LOCUS       pT3-EF1aH-PIK3CA H1047R        8867 bp    DNA     circular SYN 14-APR-2025
DEFINITION  This plasmid is in the pT3-EF1a vector without loxP sites flanking 
            the inverted repeats of SB (sleeping beauty) sequence. Therefore 
            this plasmid can be used in LoxP mice..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8867)
  AUTHORS   Wang C, Che L, Hu J, Zhang S, Jiang L, Latte G, Demartis MI, Tao J, 
            Gui B, Pilo MG, Ribback S, Dombrowski F, Evert M, Calvisi DF, Chen X
  TITLE     Activated mutant forms of PIK3CA cooperate with RasV12 or c-Met to 
            induce liver tumor formation in mice via AKT2/mTORC1 cascade.
  JOURNAL   Liver Int. 2015 Dec 30. doi: 10.1111/liv.13055.
  PUBMED    26716908
REFERENCE   2  (bases 1 to 8867)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            //gt12365.cn
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Liver Int. 
            2015 Dec 30. doi: 10.1111/liv.13055."
COMMENT     Sequence Label: pT3-EF1aH-PIK3CA H1047R
FEATURES             Location/Qualifiers
     source          1..8867
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     132..151
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     385..402
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    519..540
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(555..560,561..578,579..585)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    593..609
                     /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     598..620
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     617..633
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     617..633
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1059..1078)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1131..1164
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1305..2483
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1536..2474
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2431..2451
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     2500..2519
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2500..2518
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2597..2621
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2668..2694
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    complement(5937..5961)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             6014..6055
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(6093..6110)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6099..6323
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    6393..6426
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     6482..6502
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     complement(6884..6901)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6884..6900)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6893..6915)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(7109..7128)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7228..7250
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7288..7306)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7374..7478
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7479..7547,7548..8339)
                     /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(7697..7716)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(8510..8867,1..231)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
       61 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
      121 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
      181 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
      241 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
      301 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
      361 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
      421 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
      481 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
      541 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
      601 ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct tgcatgcctg
      661 caggtcgact ctagacctat acagttgaag tcggaagttt acatacactt aagttggagt
      721 cattaaaact cgtttttcaa ctactccaca aatttcttgt taacaaacaa tagttttggc
      781 aagtcagtta ggacatctac tttgtgcatg acacaagtca tttttccaac aattgtttac
      841 agacagatta tttcacttat aattcactgt atcacaattc cagtgggtca gaagtttaca
      901 tacactaagt tgactgtgcc tttaaacagc ttggaaaatt ccagaaaatg atgtcatggc
      961 tttagaagct tctgatagac taattgacat catttgagtc aattggaggt gtacctgtgg
     1021 atgtatttca aggggatcca gacatgataa gatacattga tgagtttgga caaaccacaa
     1081 ctagaatgca gcggccgcta gctcgagatc ttaattaaac tagtatcgat ataacttcgt
