LOCUS       sequence-139737-        9464 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 9464)
  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 9464)
  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 Depdc5 pRK5
FEATURES             Location/Qualifiers
     source          1..9464
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        250..629
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        630..833
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     784..804
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     830..854
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        1064..1082
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1064..1081
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1095..1114
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             1173..1196
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      6000..6009
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    6013..6147
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6050..6069)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6104..6123
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        6216..6573
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6216..6407
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6239..6259)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6424..6559
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6593..6610)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6593..6609)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6602..6624)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6822..7277
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6909..6928)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7119..7140
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7294..7313)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7413..7435
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7473..7491)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7559..7663
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7664..7732,7733..8524)
                     /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(7882..7901)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8695..9283
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9184..9203
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9437..9454
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga catgattacg aattaattcg
      241 agctcgcccg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
      301 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      361 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      421 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      481 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      541 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      601 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      661 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      721 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      781 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
      841 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc
      901 gggaccgatc cagcctccgc ggccgggaac ggtgcattgg aacgcggatt ccccgtgcca
      961 agagtgacgt aagtaccgcc tatagagtct ataggcccac ccccttggct tcgttagaac
     1021 gcggctacaa ttaatacata accttatgta tcatacacat acgatttagg tgacactata
     1081 gaataacatc cactttgcct ttctctccac aggtgtccac tcccaggtcc aactgcacct
     1141 cggttctatc gattgaattc accatggaaa tggactacaa ggatgacgat gacaagggtg
     1201 gtggtggtgg tgcgtcgacg atgagaacaa caaaggtcta caaactcgtc atccacaaga
     1261 agggctttgg gggcagtgat gatgagctag ttgtgaaccc caaagtgttc cctcacatca
     1321 agcttggaga cattgtagag attgcacacc ccaacgatga atacagccct ctgcttttgc
     1381 aggtcaagtc tcttaaggaa gatttacaga aggaaactat cagtgtggac cagactgtga
     1441 ctcaagtgtt ccggctgaga ccttatcagg atgtctatgt taatgtcgta gaccctaagg
     1501 atgtgaccct tgacctagtg gaattaactt ttaaggatca gtatattggc cgtggggata
     1561 tgtggcgact aaagaaaagt ttggtcagca catgtgccta tatcacccag aaggtggagt
     1621 ttgctggcat cagagcacag gctggtgaac tgtgggttaa gaatgagaag gtcatgtgtg
     1681 gctacatcag tgaagatacc agggtggtgt ttcgttctac gtcggctatg gtttacatat
     1741 ttattcagat gagctgtgaa atgtgggatt ttgatattta tggggatttg tattttgaga
     1801 aagctgtgaa tggtttcctt gctgatctat ttaccaagtg gaaggagaag aactgtagtc
     1861 atgaagtgac agtggtcctg ttttctagaa ctttctatga tgcaaaatct gttgatgaat
