LOCUS       sequence-139749-        7091 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 7091)
  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 7091)
  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 WDR24 pRK5
FEATURES             Location/Qualifiers
     source          1..7091
                     /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      3378..3387
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3391..3525
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3428..3447)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3482..3501
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        3594..3951
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3594..3785
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3617..3637)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3802..3937
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3971..3988)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3971..3987)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3980..4002)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4200..4655
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4287..4306)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4497..4518
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4672..4691)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4791..4813
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4851..4869)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4937..5041
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5042..5110,5111..5902)
                     /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(5260..5279)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6073..6661
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6562..6581
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6815..6832
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6949..6970
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6985..6990,6991..7008,7009..7015)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7023..7039
                     /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     7028..7050
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7047..7063
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7047..7063
                     /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 gatggagaag atgtcccgtg tgaccacagc cctgggtggc agcgtgctga
     1021 caggccgcac catgcactgc cacctggatg ctcccgccaa tgccatcagt gtgtgccgcg
     1081 acgcagccca ggtggtcgtg gcaggccgta gcatcttcaa gatctatgcc atcgaggagg
     1141 aacagttcgt ggaaaagctg aacctgcgtg tggggcgcaa gccttcgctt aacctgagct
     1201 gtgctgacgt ggtctggcac cagatggatg agaacctgct ggccacagca gccaccaatg
     1261 gcgtggtggt cacgtggaac ctgggccggc catcccgcaa caagcaggac cagctgttca
     1321 cagaacacaa gcgcacggta aacaaagtct gcttccaccc caccgaagcc cacgtgctgc
     1381 tcagtggctc ccaggatggc ttcatgaagt gctttgacct ccgcagaaag gactctgtca
     1441 gcaccttctc gggccagtcg gagagcgtgc gggacgtgca gttcagtatc cgggactact
     1501 tcaccttcgc ctccaccttt gagaacggca atgtgcagct ctgggacatc cggcgtcccg
     1561 accggtgcga gaggatgttc acagcccaca acggacccgt cttctgctgc gactggcacc
     1621 ccgaggacag gggctggttg gccactggag ggcgcgacaa gatggtgaag gtctgggaca
     1681 tgaccacgca ccgtgccaag gagatgcact gtgtgcagac catcgcctcg gtggcccgtg
     1741 tgaagtggcg gccagagtgc cgccaccacc tggccacgtg ctccatgatg gtggaccaca
     1801 acatctatgt ttgggacgtg cgccggccct tcgtgccagc tgccatgttt gaggaacacc
     1861 gagacgtcac cacgggaatt gcctggcgcc acccccacga cccctccttc ctgctgtctg
     1921 gctccaagga cagctcgctg tgccagcacc tgttccgcga cgccagccag cccgtcgagc
     1981 gcgccaaccc tgagggcctc tgctacggcc tcttcgggga cctggccttc gccgccaagg
     2041 agagcctcgt ggctgccgag tcggggcgca agccctacac tggcgaccgg cgccacccca
     2101 tcttctttaa gcgcaagctg gaccctgccg agcccttcgc aggcctcgcc tccagtgccc
     2161 tcagtgtctt tgagacggag ccaggtggcg gcggcatgcg ctggtttgtg gacacagctg
     2221 agcgttatgc gctggctggc cggccactgg ccgagctctg tgaccacaac gcaaaggtgg
     2281 ctcgagagct tggccgcaac caggtggcgc aaacgtggac catgctgcgg atcatctact
     2341 gcagccctgg cctagtgccc actgcaaacc tcaaccacag tgtgggcaag ggtggctcct
     2401 gtggcctccc gctcatgaac agtttcaacc tgaaggatat ggccccaggg ttgggcagtg
     2461 agacgcggct ggaccgcagc aaaggagatg cgcggagcga cacagttctg ctcgactcct
     2521 cggccacact catcaccaat gaggataacg aggaaaccga gggcagcgac gtacctgccg
     2581 actacctgct gggtgacgtg gaaggtgagg aggacgagct gtacctgctg gatccggaac
     2641 acgcgcaccc cgaggaccct gagtgcgtgc tgccgcagga ggcctttccg ctgcgccacg
     2701 agatcgtgga cacgcctccc gggcccgagc acctgcagga caaggccgac tccccgcacg
     2761 tgagcggcag cgaggcggat gtggcctccc tggcccccgt ggactcctcc ttctcgctcc
     2821 tgtctgtctc acacgcgctc tacgacagcc gcctgccgcc cgacttcttc ggcgtgctgg
     2881 tgcgcgacat gctgcacttc tacgctgagc agggcgacgt gcagatggct gtgtctgtgc
     2941 tcatcgtcct gggtgaacgg gtgcgcaagg acatcgacga gcagacccag gagcactggt
     3001 acacttccta catcgacctg ctgcagcgct tccgcctctg gaacgtgtcc aacgaggtgg
     3061 tcaagctgag caccagccgc gccgtcagct gcctcaacca ggcctccacc accctgcacg
     3121 tcaactgcag ccactgcaag cggcccatga gcagccgggg ctgggtctgc gacaggtgcc
     3181 accgctgcgc cagcatgtgt gccgtctgcc accacgtagt caagggtctc ttcgtgtggt
     3241 gccagggctg cagccacggc ggccacctgc agcacatcat gaagtggctg gaaggcagct
     3301 cccactgtcc cgcaggctgc ggccacctct gcgagtactc ctgagcggcc gctaagtaag
     3361 taaggatccc cagcttggcc gccatggccc aacttgttta ttgcagctta taatggttac
     3421 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3481 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatcgggaa ttaattcggc
     3541 gcagcaccat ggcctgaaat aacctctgaa agaggaactt ggttaggtac cttctgaggc
     3601 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3661 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3721 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     3781 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     3841 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     3901 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctgtta
     3961 acagcttggc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     4021 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     4081 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt
     4141 attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta
     4201 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     4261 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     4321 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4381 tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc
     4441 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4501 actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt ttgatttata
     4561 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4621 cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg
     4681 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     4741 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     4801 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     4861 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     4921 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     4981 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     5041 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     5101 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     5161 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5221 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5281 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5341 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5401 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5461 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5521 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5581 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     5641 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     5701 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     5761 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     5821 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     5881 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     5941 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     6001 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     6061 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     6121 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6181 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     6241 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6301 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6361 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6421 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6481 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6541 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6601 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     6661 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     6721 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     6781 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     6841 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     6901 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
     6961 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     7021 aattgtgagc ggataacaat ttcacacagg aaacagctat gacatgatta cgaattaatt
     7081 cgagctcgcc c
//