     1141 ataatgtatg ctatacgaag ttatatgtac gggccagata tacgcgttga cattgattat
     1201 tgactaggct tttgcaaaaa gctttgcaaa gatggataaa gttttaaaca gagaggaatc
     1261 tttgcagcta atggaccttc taggtcttga aaggagtggg aattggctcc ggtgcccgtc
     1321 agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt
     1381 gaaccggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc
     1441 tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg
     1501 ttctttttcg caacgggttt gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg
     1561 ggcctggcct ctttacgggt tatggccctt gcgtgccttg aattacttcc acctggctgc
     1621 agtacgtgat tcttgatccc gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt
     1681 gcgcttaagg agccccttcg cctcgtgctt gagttgaggc ctggcctggg cgctggggcc
     1741 gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc tgctttcgat aagtctctag
     1801 ccatttaaaa tttttgatga cctgctgcga cgcttttttt ctggcaagat agtcttgtaa
     1861 atgcgggcca agatctgcac actggtattt cggtttttgg ggccgcgggc ggcgacgggg
     1921 cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa
     1981 tcggacgggg gtagtctcaa gctggccggc ctgctctggt gcctggcctc gcgccgccgt
     2041 gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc accagttgcg tgagcggaaa
     2101 gatggccgct tcccggccct gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag
     2161 agcgggcggg tgagtcaccc acacaaagga aaagggcctt tccgtcctca gccgtcgctt
     2221 catgtgactc cacggagtac cgggcgccgt ccaggcacct cgattagttc tcgagctttt
     2281 ggagtacgtc gtctttaggt tggggggagg ggttttatgc gatggagttt ccccacactg
     2341 agtgggtgga gactgaagtt aggccagctt ggcacttgat gtaattctcc ttggaatttg
     2401 ccctttttga gtttggatct tggttcattc tcaagcctca gacagtggtt caaagttttt
     2461 ttcttccatt tcaggtgtcg tgaggaatta gcttggtact aatacgactc actataggga
     2521 gacccaagct ggctaggtaa gcttggtacc gagctcggat ccactagtcc agtgtggtgg
     2581 aattctgcag atatcaacaa gtttgtacaa aaaagcaggc tttaaaggaa ccaattcagt
     2641 cgactggatc caagcttcac catggggtac ccatacgacg tgccagatta cgccagatcc
     2701 cagtgtggtg gtacgtaccc tccaagacca tcatcaggtg aactgtgggg catccacttg
     2761 atgcccccaa gaatcctagt agaatgttta ctaccaaatg gaatgatagt gactttagaa
     2821 tgcctccgtg aggctacatt aataaccata aagcatgaac tatttaaaga agcaagaaaa
     2881 taccccctcc atcaacttct tcaagatgaa tcttcttaca ttttcgtaag tgttactcaa
     2941 gaagcagaaa gggaagaatt ttttgatgaa acaagacgac tttgtgacct tcggcttttt
     3001 caaccctttt taaaagtaat tgaaccagta ggcaaccgtg aagaaaagat cctcaatcga
     3061 gaaattggtt ttgctatcgg catgccagtg tgtgaatttg atatggttaa agatccagaa
     3121 gtacaggact tccgaagaaa tgttctgaac gtttgtaaag aagctgtgga tcttagggac
     3181 ctcaattcac ctcatagtag agcaatgtat gtctatcctc caaatgtaga atcttcacca
     3241 gaattgccaa agcacatata taataaatta gataaagggc aaataatagt ggtgatctgg
     3301 gtaatagttt ctccaaataa tgacaagcag aagtatactc tgaaaatcaa ccatgactgt
     3361 gtaccagaac aagtaattgc tgaagcaatc aggaaaaaaa ctcgaagtat gttgctatcc
     3421 tctgaacaac taaaactctg tgttttagaa tatcagggca agtatatttt aaaagtgtgt
     3481 ggatgtgatg aatacttcct agaaaaatat cctctgagtc agtataagta tataagaagc
     3541 tgtataatgc ttgggaggat gcccaatttg atgttgatgg ctaaagaaag cctttattct
     3601 caactgccaa tggactgttt tacaatgcca tcttattcca gacgcatttc cacagctaca
     3661 ccatatatga atggagaaac atctacaaaa tccctttggg ttataaatag tgcactcaga
     3721 ataaaaattc tttgtgcaac ctacgtgaat gtaaatattc gagacattga taagatctat
     3781 gttcgaacag gtatctacca tggaggagaa cccttatgtg acaatgtgaa cactcaaaga
     3841 gtaccttgtt ccaatcccag gtggaatgaa tggctgaatt atgatatata cattcctgat
     3901 cttcctcgtg ctgctcgact ttgcctttcc atttgctctg ttaaaggccg aaagggtgct
     3961 aaagaggaac actgtccatt ggcatgggga aatataaact tgtttgatta cacagacact
     4021 ctagtatctg gaaaaatggc tttgaatctt tggccagtac ctcatggatt agaagatttg
     4081 ctgaacccta ttggtgttac tggatcaaat ccaaataaag aaactccatg cttagagttg
     4141 gagtttgact ggttcagcag tgtggtaaag ttcccagata tgtcagtgat tgaagagcat
     4201 gccaattggt ctgtatcccg agaagcagga tttagctatt cccacgcagg actgagtaac
     4261 agactagcta gagacaatga attaagggaa aatgacaaag aacagctcaa agcaatttct
     4321 acacgagatc ctctctctga aatcactgag caggagaaag attttctatg gagtcacaga
     4381 cactattgtg taactatccc cgaaattcta cccaaattgc ttctgtctgt taaatggaat
     4441 tctagagatg aagtagccca gatgtattgc ttggtaaaag attggcctcc aatcaaacct
     4501 gaacaggcta tggaacttct ggactgtaat tacccagatc ctatggttcg aggttttgct
     4561 gttcggtgct tggaaaaata tttaacagat gacaaacttt ctcagtattt aattcagcta
     4621 gtacaggtcc taaaatatga acaatatttg gataacttgc ttgtgagatt tttactgaag
     4681 aaagcattga ctaatcaaag gattgggcac tttttctttt ggcatttaaa atctgagatg
     4741 cacaataaaa cagttagcca gaggtttggc ctgcttttgg agtcctattg tcgtgcatgt