     1921 ttcctgaaat aaaccgagcc tcaattcgac aggatcacaa ggggagattc tatgaagact
     1981 tttacaaagt ggtggtgcag aatgagagaa gagaagaatg gacttcactt ctcgtaacca
     2041 ttaaaaaact cttcatccag tatccagtgt tggtgcgact ggaacaggca gagggctttc
     2101 ctcaaggaga taattctacc tcagcacaag gaaactacct ggaggccatc aatctgtcat
     2161 tcaatgtgtt tgataagcac tacatcaacc gcaactttga ccgaactggg cagatgtcag
     2221 tggtgatcac gcccggggtg ggtgtctttg aagtggaccg cctactcatg atcctgacca
     2281 agcagcggat gatagataat ggaattggtg tggatttggt gtgcatggga gagcaaccgt
     2341 tacatgctgt cccattgttc aagctccata atcggagtgc tccccgtgat tctcgtctgg
     2401 gcgatgacta taatatccct cactggataa accacagttt ctacacatcc aaaagccagc
     2461 tcttttgtaa tagtttcacc ccacgaataa aactggcagg aaagaagccc gcctctgaga
     2521 aagcaaaaaa tggccgtgat acatctctcg ggagtccaaa agaatctgag aacgcccttc
     2581 ccatccaagt agattatgac gcctatgacg ctcaagtgtt caggctgccc ggcccatccc
     2641 gggcccagtg cctcaccacc tgcagatctg tgcgagagcg agagagtcac agtcgaaaga
     2701 gtgccagctc ctgtgatgtt tcatccagcc cttccctacc aagccgcaca ctgcccactg
     2761 aggaagtgag gagccaggct tctgacgaca gctccctagg caagagtgcc aacatcctga
     2821 tgatcccaca cccccacctg caccagtatg aagtcagcag ctccttggga tacaccagca
     2881 ctcgagatgt cctggagaac atgatggagc caccacagcg agactccagt gcaccaggga
     2941 ggtttcacgt tggcagtgca gaatccatgc tgcatgttcg acctggtgga tacacgcccc
     3001 agagagcact gattaacccc ttcgctccct ctcggatgcc catgaagctt acgtccaaca
     3061 gaaggcgctg gatgcacact tttcctgtgg ggccatccgg agaagccatc cagatccacc
     3121 accagacccg acagaatatg gcggagctac aaggcagcgg gcagagggat ccaactcact
     3181 cctctgcaga gctgctggag ttagcatatc atgaagctgc tggaaggcac agcaattccc
     3241 gccagcctgg tgacggcatg tccttcttga acttcagtgg aacagaggag ctttctgtcg
     3301 gcctgcttag caacagtggt gcaggtatga atcctaggac ccagaataag gattctctag
     3361 aggacagtgt ttctacctct ccagacccaa ttctgacact gtctgctccc cctgtagtgc
     3421 caggcttctg ttgcacagtt ggagtggact ggaagtctct cactactccg gcgtgcctcc
     3481 cccttaccac cgactacttc cctgaccgcc agggcctgca gaatgactac acagagggct
     3541 gttatgatct ccttccagaa gcagacatcg acaggaggga cgaagatggt gtgcagatga
     3601 cagcccagca ggtatttgaa gagtttattt gccaacgtct catgcagggc taccaaatca
     3661 tagtgcagcc caagacacag aaacccaatc ctgctgtccc gcccccgctg agcagtagcc
     3721 cactctatag ccgaggcctt gtgtcccgaa accgccctga ggaggaggac cagtattggc
     3781 tgagtatggg cagaacgttc cacaaagtga cgctgaagga taagatgatc acagtgacgc
     3841 gataccttcc caagtatcct tatgaatctg cccagatcca ctacacctac agcctctgtc
     3901 cttcccactc agactcagag ttcgtctcct gctgggtgga attctcccac gaacggctgg
     3961 aggagtacaa gtggaattac ttagatcagt atatctgttc tgccggctct gaagacttca
     4021 gcttaattga gtccctgaag ttctggagga cccgcttcct gctgctgcca gcctgtgtca
     4081 ccgccaccaa gcgcatcacg gagggggagg cccactgcga catctatggg gacaggcccc
     4141 gtgcagacga ggacgagtgg caactcctgg atggttttgt ccgctttgtg gagggcttga
     4201 atcgcattcg caggcggcat cgctcggatc gcatgatgcg gaaagggacc gccatgaaag
     4261 gcttgcagat gactgggccc atttccacgc attctctgga gtcaactgca cccccagtgg
     4321 ggaagaaggg aacctcagct ctctctgccc tgttggagat ggaggccagt cagaagtgcc
     4381 tgggagaaca gcaggcagct gtgcatggtg ggaagagctc cgcccagtca gccgagagca
     4441 gcagcgttgc catgactccc acctacatgg acagcccacg aaaggtatct gtggaccaaa
     4501 cagccactcc tatgttggac ggcaccagtt tgggcatatg cacaggccaa tccatggaca
     4561 gaggcaacag ccagaccttt gggaactccc agaacatagg agaacagggc tactcctcca
     4621 caaactccag tgacagcagc tctcagcagc tggtggcaag ctccttgacc tcatcctcta
     4681 ccctgacaga gatcctggaa gccatgaagc acccctcgac aggagtccag ctgctctctg
     4741 aacagaaggg cctctcaccg tactgcttca tcagcgcgga ggtggtacac tggttggtga
     4801 accacgtgga ggggatccag acacaggcga tggccattga catcatgcag aaaatgctgg
     4861 aagagcagct catcacacat gcatctggcg aagcctggcg gaccttcatc tacggcttct
     4921 atttctacaa gatagtaacg gacaaagagc ccgaccgagt ggccatgcag cagcccgcca
     4981 ccacctggca cacagcagga gtggacgact tcgccagctt ccagcgcaag tggtttgagg
     5041 tggcctttgt ggcagaagag ctcgtgcact ctgagattcc tgcctttctc ctgccctggc
     5101 tgcctagccg gccagcctcc tatgcaagta ggcacagctc ctttagccga agttttggag
     5161 gacggagcca ggcggcagca cttttagctg ccactgtccc agagcagagg actgtgaccc
     5221 tggatgttga cgtgaacaac cgcacagacc ggctggagtg gtgcagctgt tattaccatg
     5281 gcaacttttc tctgaatgca gcctttgaga tcaagctgca ctggatggcg gtgaccgcag
     5341 cagtactctt cgagatggtc caaggttggc atcggaaagc cacctcctgt ggcttcttgt
     5401 tagtcccagt tttggagggg ccttttgcac tgcccagtta cctgtatggc gacccccttc
