LOCUS       sequence-194178-        5983 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HA tagged SAMTOR D190A mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5983)
  AUTHORS   Gu X, Orozco JM, Saxton RA, Condon KJ, Liu GY, Krawczyk PA, Scaria 
            SM, Harper JW, Gygi SP, Sabatini DM
  TITLE     SAMTOR is an S-adenosylmethionine sensor for the mTORC1 pathway.
  JOURNAL   Science. 2017 Nov 10;358(6364):813-818. doi: 
            10.1126/science.aao3265.
  PUBMED    29123071
REFERENCE   2  (bases 1 to 5983)
  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"; 
            journalName: "Science"; date: "2017-11-10- 10"; volume: "358"; 
            issue: "6364"; pages: "813-818"
COMMENT     Sequence Label: pRK5-HA-SAMTOR (D190A)
FEATURES             Location/Qualifiers
     source          1..5983
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      950..959
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             962..988
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2270..2279
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2283..2417
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2320..2339)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2374..2393
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2486..2843
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2486..2677
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2509..2529)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2694..2829
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2863..2880)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2863..2879)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2872..2894)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3092..3547
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3179..3198)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3389..3410
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3564..3583)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3683..3705
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3743..3761)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3829..3933
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3934..4002,4003..4794)
                     /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(4152..4171)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4965..5553
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5454..5473
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5707..5724
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5841..5862
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5877..5882,5883..5900,5901..5907)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5915..5931
                     /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     5920..5942
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5939..5955
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5939..5955
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgattgaatt cccgaaccga cagtcggtct cttcaccaag gccattcgcg ccaccatggg
      961 ctacccctat gatgtgcctg actacgcaga tctcaatggt ggtggtggtg ggtcgacgat
     1021 ggagccaggg gccggcggcc gaaatactgc tcgtgcgcag agggccgggt ccccgaacac
     1081 tccgccgccc cgggagcagg agcggaaact ggagcaggag aagctctccg gtgtggtgaa
     1141 gagcgtccac cggcggctcc gcaagaagta ccgagaagtg ggagattttg ataagatctg
     1201 gcgagaacac tgtgaggatg aagaaactct ttgtgaatat gccgttgcaa tgaaaaattt
     1261 ggcagataat cattgggcaa aaacttgtga gggcgaaggt cgtattgaat ggtgttgtag
     1321 tgtatgcaga gaatatttcc aaaatggtgg gaagagaaaa gcacttgaga aagatgaaaa
     1381 aagagctgta cttgccacta agaccactcc agccttaaat atgcatgagt cttctcaact
     1441 tgaaggtcat ttaaccaacc tcagctttac aaaccctgaa tttataactg agttgctaca
     1501 agcctcagga aaaatcagat tacttgatgt tggcagctgc tttaacccat ttctgaagtt
     1561 tgaagaattt ctaactgttg gcatagctat tgtacctgct gtagagagtg tctataaatg
     1621 tgatttcctg aacttacagc ttcagcaacc actccagctt gcacaggatg ctatagatgc
     1681 ttttttgaag cagctgaaaa accctattga ttctcttcct ggagagcttt tccatgtggt
     1741 tgttttctct ctccttcttt cttattttcc atctccttac cagcgatgga tttgctgcaa
     1801 gaaagcccat gaactgttag tgttaaatgg tttattacta atcatcacac ctgattcctc
     1861 ccatcagaac cgtcatgcta tgatgatgaa aagctggaag attgctatag agtccctggg
     1921 ctttaaacgc ttcaagtact caaaattttc acatatgcat ctgatggcat ttaggaaaat
     1981 ctctctaaaa accacaagtg acttggttag taggaactac ccaggaatgt tatatattcc
     2041 tcaagatttc aacagtatag aagatgagga atattctaac ccttcctgct atgttcgatc
     2101 agatatagaa gatgaacaac tagcatatgg tttcacagaa ctccctgatg cgccatatga
     2161 ctcagattct ggagaaagtc aagccagctc tattcctttc tatgagctag aagaccccat
     2221 attactttta agttaagcgg ccgctaagta agtaaggatc cccagcttgg ccgccatggc
     2281 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     2341 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     2401 cttatcatgt ctggatcggg aattaattcg gcgcagcacc atggcctgaa ataacctctg
     2461 aaagaggaac ttggttaggt accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc
     2521 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     2581 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2641 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     2701 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     2761 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     2821 ttggaggcct aggcttttgc aaaaagctgt taacagcttg gcactggccg tcgttttaca
     2881 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     2941 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     3001 cagcctgaat ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     3061 ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg
     3121 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     3181 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     3241 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     3301 cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     3361 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     3421 aaccctatct cgggctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     3481 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt
     3541 acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     3601 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     3661 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     3721 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     3781 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     3841 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     3901 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     3961 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     4021 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     4081 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     4141 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     4201 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     4261 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     4321 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     4381 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     4441 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     4501 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     4561 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     4621 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     4681 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     4741 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     4801 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     4861 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     4921 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     4981 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     5041 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     5101 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     5161 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     5221 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     5281 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     5341 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     5401 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     5461 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     5521 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     5581 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     5641 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     5701 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     5761 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg
     5821 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta
     5881 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca
     5941 ggaaacagct atgacatgat tacgaattaa ttcgagctcg ccc
//
