LOCUS       sequence-205812-        8003 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector that codes for FLAG-RAB35 wildtype (human) 
            protein. Puromycin resistance for mammalian selection and ampicillin
            resistance for growth in Stbl3 cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8003)
  AUTHORS   Wheeler DB, Zoncu R, Root DE, Sabatini DM, Sawyers CL
  TITLE     Identification of an oncogenic RAB protein.
  JOURNAL   Science. 2015 Oct 9;350(6257):211-7. doi: 10.1126/science.aaa4903. 
            Epub 2015 Sep 3.
  PUBMED    26338797
REFERENCE   2  (bases 1 to 8003)
  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: 
            "10.1126/science.aaa4903"; journalName: "Science"; date: "2015-10-9-
            9"; volume: "350"; issue: "6257"; pages: "211-7"
COMMENT     Sequence Label: pLJM60 FLAG-RAB35 wt
FEATURES             Location/Qualifiers
     source          1..8003
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..227
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             228..408
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    455..580
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1077..1310
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1495..1539
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        1846..2049
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2000..2020
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2046..2070
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2280..2298
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2280..2297
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2311..2330
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2389..2412
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    3097..3214
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3263..3773
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3692..3710
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3795..4394
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3795..3814)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4291..4311
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4522..4755
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4827..4948
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4864..4883)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4918..4937
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4988..5123
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5143..5162)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5144..5162)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5172..5189)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5172..5188)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5181..5203)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5330..5785
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5417..5436)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5627..5648
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5811..5915
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5916..5984,5985..6776)
                     /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     complement(6134..6153)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6947..7535
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7436..7455
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7689..7706
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7823..7844
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7859..7864,7865..7882,7883..7889)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7897..7913
                     /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)."
     primer_bind     7902..7924
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7921..7937
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7921..7937
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7956..7976
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7958..7976
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 tgtagtctta tgcaatactc ttgtagtctt gcaacatggt aacgatgagt tagcaacatg
       61 ccttacaagg agagaaaaag caccgtgcat gccgattggt ggaagtaagg tggtacgatc
      121 gtgccttatt aggaaggcaa cagacgggtc tgacatggat tggacgaacc actgaattgc
      181 cgcattgcag agatattgta tttaagtgcc tagctcgata cataaacggg tctctctggt
      241 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
      301 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
      361 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
      421 cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc
      481 tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac
      541 tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat
      601 tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa
      661 acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga
      721 aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc
      781 agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat
      841 agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa
      901 gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg agatatgagg
      961 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     1021 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     1081 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     1141 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     1201 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     1261 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     1321 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
     1381 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
     1441 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
     1501 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
     1561 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
     1621 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
     1681 tttcagaccc acctcccaac cccgagggga ccccatatgc caagtacgcc ccctattgac
     1741 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     1801 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
     1861 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     1921 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     1981 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     2041 agcagagctc gtttagtgaa ccgtcagatc gcctggagac gccatccacg ctgttttgac
     2101 ctccatagaa gacaccggga ccgatccagc ctccgcggcc gggaacggtg cattggaacg
     2161 cggattcccc gtgccaagag tgacgtaagt accgcctata gagtctatag gcccaccccc
     2221 ttggcttcgt tagaacgcgg ctacaattaa tacataacct tatgtatcat acacatacga
