LOCUS       sequence-492228-        5816 bp    DNA     circular SYN 06-NOV-2025
DEFINITION  mammalian expression of Rab12 T56N, constitutively GDP-bound mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5816)
  AUTHORS   Xu J, Fotouhi M, McPherson PS
  TITLE     Phosphorylation of the exchange factor DENND3 by ULK in response to 
            starvation activates Rab12 and induces autophagy.
  JOURNAL   EMBO Rep. 2015 Jun;16(6):709-18. doi: 10.15252/embr.201440006. Epub 
            2015 Apr 29.
  PUBMED    25925668
REFERENCE   2  (bases 1 to 5816)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 6, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.15252/embr.201440006"; journalName: "EMBO Rep"; date: "2015-06";
            volume: "16"; issue: "6"; pages: "709-18"
COMMENT     Sequence Label: Flag Rab12 T56N
FEATURES             Location/Qualifiers
     source          1..5816
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(20..36)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(20..36)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(58..74)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(59..75)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(130..149)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(131..149)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    423..544
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(460..479)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     514..533
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(551..1006)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(688..709)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     900..919
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1033..1137
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1139..1496
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1139..1330
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1162..1182)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1347..1482
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1531..2325
                     /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             1531..2325
                     /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(1585..1604)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2195..2214
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2513..2532)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2557..2604
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2933..3521
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3422..3441
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        3696..3999
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4000..4203
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4154..4174
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        4249..4267
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      4302..4311
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4311..4334
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
ORIGIN
        1 gctctagatt gcggccgcgg tcatagctgt ttcctgaaca gatcccatca agcttatcga
       61 taccgtcgac ctcgaggggg ggcccggtac cttaattaat taaggtacca ggtaagtgta
      121 cccaattcgc cctatagtga gtcgtattac aattcactcg atcgcccttc ccaacagttg
      181 cgcagcctga atggcgaatg gagatccaat ttttaagtgt ataatgtgtt aaactactga
      241 ttctaattgt ttgtgtattt tagattcaca gtcccaaggc tcatttcagg cccctcagtc
      301 ctcacagtct gttcatgatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
      361 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
      421 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
      481 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
      541 cttaacgcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      601 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      661 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      721 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      781 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      841 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      901 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      961 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt
     1021 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     1081 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagaatcct
     1141 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     1201 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     1261 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     1321 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     1381 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     1441 ctgggctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc
     1501 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     1561 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     1621 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     1681 accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg
     1741 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     1801 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     1861 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     1921 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     1981 ggtcttgtcg atcaggatga tctggacgaa gaacatcagg ggctcgcgcc agccgaactg
     2041 ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     2101 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     2161 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     2221 gaacttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     2281 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     2341 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     2401 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     2461 agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac
     2521 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     2581 cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg
     2641 tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac
     2701 cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc
     2761 ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat
     2821 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     2881 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     2941 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     3001 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     3061 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     3121 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     3181 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     3241 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     3301 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     3361 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     3421 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     3481 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     3541 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     3601 ctgattctgt ggataaccgt attaccgcca tgcattagtt attaatagta atcaattacg
     3661 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     3721 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     3781 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     3841 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     3901 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     3961 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
     4021 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     4081 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     4141 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     4201 gctggtttag tgaaccgtca gatccgctag cgattacgcc aagctcgaaa ttaaccctca
     4261 ctaaagggaa caaaagctgg agctccaccg cggtggcggc cgccaccatg gattacaagg
     4321 atgacgacga taaggcccgg gcggatccgg gcgccgcgct gcagaggcgg gccgggggcg
     4381 gcggcggtct gggcgcgggc tccccggcgc tgtcgggcgg ccagggccgc cggaggaagc
     4441 agccccccag gccggccgac ttcaagttgc aggtcatcat tatcggctcc cgcggcgtgg
     4501 gcaagaacag cctgatggag cgcttcaccg acgacacctt ctgcgaggcc tgcaagtcca
     4561 ccgtgggtgt tgacttcaaa atcaaaactg tagagctaag aggaaagaaa attagattac
     4621 agatctggga cacagcaggt caggagagat tcaacagcat tacctcagct tattacagaa
     4681 gtgccaaggg gatcatatta gtatatgata tcactaagaa ggagacattt gatgatttgc
     4741 cgaaatggat gaagatgatt gataagtatg cttcagaaga tgcagagctt ctcttagttg
     4801 gaaataagtt ggactgtgaa acggacagag aaatcaccag gcagcagggg gaaaagtttg
     4861 cacagcagat cactgggatg cggttctgtg aagcaagtgc caaggataac ttcaatgtgg
     4921 acgagatatt tttgaaactt gtcgatgaca ttctgaaaaa gatgcctctg gatattttaa
     4981 ggaatgagtt gtccaatagt atcctgtcgt tacaaccaga gcctgagata ccgccagaac
     5041 tgcctccacc aagaccacat gtccgatgct gttgatttcc tactttggag acaaagtgga
     5101 aatgattcct ggaaagggga aaaaacgttc tattctgcac tacaatcatt ttgacaattt
     5161 cctttcgcac tttgtaatcc aagtcagagc tatacactaa cttgtaaata tgcatatatg
     5221 caatcctggg taagttttgg ttataagtta cctatttccc tccaaattat tatatttcat
     5281 tcattacccc agtgtctagt gtacatacac tgggaaacct agtacttcta atatgaagaa
     5341 tgggagaaat gaaaggtata atgtttcttg aaataaataa tataattgtc cttattaatt
     5401 atattatgag gacagaagat attctgataa gagagaacgt ggtgctttgc ttaccgtttt
     5461 aaagaaaatt tgtaaaacta aagacttttt gaaaaaaagc tatcttaagt gctttttctt
     5521 tatttacaag acatttcccc cagtggtagc atctgaagta ttggagtgtt tctgccacga
     5581 agcaaagctc cattcatggc cgtcatggaa ggttatttat taatgttaca taatggtaga
     5641 atattactag ttagagggtt ggatttgact tggtcctaag gccacagaat ctctctcatg
     5701 gcttcctaag ggatgtacct ttatgctttt aagaactaca aagattcaat aaagaaagaa
     5761 atgtttttga aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaagg gcggcc
//