     4801 gggatgtatt tgaagcacct gaataggcaa gtcgaggcaa tggaaaagct cattaactta
     4861 actgacattc tcaaacagga gaagaaggat gaaacacaaa aggtacagat gaagttttta
     4921 gttgagcaaa tgaggcgacc agatttcatg gatgctctac agggctttct gtctcctcta
     4981 aaccctgctc atcaactagg aaacctcagg cttgaagagt gtcgaattat gtcctctgca
     5041 aaaaggccac tgtggttgaa ttgggagaac ccagacatca tgtcagagtt actgtttcag
     5101 aacaatgaga tcatctttaa aaatggggat gatttacggc aagatatgct aacacttcaa
     5161 attattcgta ttatggaaaa tatctggcaa aatcaaggtc ttgatcttcg aatgttacct
     5221 tatggttgtc tgtcaatcgg tgactgtgtg ggacttattg aggtggtgcg aaattctcac
     5281 actattatgc aaattcagtg caaaggcggc ttgaaaggtg cactgcagtt caacagccac
     5341 acactacatc agtggctcaa agacaagaac aaaggagaaa tatatgatgc agccattgac
     5401 ctgtttacac gttcatgtgc tggatactgt gtagctacct tcattttggg aattggagat
     5461 cgtcacaata gtaacatcat ggtgaaagac gatggacaac tgtttcatat agattttgga
     5521 cactttttgg atcacaagaa gaaaaaattt ggttataaac gagaacgtgt gccatttgtt
     5581 ttgacacagg atttcttaat agtgattagt aaaggagccc aagaatgcac aaagacaaga
     5641 gaatttgaga ggtttcagga gatgtgttac aaggcttatc tagctattcg acagcatgcc
     5701 aatctcttca taaatctttt ctcaatgatg cttggctctg gaatgccaga actacaatct
     5761 tttgatgaca ttgcatacat tcgaaagacc ctagccttag ataaaactga gcaagaggct
     5821 ttggagtatt tcatgaaaca aatgaatgat gcacgtcatg gtggctggac aacaaaaatg
     5881 gattggatct tccacacaat taaacagcat gcattgaact gagtcgagat atctagaccc
     5941 agctttcttg tacaaagtgg ttgatatcca gcacagtggc ggccgctcga gtctagaggg
     6001 cccgcggttc gaaggtaagc ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac
     6061 cggttagtaa tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     6121 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     6181 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     6241 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     6301 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg
     6361 gtatccccac gcgccctgta gcggcgcatt aaataacttc gtataatgta tgctatacga
     6421 agttatatcg tatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
     6481 caccctaact gacacacatt ccacagaatt ccttgaaata catccacagg tacacctcca
     6541 attgactcaa atgatgtcaa ttagtctatc agaagcttct aaagccatga catcattttc
     6601 tggaattttc caagctgttt aaaggcacag tcaacttagt gtatgtaaac ttctgaccca
     6661 ctggaattgt gatacagtga attataagtg aaataatctg tctgtaaaca attgttggaa
     6721 aaatgacttg tgtcatgcac aaagtagatg tcctaactga cttgccaaaa ctattgtttg
     6781 ttaacaagaa atttgtggag tagttgaaaa acgagtttta atgactccaa cttaagtgta
     6841 tgtaaacttc cgacttcaac tgtagggtac cgagctcgaa ttcactggcc gtcgttttac
     6901 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     6961 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     7021 gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     7081 tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     7141 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     7201 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     7261 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     7321 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     7381 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     7441 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     7501 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     7561 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     7621 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     7681 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     7741 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     7801 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     7861 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     7921 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     7981 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     8041 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     8101 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     8161 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     8221 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     8281 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     8341 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     8401 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     8461 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     8521 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     8581 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     8641 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     8701 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     8761 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     8821 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacg
//