LOCUS       sequence-359-exp        6993 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 6993)
  AUTHORS   Brunet A, Bonni A, Zigmond MJ, Lin MZ, Juo P, Hu LS, Anderson MJ, 
            Arden KC, Blenis J, Greenberg ME
  TITLE     Akt promotes cell survival by phosphorylating and inhibiting a 
            Forkhead transcription factor.
  JOURNAL   Cell 1999 Mar 19;96(6):857-68.
  PUBMED    10102273
REFERENCE   2  (bases 1 to 6993)
  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"; journalName: "Cell"; 
            date: "1999-03-19"; volume: "96(6)"; pages: "857-68"
COMMENT     Sequence Label: GST-FOXO3a WT
FEATURES             Location/Qualifiers
     source          1..6993
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2966..2982)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2966..2982)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3043..3065)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3210..3228)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3296..3400
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3401..3469,3470..4261)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3619..3638)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4432..5020
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4921..4940
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5174..5191
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5264..5341
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             5342..6424
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(5361..5380)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    6437..6458
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6473..6478,6479..6496,6497..6503)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6511..6527
                     /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     6535..6551
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6535..6551
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(6547..6993,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6567..6584)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6567..6583)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6576..6598)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatccccgaa ttcggcagag gcaccggctt
      961 ccccggcccc gctctctccg ctcgaagtgg agctggaccc ggagttcgag ccccagagcc
     1021 gtccgcgatc ctgtacgtgg cccctgcaaa ggccggagct ccaagcgagc cctgccaagc
     1081 cctcggggga gacggccgct gactccatga tccccgagga ggaggacgat gaagacgacg
     1141 aggacggcgg gggacgggcc ggctcggcca tggcgatcgg cggcggcggc gggagcggca
     1201 cgctgggctc cgggctgctc cttgaggact cggcccgggt gctggcaccc ggagggcaag
     1261 accccgggtc tgggccagcc accgcggcgg gcgggctgag cgggggtaca caggcgctgc
     1321 tgcagcctca gcaaccgctg ccaccgccgc agccgggggc ggctgggggc tccgggcagc
     1381 cgaggaaatg ttcgtcgcgg cggaacgcct ggggaaacct gtcctacgcg gacctgatca
     1441 cccgcgccat cgagagctcc ccggacaaac ggctcactct gtcccagatc tacgagtgga
     1501 tggtgcgttg cgtgccctac ttcaaggata agggcgacag caacagctct gccggctgga
     1561 agaactccat ccggcacaac ctgtcactgc atagtcgatt catgcgggtc cagaatgagg
     1621 gaactggcaa gagctcttgg tggatcatca accctgatgg ggggaagagc ggaaaagccc
     1681 cccggcggcg ggctgtctcc atggacaata gcaacaagta taccaagagc cgtggccgcg
     1741 cagccaagaa gaaggcagcc ctgcagacag cccccgaatc agctgacgac agtccctccc
     1801 agctctccaa gtggcctggc agccccacgt cacgcagcag tgatgagctg gatgcgtgga
     1861 cggacttccg ttcacgcacc aattctaacg ccagcacagt cagtggccgc ctgtcgccca
     1921 tcatggcaag cacagagttg gatgaagtcc aggacgatga tgcgcctctc tcgcccatgc
     1981 tctacagcag ctcagccagc ctgtcacctt cagtaagcaa gccgtgcacg gtggaactgc
     2041 cacggctgac tgatatggca ggcaccatga atctgaatga tgggctgact gaaaacctca
     2101 tggacgacct gctggataac atcacgctcc cgccatccca gccatcgccc actgggggac
     2161 tcatgcagcg gagctctagc ttcccgtata ccaccaaggg ctcgggcctg ggctccccaa
     2221 ccagctcctt taacagcacg gtgttcggac cttcatctct gaactcccta cgccagtctc
     2281 ccatgcagac catccaagag aacaagccag ctaccttctc ttccatgtca cactatggta
     2341 accagacact ccaggacctg ctcacttcgg actcacttag ccacagcgat gtcatgatga
     2401 cacagtcgga ccccttgatg tctcaggcca gcaccgctgt gtctgcccag aattcccgcc
     2461 ggaacgtgat gcttcgcaat gatccgatga tgtcctttgc tgcccagcct aaccagggaa
     2521 gtttggtcaa tcagaacttg ctccaccacc agcaccaaac ccagggcgct cttggtggca
     2581 gccgtgcctt gtcgaattct gtcagcaaca tgggcttgag tgagtccagc agccttgggt
     2641 cagccaaaca ccagcagcag tctcctgtca gccagtctat gcaaaccctc tcggactctc
     2701 tctcaggctc ctccttgtac tcaactagtg caaacctgcc cgtcatgggc catgagaagt
     2761 tccccagcga cttggacctg gacatgttca atgggagctt ggaatgtgac atggagtcca
     2821 ttatccgtag tgaactcatg gatgctgatg ggttggattt taactttgat tccctcatct
     2881 ccacacagaa tgttgttggt ttgaacgtgg ggaacttcac tggtgctaag caggcctcat
     2941 ctcagagctg ggtgccaggc tgaaggatcc actagttcta gagcggccgc atcgtgactg
     3001 actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     3061 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     3121 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
     3181 cggagtgtat aattcttgaa gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     3241 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     3301 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     3361 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     3421 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     3481 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     3541 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     3601 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg ccgggcaaga
     3661 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     3721 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     3781 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     3841 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     3901 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa tggcaacaac
     3961 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     4021 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4081 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     4141 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     4201 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     4261 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     4321 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     4381 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4441 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     4501 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     4561 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     4621 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     4681 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     4741 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     4801 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     4861 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     4921 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     4981 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5041 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5101 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     5161 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     5221 tctccttacg catctgtgcg gtatttcaca ccgcataaat tccgacacca tcgaatggtg
     5281 caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa
     5341 tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
     5401 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
     5461 ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca
     5521 gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt
     5581 cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga
     5641 acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag
     5701 tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg
     5761 cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat
     5821 tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca
     5881 gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg
     5941 ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg
     6001 gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac
     6061 tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc
     6121 cgggctgcgc gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc
     6181 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
     6241 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
     6301 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
     6361 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     6421 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     6481 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     6541 acagctatga ccatgattac ggattcactg gccgtcgttt tacaacgtcg tgactgggaa
     6601 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
     6661 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     6721 tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg aaagctggct ggagtgcgat
     6781 cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc agatgcacgg ttacgatgcg
     6841 cccatctaca ccaacgtaac ctatcccatt acggtcaatc cgccgtttgt tcccacggag
     6901 aatccgacgg gttgttactc gctcacattt aatgttgatg aaagctggct acaggaaggc
     6961 cagacgcgaa ttatttttga tggcgttgga att
//