     2281 tttaggtgac actatagaat aacatccact ttgcctttct ctccacaggt gtccactccc
     2341 aggtccaact gcacctcggt tctatcgatt gaattcacca tggaaatgga ctacaaggat
     2401 gacgatgaca agggtggtgg tggtggtgcg tcgacgatgg cccgggacta cgaccacctc
     2461 ttcaagctgc tcatcatcgg cgacagcggt gtgggcaaga gcagtttact gttgcgtttt
     2521 gcagacaaca ctttctcagg cagctacatc accacgatcg gagtggattt caagatccgg
     2581 accgtggaga tcaacgggga gaaggtgaag ctgcagatct gggacacagc ggggcaggag
     2641 cgcttccgca ccatcacctc cacgtattat cgggggaccc acggggtcat tgtggtttac
     2701 gacgtcacca gtgccgagtc ctttgtcaac gtcaagcggt ggcttcacga aatcaaccag
     2761 aactgtgatg atgtgtgccg aatattagtg ggtaataaga atgacgaccc tgagcggaag
     2821 gtggtggaga cggaagatgc ctacaaattc gccgggcaga tgggcatcca gttgttcgag
     2881 accagcgcca aggagaatgt caacgtggaa gagatgttca actgcatcac ggagctggtc
     2941 ctccgagcaa agaaagacaa cctggcaaaa cagcagcagc aacaacagaa cgatgtggtg
     3001 aagctcacga agaacagtaa acgaaagaaa cgctgctgct aagcggccgc agtgagaatt
     3061 ctcgacctcg agacaaatgg cagtattcat ccacaatttt aaaagaaaag gggggattgg
     3121 ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac aaactaaaga
     3181 attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg acagcagaga
     3241 tccactttgg ccgcggctcg agggggttgg ggttgcgcct tttccaaggc agccctgggt
     3301 ttgcgcaggg acgcggctgc tctgggcgtg gttccgggaa acgcagcggc gccgaccctg
     3361 ggtctcgcac attcttcacg tccgttcgca gcgtcacccg gatcttcgcc gctacccttg
     3421 tgggcccccc ggcgacgctt cctgctccgc ccctaagtcg ggaaggttcc ttgcggttcg
     3481 cggcgtgccg gacgtgacaa acggaagccg cacgtctcac tagtaccctc gcagacggac
     3541 agcgccaggg agcaatggca gcgcgccgac cgcgatgggc tgtggccaat agcggctgct
     3601 cagcagggcg cgccgagagc agcggccggg aaggggcggt gcgggaggcg gggtgtgggg
     3661 cggtagtgtg ggccctgttc ctgcccgcgc ggtgttccgc attctgcaag cctccggagc
     3721 gcacgtcggc agtcggctcc ctcgttgacc gaatcaccga cctctctccc cagggggatc
     3781 caccggagct taccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg
     3841 tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca
     3901 ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc
     3961 gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct
     4021 ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg
     4081 ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc
     4141 accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg
     4201 gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg
     4261 tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct
     4321 tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca
     4381 agcccggtgc ctgacgcccg ccccacgacc cgcagcgccc gaccgaaagg agcgcacgac
     4441 cccatgcatc ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact
     4501 ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga caagatctgc
     4561 tttttgcttg tactgggtct ctctggttag accagatctg agcctgggag ctctctggct
     4621 aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt
     4681 gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt
     4741 ggaaaatctc tagcagtagt agttcatgtc atcttattat tcagtattta taacttgcaa
     4801 agaaatgaat atcagagagt gagaggaact tgtttattgc agcttataat ggttacaaat
     4861 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     4921 gtttgtccaa actcatcaat gtatcttatc atgtctggct ctagctatcc cgcccctaac
     4981 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     5041 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     5101 gtgaggaggc ttttttggag gcctagggac gtacccaatt cgccctatag tgagtcgtat
     5161 tacgcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     5221 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     5281 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggga cgcgccctgt
     5341 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     5401 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     5461 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     5521 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     5581 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     5641 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     5701 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     5761 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
     5821 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     5881 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5941 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     6001 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     6061 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     6121 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     6181 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     6241 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6301 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6361 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6421 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     6481 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6541 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6601 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6661 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6721 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     6781 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     6841 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     6901 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     6961 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     7021 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     7081 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     7141 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     7201 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     7261 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     7321 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     7381 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     7441 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     7501 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     7561 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     7621 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     7681 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     7741 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     7801 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
     7861 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
     7921 caggaaacag ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac
     7981 aaaagctgga gctgcaagct taa
//
