LOCUS       sequence-392069-        6201 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of LYCHOS (GPR155) ∆Permease.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6201)
  AUTHORS   Shin HR, Citron YR, Wang L, Tribouillard L, Goul CS, Stipp R, 
            Sugasawa Y, Jain A, Samson N, Lim CY, Davis OB, Castaneda-Carpio D, 
            Qian M, Nomura DK, Perera RM, Park E, Covey DF, Laplante M, Evers 
            AS, Zoncu R
  TITLE     Lysosomal GPCR-like protein LYCHOS signals cholesterol sufficiency 
            to mTORC1.
  JOURNAL   Science. 2022 Sep 16;377(6612):1290-1298. doi: 
            10.1126/science.abg6621. Epub 2022 Aug 25.
  PUBMED    36007018
REFERENCE   2  (bases 1 to 6201)
  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.abg6621"; journalName: "Science"; date: 
            "2022-09-16- 16"; volume: "377"; issue: "6612"; pages: "1290-1298"
COMMENT     Sequence Label: pRK5 LYCHOS (GPR155) ∆Permease-HA
FEATURES             Location/Qualifiers
     source          1..6201
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      918..927
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(1712..1723)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             2457..2483
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     regulatory      2501..2510
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2514..2648
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2551..2570)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2605..2624
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2717..3074
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2717..2908
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2740..2760)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2925..3060
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3094..3111)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3094..3110)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3103..3125)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3323..3778
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3410..3429)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3620..3641
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3795..3814)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3914..3936
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3974..3992)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4060..4164
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4165..4233,4234..5025)
                     /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(4383..4402)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5196..5784
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5685..5704
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5938..5955
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6072..6093
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6108..6113,6114..6131,6132..6138)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6146..6162
                     /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     6151..6173
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6170..6186
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6170..6186
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgatgcc accatggacc ctaagccatt ggcatatgcc atccagaatg
      961 ttagttttga tataagtatt gtcagcctga tctccttgat ctggtctctg gctattcttc
     1021 ttttgagtaa gaaatataaa cagcttcctc atatgcttac aactaattta ctcattgctc
     1081 agtctattgt ctgtgctgga atgatgatat ggaattttgt taaagaaaaa aattttgttg
     1141 gacaaatttt ggtgtttgtt ctattgtaca gctccctcta tagcacctac ctgtggacag
     1201 gccttctagc aatttctttg tttcttttga aaaagcgaga gagggtacaa attcctgttg
     1261 gaataatcat aatatctggc tggggaattc ctgctctcct tgttggtgtt cttttgataa
     1321 ctggaaaaca caatggagat agcattgact cagccttctt ttatggaaaa gaacagatga
     1381 tcaccacagc agtcaccctg ttctgcagca tcctgatagc tggcatatcc ctcatgtgca
     1441 tgaaccagac tgcccaagca ggaagctatg aaggtttcga tcagtctcag agccacaaag
     1501 tggtggagcc tggaaatact gcttttgagg agagtccagc accagtaaat gaaccagaac
     1561 tttttacaag ctctattcca gaaacaagtt gctgctcctg ctccatggga aatggtgaat
     1621 tacactgccc atcaatagag ccaatagcaa acacaagcac cagtgagcct gtgattcctt
     1681 cgtttgagaa aaacaatcat tgtgtgagtc gctgtaactc ccagagctgc atattagccc
     1741 aggaagaaga acagtatcta cagagtggag accagcaact gacccgacat gtgttgctgt
     1801 gtttacttct catcattggc ctgttcgcta atctttccag ttgtttatgg tggctattca
     1861 accaagagcc tggaagactt tatgttgagt tacagttttt ctgtgccgtg tttaactttg
     1921 gtcagggatt tatttccttt ggaatctttg gattagataa acatttaatc atcctgcctt
     1981 tcaaaagaag acttgaattc ctatggaaca ataaagacac agcagaaaac agggattctc
     2041 ctgtttcaga ggaaataaaa atgacctgtc aacaatttat ccattatcac cgtgacctct
     2101 gtatccgaaa cattgtcaaa gaaagaaggt gtggtgcaaa gacttctgct ggaactttct
     2161 gtggctgtga cctggtgagc tggctaattg aagtcggcct tgcctccgac cgtggtgaag
     2221 ctgtgatata cggagacagg ctggtacaag ggggagtcat ccaacatatt accaatgagt
     2281 atgaattccg ggatgagtac ttgttttaca gatttcttca aaagagtcct gaacagagtc
     2341 ctcctgctat taatgcaaac actctccaac aggaaagata taaagaaatt gagcattcat
     2401 ccccaccctc acattcccct aagaccagat ctaatggtgg tggtggtggg atgggctacc
     2461 cctatgatgt gcctgactac gcataaggat ccccagcttg gccgccatgg cccaacttgt
     2521 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     2581 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     2641 tctggatcgg gaattaattc ggcgcagcac catggcctga aataacctct gaaagaggaa
     2701 cttggttagg taccttctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt
     2761 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     2821 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     2881 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     2941 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     3001 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     3061 taggcttttg caaaaagctg ttaacagctt ggcactggcc gtcgttttac aacgtcgtga
     3121 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     3181 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     3241 tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3301 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     3361 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     3421 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     3481 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     3541 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     3601 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     3661 tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     3721 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     3781 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     3841 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     3901 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     3961 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     4021 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4081 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     4141 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     4201 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     4261 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     4321 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4381 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     4441 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4501 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     4561 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     4621 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     4681 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     4741 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     4801 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     4861 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     4921 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     4981 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5041 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5101 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     5161 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     5221 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5281 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     5341 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5401 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5461 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     5521 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     5581 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     5641 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     5701 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     5761 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     5821 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5881 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     5941 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     6001 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     6061 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     6121 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     6181 tatgacatga ttacgaatta a
//
