LOCUS       sequence-490911-        7516 bp    DNA     circular SYN 28-OCT-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7516)
  AUTHORS   Samuels Y, Wang Z, Bardelli A, Silliman N, Ptak J, Szabo S, Yan H, 
            Gazdar A, Powell SM, Riggins GJ, Willson JK, Markowitz S, Kinzler 
            KW, Vogelstein B, Velculescu VE
  TITLE     High frequency of mutations of the PIK3CA gene in human cancers.
  JOURNAL   Science. 2004 Apr 23. 304(5670):554.
  PUBMED    15016963
REFERENCE   2  (bases 1 to 7516)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 28, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science. 
            2004 Apr 23. 304(5670):554."
COMMENT     Sequence Label: PIK3CA-WT
FEATURES             Location/Qualifiers
     source          1..7516
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        621..639
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      674..683
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             683..706
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4018..4037)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4019..4037)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    4311..4432
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4348..4367)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4402..4421
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4439..4894)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4576..4597)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4788..4807
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4921..5025
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5027..5384
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5027..5218
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5050..5070)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5235..5370
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5419..6213
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5419..6213
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5473..5492)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6083..6102
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6401..6420)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6445..6492
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6821..7409
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7310..7329
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgattacg ccaagctcga aattaaccct cactaaaggg aacaaaagct ggagctccac
      661 cgcggtggcg gccgccacca tggattacaa ggatgacgac gataaggccc gggcggatcc
      721 cccgggctgc aggaattcga tagttcgaac catgcctcca agaccatcat caggtgaact
      781 gtggggcatc cacttgatgc ccccaagaat cctagtagaa tgtttactac caaatggaat
      841 gatagtgact ttagaatgcc tccgcgaggc tacattaata accataaagc atgaactatt
      901 taaagaagca agaaaatacc ccctccatca acttcttcaa gatgaatctt cttacatttt
      961 cgtaagtgtt actcaagaag cagaaaggga agaatttttt gatgaaacaa gacgactttg
     1021 tgaccttcgg ctttttcaac cctttttaaa agtaattgaa ccagtaggca accgtgaaga
     1081 aaagatcctc gatcgagaaa ttggttttgc tatcggcatg ccagtgtgtg aatttgatat
     1141 ggttaaagat ccagaagtac aggacttccg aagaaatatt ctgaacgttt gtaaagaagc
     1201 tgtggatctt agggacctca attcacctca tagtagagca atgtatgtct atcctccaaa
     1261 tgtagaatct tcaccagaat cgccaaagca catatacaat aaattagata aagggcaaat
     1321 aatagtggtg atctgggtaa tagtttctcc aaataatgac aagcagaagt atactctgaa
     1381 aatcaaccat gactgtgtac cagaacaagt aattgctgaa gcaatcagga aaaaaactcg
     1441 aagtatgttg ctatcctctg aacaactaaa actctgtgtt ttagaatatc agggcaagta
     1501 tattttaaaa gtgtgtggat gtgatgaata cttcctagaa aaatatcctc tgagtcagta
     1561 taagtatata agaagctgta taatgctagg gaggatgccc aatttgatgt tgatggctaa
     1621 agaaagcctt tattctcaac tgccgatgga ctgtttcaca atgccatctt attccagacg
     1681 catttccaca gctacaccat atatgaatgg agaagcatct acaaaatccc tttgggttat
     1741 aaatagtgca ctcagaataa aaattctttg tgcaacctac gtgaatgtaa atattcgaga
     1801 cattgataag atctatgttc gaacaggtat ctaccatgga ggagaaccct tatgtgacaa
     1861 tgtgaacact caaagagtac cttgttccaa tcccaggtgg aatgaatggc tgaattatga
     1921 tatatacatt cctgatcttc ctcgtgctgt tcgactttgc ccttccattt gctctgttaa
     1981 aggccgaaag ggtgctaaag aggaacactg tccattggca tggggaaata taaacttgtt
     2041 tgattacaca gacactctag tatctggaaa aatggctttg aatctttggc cagtacctca
     2101 tggattagaa gatttgctga accctattgg tgttactgga tcaaatccaa ataaagaaac
     2161 tccatgctta gagttggagt ttgactggtt cagcagtgtg gtaaagttcc cagatatgtc
     2221 agtgattgaa gagcatgcca attggtctgt atcccgagaa gcaggattta gctattccca
     2281 cgcaggactg agtaacagac tagctagaga caatgaatta agggaaaatg acaaagaaca
     2341 gctcaaagca atttctacac gagatcctct ctctgaaatc actgagcagg agaaagattt
     2401 tctatggagt cacagacact attgtgtaac tatccccgaa attctaccca aattgcttct
     2461 gtctgttaaa tggaattcta gagatgaagt agcccagatg tattgcttgg taaaagattg
     2521 gcctccaatc aaacctgaac aggctatgga acttctggac tgtaattacc cagatcctat
     2581 ggttcgaggt tttgctgttc ggtgcttgga aaaatattta acagatgaca aactttctca
     2641 gtatttaatt cagctagtac aggtcctaaa atatgaacaa tatttggata acttgcttgt
     2701 gagattttta ctgaagaaag cattgactaa tcaaaggatt gggcactttt tcttttggca
     2761 tttaaaatct gagatgcaca ataaaacagt tagccagagg tttggcctgc ttttggagtc
     2821 ctattgtcgt gcatgtggga tgtatttgaa gcacctgaat aggcaagtcg aggcaatgga
