LOCUS       sequence-139740-        5861 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5861)
  AUTHORS   Bar-Peled L, Chantranupong L, Cherniack AD, Chen WW, Ottina KA, 
            Grabiner BC, Spear ED, Carter SL, Meyerson M, Sabatini DM
  TITLE     A Tumor suppressor complex with GAP activity for the Rag GTPases 
            that signal amino acid sufficiency to mTORC1.
  JOURNAL   Science. 2013 May 31;340(6136):1100-6. doi: 10.1126/science.1232044.
  PUBMED    23723238
REFERENCE   2  (bases 1 to 5861)
  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: "2013-05-31- 31"; volume: "340"; 
            issue: "6136"; pages: "1100-6"
COMMENT     Sequence Label: Flag Nprl2 pRK5
FEATURES             Location/Qualifiers
     source          1..5861
                     /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"
     CDS             924..947
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      2148..2157
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2161..2295
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2198..2217)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2252..2271
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2364..2721
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2364..2555
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2387..2407)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2572..2707
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2741..2758)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2741..2757)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2750..2772)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2970..3425
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3057..3076)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3267..3288
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3442..3461)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3561..3583
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3621..3639)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3707..3811
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3812..3880,3881..4672)
                     /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(4030..4049)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4843..5431
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5332..5351
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5585..5602
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5719..5740
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5755..5760,5761..5778,5779..5785)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5793..5809
                     /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     5798..5820
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5817..5833
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5817..5833
                     /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 caccatggaa atggactaca aggatgacga tgacaagggt ggtggtggtg
      961 gtgcgtcgac gatgggcagc ggctgccgca tcgaatgcat attcttcagc gagttccacc
     1021 ccacgctggg acccaagatc acctatcagg tccctgaaga cttcatctcc cgagagctgt
     1081 ttgacacagt ccaagtgtac atcatcacca agccagagct gcagaacaag cttatcactg
     1141 tcacagctat ggaaaagaag ctgatcggct gtcctgtgtg catcgaacac aagaagtaca
     1201 gccgcaatgc tctcctcttc aacctgggct tcgtgtgtga tgcccaggcc aagacctgcg
     1261 ccctcgagcc cattgttaaa aagctggctg gctatctgac cacactagag ctagagagca
     1321 gcttcgtgtc catggaggag agcaagcaga agttggtgcc catcatgacc atcttgctgg
     1381 aggagctaaa tgcctcaggc cggtgcactc tgcccattga tgagtccaac accatccact
     1441 tgaaggtgat tgagcagcgg ccagaccctc cggtggccca ggagtatgat gtacctgtct
     1501 ttaccaaaga caaggaggat ttcttcaact cacagtggga cctcactaca caacaaatcc
     1561 tgccctacat tgatgggttc cgccacatcc agaagatttc agcagaggca gatgtggagc
     1621 tcaacctggt gcgcattgct atccagaacc tgctgtacta cggcgttgtg acactggtgt
     1681 ccatcctcca gtactccaat gtatactgcc caacgcccaa ggtccaggac ctggtagatg
     1741 acaagtccct gcaagaggca tgtctatcct acgtgaccaa gcaagggcac aagagggcca
     1801 gtctccggga tgtgttccag ctatactgca gcctgagccc tggcactacc gtgcgagacc
     1861 tcattggccg ccacccccag cagctgcagc atgttgatga acggaagctg atccagttcg
     1921 ggcttatgaa gaacctcatc aggcgactac agaagtatcc tgtgcgggtg actcgggaag
     1981 agcagagcca ccctgcccgg ctttatacag gctgccacag ctatgacgag atctgctgca
     2041 agacaggcat gagctaccat gagctggatg agcggcttga aaatgacccc aacatcatca
     2101 tctgctggaa gtgagcggcc gctaagtaag taaggatccc cagcttggcc gccatggccc
     2161 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     2221 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     2281 tatcatgtct ggatcgggaa ttaattcggc gcagcaccat ggcctgaaat aacctctgaa
     2341 agaggaactt ggttaggtac cttctgaggc ggaaagaacc agctgtggaa tgtgtgtcag
     2401 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2461 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2521 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     2581 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     2641 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     2701 ggaggcctag gcttttgcaa aaagctgtta acagcttggc actggccgtc gttttacaac
     2761 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     2821 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     2881 gcctgaatgg cgaatggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     2941 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     3001 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     3061 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     3121 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     3181 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     3241 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     3301 ccctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     3361 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     3421 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     3481 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     3541 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     3601 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     3661 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3721 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3781 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     3841 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     3901 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3961 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     4021 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     4081 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     4141 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     4201 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     4261 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     4321 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     4381 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     4441 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     4501 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     4561 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     4621 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     4681 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4741 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4801 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4861 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4921 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4981 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     5041 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     5101 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     5161 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     5221 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     5281 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     5341 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     5401 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5461 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     5521 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     5581 gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc
     5641 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
     5701 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
     5761 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
     5821 aaacagctat gacatgatta cgaattaatt cgagctcgcc c
//
