LOCUS       sequence-363194-        7555 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Indicator of Phosphoinositides using Akt; targeted to the plasma 
            membrane..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7555)
  AUTHORS   Ananthanarayanan B, Ni Q, Zhang J
  TITLE     Signal propagation from membrane messengers to nuclear effectors 
            revealed by reporters of phosphoinositide dynamics and Akt activity.
  JOURNAL   Proc Natl Acad Sci U S A. 2005 Oct 18;102(42):15081-6. doi: 
            10.1073/pnas.0502889102. Epub 2005 Oct 7.
  PUBMED    16214892
REFERENCE   2  (bases 1 to 7555)
  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: 
            "10.1073/pnas.0502889102"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2005-10-18- 18"; volume: "102"; issue: "42"; pages: "15081-6"
COMMENT     Sequence Label: pcDNA3-PM-InPAkt
FEATURES             Location/Qualifiers
     source          1..7555
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             922..939
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             943..975
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(979..987,988..1002)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(1008..1010,1011..1013,1014..1526)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIE"
     CDS             join(2271..2273,2274..2276,2277..2789)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGVVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     promoter        complement(3108..3126)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3109..3126)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3146..3163)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3152..3376
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3422..3850
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3509..3528)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3719..3740
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3859..3879)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3864..4193
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4044..4179
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4106..4125
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4260..5054
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4314..4333)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4924..4943
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5228..5349
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5265..5284)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5319..5338
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5398..5414)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5398..5414)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5411..5433)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5422..5438
                     /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)."
     promoter        complement(join(5446..5452,5453..5470,5471..5476))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5491..5512
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5629..5646)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5800..6388)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5880..5899)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6559..7350,7351..7419))
                     /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     7182..7201
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7420..7524)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgg tgcggggttc tcatcatcat catcatcatg gtatggctag catgactggt
      961 ggacagcaaa tgggtcggga tctgtacgac gatgacgata aggatccatg gtgagcaagg
     1021 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     1081 gccacaggtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     1141 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
     1201 tgacctgggg cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct
     1261 tcaagtccgc catgcccgaa ggctacgtcc aggagcgtac catcttcttc aaggacgacg
     1321 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     1381 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     1441 actacatcag ccacaacgtc tatatcaccg ccgacaagca gaagaacggc atcaaggccc
     1501 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
     1561 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc
     1621 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     1681 tgaccgccgc ccgcatgcat atgagcgacg tggctattgt gaaggagggt tggctgcaca
     1741 aacgagggga gtacatcaag acctggcggc cacgctactt cctcctcaag aatgatggca
     1801 ccttcattgg ctacaaggag cggccgcagg atgtggacca acgtgaggct cccctcaaca
     1861 acttctctgt ggcgcagtgc cagctgatga agacggagcg gccccggccc aacaccttca
     1921 tcatccgctg ccagcagtgg accactgtca tcgaacgcac cttccatgtg gagactcctg
     1981 aggagcggga ggagtggaca accgccatcc agactgtggc tgacggcctc aagaagcagg
     2041 aggaggagga gggtaccatg gacttccggt cgggctcacc cagtgacaac tcaggggctg
     2101 aagagatgga ggtgtccctg gccaagccca agcaccgcgt gaccatgaac gagtttgagt
     2161 acctgaagct gctgggcaag ggcactttcg gcaaggtgtc tgcaggcggt agcgtggctg
     2221 aggaagagga tgacgaggag gaagacgagg acgatggcgg cagcgagctc atggtgagca
     2281 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgtcgtaa
     2341 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     2401 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     2461 ccttcggcta cggcctgatg tgcttcgccc gctaccccga ccacatgaag cagcacgact
     2521 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     2581 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     2641 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     2701 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     2761 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     2821 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagct
     2881 accagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     2941 tcgtgaccgc cgccgggatc actctcggag ggagcggagg gacaggcatg gacgagctgt
     3001 acaagaagaa gaagaagaag agcaagacca agtgcgtgat catgtaagaa ttctgcagat
     3061 atccatcaca ctggcggccg ctcgagcatg catctagagg gccctattct atagtgtcac
     3121 ctaaatgcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3181 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3241 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3301 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     3361 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     3421 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3481 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3541 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3601 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3661 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3721 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     3781 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3841 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     3901 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     3961 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4021 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     4081 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     4141 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     4201 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca
     4261 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     4321 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     4381 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     4441 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     4501 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     4561 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     4621 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     4681 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     4741 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     4801 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     4861 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     4921 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     4981 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     5041 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     5101 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     5161 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     5221 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     5281 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     5341 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     5401 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     5461 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     5521 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     5581 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     5641 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5701 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5761 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5821 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5881 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5941 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6001 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6061 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6121 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     6181 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     6241 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     6301 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6361 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6421 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6481 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     6541 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6601 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6661 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6721 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6781 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6841 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     6901 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6961 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7021 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7081 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7141 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7201 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7261 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7321 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7381 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7441 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7501 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