     2881 aaagctcatt aacttaactg acattctcaa acaggagaag aaggatgaaa cacaaaaggt
     2941 acagatgaag tttttagttg agcaaatgag gcgaccagat ttcatggatg ctctacaggg
     3001 ctttctgtct cctctaaacc ctgctcatca actaggaaac ctcaggcttg aagagtgtcg
     3061 aattatgtcc tctgcaaaaa ggccactgtg gttgaattgg gagaacccag acatcatgtc
     3121 agagttactg tttcagaaca atgagatcat ctttaaaaat ggggatgatt tacggcaaga
     3181 tatgctaaca cttcaaatta ttcgtattat ggaaaatatc tggcaaaatc aaggtcttga
     3241 tcttcgaatg ttaccttatg gttgtctgtc aatcggtgac tgtgtgggac ttattgaggt
     3301 ggtgcgaaat tctcacacta ttatgcaaat tcagtgcaaa ggcggcttga aaggtgcact
     3361 gcagttcaac agccacacac tacatcagtg gctcaaagac aagaacaaag gagaaatata
     3421 tgatgcagcc attgacctgt ttacacgttc atgtgctgga tactgtgtag ctaccttcat
     3481 tttgggaatt ggagatcgtc acaatagtaa catcatggtg aaagacgatg gacaactgtt
     3541 tcatatagat tttggacact ttttggatca caagaagaaa aaatttggtt ataaacgaga
     3601 acgtgtgcca tttgttttga cacaggattt cttaatagtg attagtaaag gagcccaaga
     3661 atgcacaaag acaagagaat ttgagaggtt tcaggagatg tgttacaagg cttatctagc
     3721 tattcgacag catgccaatc tcttcataaa tcttttctca atgatgcttg gctctggaat
     3781 gccagaacta caatcttttg atgacattgc atacattcga aagaccctag ccttagataa
     3841 aactgagcaa gaggctttgg agtatttcat gaaacaaatg aatgatgcac atcatggtgg
     3901 ctggacaaca aaaatggatt ggatcttcca cacaattaaa cagcatgcat tgaactagct
     3961 cgaggggggg cccggtacct taattaatta aggtaccagg taagtgtacc caattcgccc
     4021 tatagtgagt cgtattacaa ttcactcgat cgcccttccc aacagttgcg cagcctgaat
     4081 ggcgaatgga gatccaattt ttaagtgtat aatgtgttaa actactgatt ctaattgttt
     4141 gtgtatttta gattcacagt cccaaggctc atttcaggcc cctcagtcct cacagtctgt
     4201 tcatgatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     4261 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     4321 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     4381 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct taacgcgtaa
     4441 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     4501 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     4561 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     4621 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     4681 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     4741 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     4801 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     4861 cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg
     4921 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     4981 caataaccct gataaatgct tcaataatat tgaaaaagga agaatcctga ggcggaaaga
     5041 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     5101 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     5161 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     5221 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     5281 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     5341 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
     5401 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     5461 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     5521 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     5581 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
     5641 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     5701 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     5761 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     5821 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     5881 caggatgatc tggacgaaga acatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     5941 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     6001 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     6061 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga acttggcggc
     6121 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     6181 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     6241 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     6301 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     6361 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     6421 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     6481 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     6541 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     6601 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     6661 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     6721 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     6781 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6841 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6901 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6961 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     7021 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     7081 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     7141 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     7201 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     7261 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     7321 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     7381 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     7441 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     7501 ataaccgtat taccgc
//
