LOCUS       sequence-226770-        7516 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 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 Dec 15, 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 - Exon9 E545K
FEATURES             Location/Qualifiers
     source          1..7516
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        554..572
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             616..639
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3951..3970)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3952..3970)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    4244..4365
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4281..4300)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4335..4354
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4372..4827)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4509..4530)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4721..4740
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4854..4958
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4960..5317
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4960..5151
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4983..5003)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5168..5303
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5352..6146
                     /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             5352..6146
                     /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(5406..5425)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6016..6035
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6334..6353)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6378..6425
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6754..7342
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7243..7262
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgatt
      541 acgccaagct cgaaattaac cctcactaaa gggaacaaaa gctggagctc caccgcggtg
      601 gcggccgcca ccatggatta caaggatgac gacgataagg cccgggcgga tcccccgggc
      661 tgcaggaatt cgatagttcg aaccatgcct ccaagaccat catcaggtga actgtggggc
      721 atccacttga tgcccccaag aatcctagta gaatgtttac taccaaatgg aatgatagtg
      781 actttagaat gcctccgcga ggctacatta ataaccataa agcatgaact atttaaagaa
      841 gcaagaaaat accccctcca tcaacttctt caagatgaat cttcttacat tttcgtaagt
      901 gttactcaag aagcagaaag ggaagaattt tttgatgaaa caagacgact ttgtgacctt
      961 cggctttttc aacccttttt aaaagtaatt gaaccagtag gcaaccgtga agaaaagatc
     1021 ctcgatcgag aaattggttt tgctatcggc atgccagtgt gtgaatttga tatggttaaa
     1081 gatccagaag tacaggactt ccgaagaaat attctgaacg tttgtaaaga agctgtggat
     1141 cttagggacc tcaattcacc tcatagtaga gcaatgtatg tctatcctcc aaatgtagaa
     1201 tcttcaccag aatcgccaaa gcacatatac aataaattag ataaagggca aataatagtg
     1261 gtgatctggg taatagtttc tccaaataat gacaagcaga agtatactct gaaaatcaac
     1321 catgactgtg taccagaaca agtaattgct gaagcaatca ggaaaaaaac tcgaagtatg
     1381 ttgctatcct ctgaacaact aaaactctgt gttttagaat atcagggcaa gtatatttta
     1441 aaagtgtgtg gatgtgatga atacttccta gaaaaatatc ctctgagtca gtataagtat
     1501 ataagaagct gtataatgct agggaggatg cccaatttga tgttgatggc taaagaaagc
     1561 ctttattctc aactgccgat ggactgtttc acaatgccat cttattccag acgcatttcc
     1621 acagctacac catatatgaa tggagaagca tctacaaaat ccctttgggt tataaatagt
     1681 gcactcagaa taaaaattct ttgtgcaacc tacgtgaatg taaatattcg agacattgat
     1741 aagatctatg ttcgaacagg tatctaccat ggaggagaac ccttatgtga caatgtgaac
     1801 actcaaagag taccttgttc caatcccagg tggaatgaat ggctgaatta tgatatatac
     1861 attcctgatc ttcctcgtgc tgttcgactt tgcccttcca tttgctctgt taaaggccga
     1921 aagggtgcta aagaggaaca ctgtccattg gcatggggaa atataaactt gtttgattac
     1981 acagacactc tagtatctgg aaaaatggct ttgaatcttt ggccagtacc tcatggatta
     2041 gaagatttgc tgaaccctat tggtgttact ggatcaaatc caaataaaga aactccatgc
     2101 ttagagttgg agtttgactg gttcagcagt gtggtaaagt tcccagatat gtcagtgatt
     2161 gaagagcatg ccaattggtc tgtatcccga gaagcaggat ttagctattc ccacgcagga
     2221 ctgagtaaca gactagctag agacaatgaa ttaagggaaa atgacaaaga acagctcaaa
     2281 gcaatttcta cacgagatcc tctctctgaa atcactaagc aggagaaaga ttttctatgg
     2341 agtcacagac actattgtgt aactatcccc gaaattctac ccaaattgct tctgtctgtt
     2401 aaatggaatt ctagagatga agtagcccag atgtattgct tggtaaaaga ttggcctcca
     2461 atcaaacctg aacaggctat ggaacttctg gactgtaatt acccagatcc tatggttcga
     2521 ggttttgctg ttcggtgctt ggaaaaatat ttaacagatg acaaactttc tcagtattta
     2581 attcagctag tacaggtcct aaaatatgaa caatatttgg ataacttgct tgtgagattt
     2641 ttactgaaga aagcattgac taatcaaagg attgggcact ttttcttttg gcatttaaaa
     2701 tctgagatgc acaataaaac agttagccag aggtttggcc tgcttttgga gtcctattgt
     2761 cgtgcatgtg ggatgtattt gaagcacctg aataggcaag tcgaggcaat ggaaaagctc
     2821 attaacttaa ctgacattct caaacaggag aagaaggatg aaacacaaaa ggtacagatg
     2881 aagtttttag ttgagcaaat gaggcgacca gatttcatgg atgctctaca gggctttctg
     2941 tctcctctaa accctgctca tcaactagga aacctcaggc ttgaagagtg tcgaattatg