     5461 gtgcccagct cttcatccca ctcaacatca gctgcttgct caaggagggc agcgagcacc
     5521 tgtttgatag ctttgaaccc gaaacgtact gggatcgaat gcacctcttc caggaagcca
     5581 ttgcacacag gtttgggttt gtacaagata aatattctgc ctctgctttt aacttccctg
     5641 ctgagaacaa gcctcagtat atccacgtta caggaacagt gtttctgcag ctgccctact
     5701 ccaagcgcaa gttctcaggg cagcagcggc ggcggcggaa ctccaccagc tccaccaacc
     5761 agaacatgtt ctgcgaggag cgggtcggct acaactgggc ctacaacacc atgctcacca
     5821 aaacatggcg ctccagcgcc acaggggatg aaaagtttgc tgatcggctg ctgaaggact
     5881 tcacggactt ctgcatcaac cgtgacaacc ggctggtcac gttctggaca agttgcctgg
     5941 agaagatgca tgccagtgcc ccgtgagcgg ccgctaagta agtaaggatc cccagcttgg
     6001 ccgccatggc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     6061 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     6121 atcaatgtat cttatcatgt ctggatcggg aattaattcg gcgcagcacc atggcctgaa
     6181 ataacctctg aaagaggaac ttggttaggt accttctgag gcggaaagaa ccagctgtgg
     6241 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     6301 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     6361 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     6421 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     6481 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     6541 ggaggctttt ttggaggcct aggcttttgc aaaaagctgt taacagcttg gcactggccg
     6601 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
     6661 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     6721 aacagttgcg cagcctgaat ggcgaatggc gcctgatgcg gtattttctc cttacgcatc
     6781 tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc
     6841 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     6901 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     6961 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     7021 ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     7081 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     7141 aacaacactc aaccctatct cgggctattc ttttgattta taagggattt tgccgatttc
     7201 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     7261 attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     7321 aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     7381 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     7441 accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt
     7501 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     7561 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     7621 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     7681 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     7741 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     7801 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     7861 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     7921 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     7981 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     8041 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     8101 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     8161 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     8221 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     8281 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     8341 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     8401 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     8461 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     8521 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     8581 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     8641 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     8701 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     8761 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     8821 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     8881 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     8941 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     9001 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     9061 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     9121 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     9181 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     9241 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     9301 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     9361 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     9421 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaag
//
