LOCUS       sequence-456576-        5954 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  overexpression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5954)
  AUTHORS   Wang S, Tsun ZY, Wolfson RL, Shen K, Wyant GA, Plovanich ME, Yuan 
            ED, Jones TD, Chantranupong L, Comb W, Wang T, Bar-Peled L, Zoncu R,
            Straub C, Kim C, Park J, Sabatini BL, Sabatini DM
  TITLE     Metabolism. Lysosomal amino acid transporter SLC38A9 signals 
            arginine sufficiency to mTORC1.
  JOURNAL   Science. 2015 Jan 9;347(6218):188-94. doi: 10.1126/science.1257132. 
            Epub 2015 Jan 7.
  PUBMED    25567906
REFERENCE   2  (bases 1 to 5954)
  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.1257132"; journalName: "Science"; date: "2015-01-9-
            9"; volume: "347"; issue: "6218"; pages: "188-94"
COMMENT     Sequence Label: pRK5-LAMP1-FLAG
FEATURES             Location/Qualifiers
     source          1..5954
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        7..386
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        387..590
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     541..561
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     587..611
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        821..839
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     821..838
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     852..871
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      925..934
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      929..938
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(931..1014,1015..2181)
                     /codon_start=1
                     /product="human lysosome-associated membrane glycoprotein 
                     1"
                     /label=LAMP1
                     /translation="MAAPGSARRPLLLLLLLLLLGLMHCASAAMFMVKNGNGTACIMAN
                     FSAAFSVNYDTKSGPKNMTFDLPSDATVVLNRSSCGKENTSDPSLVIAFGRGHTLTLNF
                     TRNATRYSVQLMSFVYNLSDTHLFPNASSKEIKTVESITDIRADIDKKYRCVSGTQVHM
                     NNVTVTLHDATIQAYLSNSSFSRGETRCEQDRPSPTTAPPAPPSPSPSPVPKSPSVDKY
                     NVSGTNGTCLLASMGLQLNLTYERKDNTTVTRLLNINPNKTSASGSCGAHLVTLELHSE
                     GTTVLLFQFGMNASSSRFFLQGIQLNTILPDARDPAFKAANGSLRALQATVGNSYKCNA
                     EEHVRVTKAFSVNIFKVWVQAFKVEGGQFGSVEECLLDENSMLIPIAVGGALAGLVLIV
                     LIAYLVGRKRSHAGYQTI"
     CDS             2203..2226
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      2247..2256
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2260..2394
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2297..2316)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2351..2370
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2463..2820
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2463..2654
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2486..2506)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2671..2806
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2840..2857)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2840..2856)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2849..2871)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3069..3524
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3156..3175)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3366..3387
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3541..3560)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3660..3682
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3720..3738)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3806..3910
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3911..3979,3980..4771)
                     /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(4129..4148)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4942..5530
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5431..5450
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5684..5701
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5818..5839
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5854..5859,5860..5877,5878..5884)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5892..5908
                     /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     5897..5919
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5916..5932
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5916..5932
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 tcgcccgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
       61 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      121 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      181 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      241 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      301 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      361 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      421 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      481 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      541 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga
      601 accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg
      661 accgatccag cctccgcggc cgggaacggt gcattggaac gcggattccc cgtgccaaga
      721 gtgacgtaag taccgcctat agagtctata ggcccacccc cttggcttcg ttagaacgcg
      781 gctacaatta atacataacc ttatgtatca tacacatacg atttaggtga cactatagaa
      841 taacatccac tttgcctttc tctccacagg tgtccactcc caggtccaac tgcacctcgg
      901 ttctatcgat tgaattcgtc gacggccacc atggcggccc ccggcagcgc ccggcgaccc
      961 ctgctgctgc tactgctgtt gctgctgctc ggcctcatgc attgtgcgtc agcagcaatg
     1021 tttatggtga aaaatggcaa cgggaccgcg tgcataatgg ccaacttctc tgctgccttc
     1081 tcagtgaact acgacaccaa gagtggccct aagaacatga cctttgacct gccatcagat
     1141 gccacagtgg tgctcaaccg cagctcctgt ggaaaagaga acacttctga ccccagtctc
     1201 gtgattgctt ttggaagagg acatacactc actctcaatt tcacgagaaa tgcaacacgt
     1261 tacagcgtcc agctcatgag ttttgtttat aacttgtcag acacacacct tttccccaat
     1321 gcgagctcca aagaaatcaa gactgtggaa tctataactg acatcagggc agatatagat
     1381 aaaaaataca gatgtgttag tggcacccag gtccacatga acaacgtgac cgtaacgctc
     1441 catgatgcca ccatccaggc gtacctttcc aacagcagct tcagcagggg agagacacgc
     1501 tgtgaacaag acaggccttc cccaaccaca gcgccccctg cgccacccag cccctcgccc
     1561 tcacccgtgc ccaagagccc ctctgtggac aagtacaacg tgagcggcac caacgggacc
     1621 tgcctgctgg ccagcatggg gctgcagctg aacctcacct atgagaggaa ggacaacacg
     1681 acggtgacaa ggcttctcaa catcaacccc aacaagacct cggccagcgg gagctgcggc
     1741 gcccacctgg tgactctgga gctgcacagc gagggcacca ccgtcctgct cttccagttc
     1801 gggatgaatg caagttctag ccggtttttc ctacaaggaa tccagttgaa tacaattctt
     1861 cctgacgcca gagaccctgc ctttaaagct gccaacggct ccctgcgagc gctgcaggcc
     1921 acagtcggca attcctacaa gtgcaacgcg gaggagcacg tccgtgtcac gaaggcgttt
     1981 tcagtcaata tattcaaagt gtgggtccag gctttcaagg tggaaggtgg ccagtttggc
     2041 tctgtggagg agtgtctgct ggacgagaac agcatgctga tccccatcgc tgtgggtggt
     2101 gccctggcgg ggctggtcct catcgtcctc atcgcctacc tcgtcggcag gaagaggagt
     2161 cacgcaggct accagactat cgcggccgca ggaggtggag gtgactacaa ggatgacgat
     2221 gacaagtgat aaggatcccc agcttggccg ccatggccca acttgtttat tgcagcttat
     2281 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     2341 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatcgggaat
     2401 taattcggcg cagcaccatg gcctgaaata acctctgaaa gaggaacttg gttaggtacc
     2461 ttctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     2521 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     2581 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     2641 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     2701 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     2761 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     2821 aagctgttaa cagcttggca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     2881 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     2941 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc
     3001 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca
     3061 accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     3121 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     3181 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     3241 ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg
     3301 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     3361 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg gctattcttt
     3421 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     3481 aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag
     3541 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     3601 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     3661 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     3721 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc
     3781 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     3841 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     3901 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     3961 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     4021 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     4081 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     4141 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     4201 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     4261 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     4321 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     4381 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     4441 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     4501 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     4561 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     4621 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     4681 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     4741 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     4801 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     4861 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     4921 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     4981 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5041 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5101 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5161 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5221 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     5281 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5341 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5401 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     5461 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     5521 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     5581 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     5641 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     5701 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     5761 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     5821 tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     5881 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg acatgattac
     5941 gaattaattc gagc
//
