LOCUS       sequence-163546-        5228 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  overexpression of CASTOR1 ACT1 domain (aa 1-169).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5228)
  AUTHORS   Chantranupong L, Scaria SM, Saxton RA, Gygi MP, Shen K, Wyant GA, 
            Wang T, Harper JW, Gygi SP, Sabatini DM
  TITLE     The CASTOR Proteins Are Arginine Sensors for the mTORC1 Pathway.
  JOURNAL   Cell. 2016 Mar 24;165(1):153-64. doi: 10.1016/j.cell.2016.02.035. 
            Epub 2016 Mar 10.
  PUBMED    26972053
REFERENCE   2  (bases 1 to 5228)
  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: 
            "10.1016/j.cell.2016.02.035"; journalName: "Cell"; date: 
            "2016-03-24- 24"; volume: "165"; issue: "1"; pages: "153-64"
COMMENT     Sequence Label: pRK5-Flag CASTOR1 ACT1
FEATURES             Location/Qualifiers
     source          1..5228
                     /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"
     CDS             937..960
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      1528..1537
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1541..1675
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1578..1597)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1632..1651
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        1744..2101
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1744..1935
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1767..1787)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1952..2087
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2121..2138)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2121..2137)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2130..2152)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2350..2805
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2437..2456)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2647..2668
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2822..2841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2941..2963
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3001..3019)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3087..3191
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3192..3260,3261..4052)
                     /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(3410..3429)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4223..4811
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4712..4731
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4965..4982
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5099..5120
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5135..5140,5141..5158,5159..5165)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5173..5189
                     /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     5178..5200
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5197..5213
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5197..5213
                     /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 tatcgattga attcaccatg gaaatggact acaaggatga cgatgacaag
      961 ggtggtggtg gtggtgcgtc gacgatggag ctgcacatcc tagaacaccg ggtgcgggtg
     1021 ctgagcgtcg cccgtcccgg tctctggctc tacacccacc cgctcatcaa gctgctcttc
     1081 ctgccccgcc gcagccggtg caagttcttc agcctgacgg agacccctga ggattacacg
     1141 cttatggtgg acgaggaggg ctttaaagag ctgcccccat ctgagttcct gcaagtagct
     1201 gaggccacat ggctggtgct gaacgtgtcg tctcacagcg gtgcggcagt gcaggctgct
     1261 ggggtcacca agatcgcccg ttcggtcatc gcgccactgg ccgagcacca cgtgtctgtg
     1321 ctgatgctgt ccacttacca gacggacttc atcctggtgc gggagcagga cctgtccgtg
     1381 gtgatccaca cgctggccca ggagttcgac atttaccgcg aggtgggcgg agagcctgtg
     1441 cctgtgacga gggatgattc cagcaatggc tttccccgca ctcagcatgg gtgagcggcc
     1501 gctaagtaag taaggatccc cagcttggcc gccatggccc aacttgttta ttgcagctta
     1561 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     1621 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatcgggaa
     1681 ttaattcggc gcagcaccat ggcctgaaat aacctctgaa agaggaactt ggttaggtac
     1741 cttctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     1801 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     1861 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     1921 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     1981 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2041 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     2101 aaagctgtta acagcttggc actggccgtc gttttacaac gtcgtgactg ggaaaaccct
     2161 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc
     2221 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc
     2281 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc
     2341 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     2401 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     2461 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     2521 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac
     2581 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     2641 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt
     2701 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     2761 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca
     2821 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     2881 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     2941 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     3001 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     3061 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     3121 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     3181 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     3241 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     3301 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     3361 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     3421 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     3481 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     3541 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     3601 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     3661 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     3721 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     3781 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     3841 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     3901 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     3961 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     4021 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     4081 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     4141 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     4201 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4261 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     4321 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     4381 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     4441 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4501 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     4561 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     4621 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     4681 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     4741 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     4801 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     4861 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     4921 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     4981 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     5041 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     5101 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     5161 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gacatgatta
     5221 cgaattaa
//
