LOCUS       sequence-472438-        7683 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of FL human VPS34 REIE > AAAA mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7683)
  AUTHORS   Tremel S, Ohashi Y, Morado DR, Bertram J, Perisic O, Brandt LTL, von
            Wrisberg MK, Chen ZA, Maslen SL, Kovtun O, Skehel M, Rappsilber J, 
            Lang K, Munro S, Briggs JAG, Williams RL
  TITLE     Structural basis for VPS34 kinase activation by Rab1 and Rab5 on 
            membranes.
  JOURNAL   Nat Commun. 2021 Mar 10;12(1):1564. doi: 10.1038/s41467-021-21695-2.
  PUBMED    33692360
REFERENCE   2  (bases 1 to 7683)
  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: "Nat 
            Commun."; date: "2021-03-10"; volume: "12(1):1564. doi"; pages: " 
            10.1038/s41467-021-21695-2"
COMMENT     Sequence Label: pYO1274
FEATURES             Location/Qualifiers
     source          1..7683
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     complement(3691..3708)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3697..3921
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3967..4395
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4054..4073)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4264..4285
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4404..4424)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4409..4738
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4589..4724
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4651..4670
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4786..4833
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4852..5226
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    5356..5477
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5393..5412)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5447..5466
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5526..5542)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5526..5542)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5539..5561)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5550..5566
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5574..5580,5581..5598,5599..5604))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5619..5640
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5757..5774)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5928..6516)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6008..6027)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6687..7478,7479..7547))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7310..7329
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7548..7652)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtacca tgggggaagc agagaagttt cactacatct
     1021 atagttgtga cctggatatc aacgtccagc ttaagatagg aagcttggaa gggaagagag
     1081 aacaaaagag ttataaagct gtcctggaag acccaatgtt gaagttctca ggactatatc
     1141 aagagacatg ctctgatctt tatgttactt gtcaagtttt tgcagaaggg aagcctttgg
     1201 ccttgccagt gagaacatcc tacaaagcat ttagtacaag atggaactgg aatgaatggc
     1261 tgaaactacc agtaaaatac cctgacctgc ccaggaatgc ccaagtggcc ctcaccatct
     1321 gggatgtgta tggtcccgga aaagcagtgc ctgtaggagg aacaacggtt tcgctctttg
     1381 gaaaatacgg catgtttcgc caagggatgc atgacttgaa agtctggcct aatgtagaag
     1441 cagatggatc agaacccaca aaaactcctg gcagaacaag tagcactctc tcagaagatc
     1501 agatgagccg tcttgccaag ctcaccaaag ctcatcgaca aggacacatg gtgaaagtag
     1561 attggctgga tagattgaca tttgctgcag ctgcaatgat aaatgagagt gaaaaacgaa
     1621 gttctaattt catgtacctg atggttgaat ttcgatgtgt caagtgtgat gataaggaat
     1681 atggtattgt ttattatgaa aaggacggtg atgaatcatc tccaatttta acaagttttg
     1741 aattagtgaa agttcctgac ccccagatgt ctatggagaa tttagttgag agcaaacacc
     1801 acaagcttgc ccggagttta agaagtggac cttctgacca cgatctgaaa cccaatgctg
     1861 ccacgagaga tcagttaaat attattgtga gttatccacc aaccaagcaa cttacctatg
     1921 aagaacaaga tcttgtttgg aagtttagat attatcttac gaatcaagaa aaagccttga
     1981 caaaattctt gaaatgtgtt aattgggatc tacctcaaga ggccaaacag gccttggaac
     2041 ttctgggaaa atggaagccg atggatgtag aggactcctt ggagctgtta tcctctcatt
     2101 acaccaaccc aactgtgagg cgttatgctg ttgcccggtt gcgacaggcc gatgatgagg
     2161 atttgttgat gtacctatta caattggtcc aggctctcaa atatgaaaat tttgatgata
     2221 taaagaatgg attggaacct accaagaagg atagtcagag ttcagtgtca gaaaatgtgt
     2281 caaattctgg aataaattct gcagaaatag atagctccca aattataacc agcccccttc
     2341 cttcagtctc ttcacctcct cctgcatcaa aaacaaaaga agttccagat ggcgaaaatc
     2401 tggaacaaga tctctgtacc ttcttgatat cgagagcctg caaaaactca acactggcta
     2461 attatttata ctggtatgtg atagtggaat gtgaagatca agatactcag cagagagatc
     2521 caaagaccca tgagatgtac ttgaacgtaa tgagaagatt cagccaagca ttgttgaagg
     2581 gtgataagtc tgtcagagtt atgcgttctt tgctggctgc acaacagaca tttgtagatc
     2641 ggttggtgca tctaatgaag gcagtacaac gcgaaagtgg aaatcgtaag aaaaagaatg
     2701 agagactaca ggcattgctt ggagataatg aaaagatgaa tttgtcagat gtggaactta
     2761 tcccgttgcc tttagaaccc caagtgaaaa ttagaggaat aattccggaa acagctacac
     2821 tgtttaaaag tgcccttatg cctgcacagt tgttttttaa gacggaagat ggaggcaaat
     2881 atccagttat atttaagcat ggagatgatt tacgtcaaga tcaacttatt cttcaaatca
     2941 tttcactcat ggacaagctg ttacggaaag aaaatctgga cttgaaattg acaccttata
     3001 aggtgttagc caccagtaca aaacatggct tcatgcagtt tatccagtca gttcctgtgg
     3061 ctgaagttct tgatacagag ggaagcattc agaacttttt tagaaaatat gcaccaagtg
     3121 agaatgggcc aaatgggatt agtgctgagg tcatggacac ttacgttaaa agctgtgctg
     3181 gatattgcgt gatcacctat atacttggag ttggagacag gcacctggat aaccttttgc
     3241 taacaaaaac aggcaaactc ttccacatag actttggata tattctgggt cgtgatccaa
     3301 agcctcttcc tccaccaatg aagctgaata aagaaatggt agaaggaatg gggggcacac
     3361 agagtgagca gtaccaagag ttccgtaaac agtgttacac ggctttcctc cacctgcgaa
     3421 ggtattctaa tctgattttg aacttgtttt ccttgatggt tgatgcaaac attccagata
     3481 ttgcacttga accagataaa actgtgaaaa aggttcagga taaattccgc ttagacctgt
     3541 cggatgaaga ggctgtgcat tacatgcaga gtctgattga tgagagtgtc catgctcttt
     3601 ttgctgcagt ggtggaacag attcacaagt ttgcccagta ctggcgcaaa tgagtcgagt
     3661 ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat
     3721 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     3781 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     3841 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     3901 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     3961 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     4021 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     4081 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     4141 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     4201 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     4261 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     4321 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     4381 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     4441 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     4501 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     4561 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     4621 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     4681 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     4741 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
     4801 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag
     4861 ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg
     4921 accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg
     4981 gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg
     5041 gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc
     5101 acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg
     5161 cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag
     5221 gactgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     5281 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     5341 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5401 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5461 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     5521 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5581 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5641 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5701 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5761 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5821 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5881 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5941 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     6001 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     6061 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     6121 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     6181 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     6241 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     6301 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     6361 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     6421 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     6481 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     6541 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     6601 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     6661 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     6721 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     6781 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6841 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6901 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6961 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     7021 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     7081 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     7141 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     7201 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     7261 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     7321 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     7381 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     7441 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     7501 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     7561 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     7621 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     7681 gtc
//