     3001 tcctctgcaa aaaggccact gtggttgaat tgggagaacc cagacatcat gtcagagtta
     3061 ctgtttcaga acaatgagat catctttaaa aatggggatg atttacggca agatatgcta
     3121 acacttcaaa ttattcgtat tatggaaaat atctggcaaa atcaaggtct tgatcttcga
     3181 atgttacctt atggttgtct gtcaatcggt gactgtgtgg gacttattga ggtggtgcga
     3241 aattctcaca ctattatgca aattcagtgc aaaggcggct tgaaaggtgc actgcagttc
     3301 aacagccaca cactacatca gtggctcaaa gacaagaaca aaggagaaat atatgatgca
     3361 gccattgacc tgtttacacg ttcatgtgct ggatactgtg tagctacctt cattttggga
     3421 attggagatc gtcacaatag taacatcatg gtgaaagacg atggacaact gtttcatata
     3481 gattttggac actttttgga tcacaagaag aaaaaatttg gttataaacg agaacgtgtg
     3541 ccatttgttt tgacacagga tttcttaata gtgattagta aaggagccca agaatgcaca
     3601 aagacaagag aatttgagag gtttcaggag atgtgttaca aggcttatct agctattcga
     3661 cagcatgcca atctcttcat aaatcttttc tcaatgatgc ttggctctgg aatgccagaa
     3721 ctacaatctt ttgatgacat tgcatacatt cgaaagaccc tagccttaga taaaactgag
     3781 caagaggctt tggagtattt catgaaacaa atgaatgatg cacatcatgg tggctggaca
     3841 acaaaaatgg attggatctt ccacacaatt aaacagcatg cattgaacta gctcgagggg
     3901 gggcccggta ccttaattaa ttaaggtacc aggtaagtgt acccaattcg ccctatagtg
     3961 agtcgtatta caattcactc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
     4021 ggagatccaa tttttaagtg tataatgtgt taaactactg attctaattg tttgtgtatt
     4081 ttagattcac agtcccaagg ctcatttcag gcccctcagt cctcacagtc tgttcatgat
     4141 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
     4201 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     4261 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4321 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaacgcg taaattgtaa
     4381 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     4441 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     4501 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     4561 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     4621 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     4681 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     4741 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     4801 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4861 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4921 cctgataaat gcttcaataa tattgaaaaa ggaagaatcc tgaggcggaa agaaccagct
     4981 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     5041 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     5101 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     5161 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     5221 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     5281 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     5341 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     5401 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     5461 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     5521 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     5581 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     5641 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     5701 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     5761 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     5821 atctggacga agaacatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     5881 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     5941 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     6001 gctatcagga catagcgttg gctacccgtg atattgctga agaacttggc ggcgaatggg
     6061 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     6121 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     6181 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     6241 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     6301 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     6361 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     6421 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     6481 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     6541 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     6601 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     6661 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     6721 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     6781 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     6841 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     6901 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     6961 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     7021 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     7081 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     7141 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     7201 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     7261 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     7321 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     7381 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     7441 tattaccgcc atgcattagt tattaatagt aatcaattac ggggtcatta gttcatagcc
     7501 catatatgga gttccg
//
