcgcc agccgaactg ttcgccaggc
     3421 tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     3481 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     3541 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gaacttggcg
     3601 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     3661 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     3721 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     3781 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     3841 tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac
     3901 aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt
     3961 tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc
     4021 accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc
     4081 aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc
     4141 ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4201 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4261 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4321 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4381 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4441 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4501 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4561 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4621 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4681 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4741 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4801 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4861 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4921 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4981 ggataaccgt attaccgcc
//
