LOCUS       sequence-173552-        8405 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses cleavable human full-length Akt1(S473D) phosphomimetic.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8405)
  AUTHORS   Ebner M, Lucic I, Leonard TA, Yudushkin I
  TITLE     PI(3,4,5)P3 Engagement Restricts Akt Activity to Cellular Membranes.
  JOURNAL   Mol Cell. 2017 Feb 2;65(3):416-431.e6. doi: 
            10.1016/j.molcel.2016.12.028.
  PUBMED    28157504
REFERENCE   2  (bases 1 to 8405)
  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: 
            "10.1016/j.molcel.2016.12"; journalName: "Mol Cell"; date: 
            "2017-02-2- 2"; volume: "65"; issue: "3"; pages: "416-431.e6"
COMMENT     Sequence Label: pFBD-Akt13C(S473D)
FEATURES             Location/Qualifiers
     source          1..8405
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      23..611
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     512..531
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     765..782
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  916..1140
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(1207..1740)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1929..1934,1935..1951,1952..1957))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    2465..2513
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2538..2557
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     promoter        complement(4302..4411)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        4435..4526
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     4451..4468
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4575..4604
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4611..4634
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             4644..4664
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQA"
     CDS             5055..5078
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    6300..6434
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6337..6356)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6391..6410
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6463..6628
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      6812..7266
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6899..6918)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7108..7129
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        7292..7396
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7397..7465,7466..8257)
                     /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(7615..7634)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
       61 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
      121 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
      181 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
      241 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
      301 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
      361 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
      421 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
      481 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
      541 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
      601 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
      661 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
      721 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
      781 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
      841 accgcataga ccagccgcgt aacctggcaa aatcggttac ggttgagtaa taaatggatg
      901 ccctgcgtaa gcgggtgtgg gcggacaata aagtcttaaa ctgaacaaaa tagatctaaa
      961 ctatgacaat aaagtcttaa actagacaga atagttgtaa actgaaatca gtccagttat
     1021 gctgtgaaaa agcatactgg acttttgtta tggctaaagc aaactcttca ttttctgaag
     1081 tgcaaattgc ccgtcgtatt aaagaggggc gtggccaagg gcatggtaaa gactatattc
     1141 gcggcgttgt gacaatttac cgaacaactc cgcggccggg aagccgatct cggcttgaac
     1201 gaattgttag gtggcggtac ttgggtcgat atcaaagtgc atcacttctt cccgtatgcc
     1261 caactttgta tagagagcca ctgcgggatc gtcaccgtaa tctgcttgca cgtagatcac
     1321 ataagcacca agcgcgttgg cctcatgctt gaggagattg atgagcgcgg tggcaatgcc
     1381 ctgcctccgg tgctcgccgg agactgcgag atcatagata tagatctcac tacgcggctg
     1441 ctcaaacttg ggcagaacgt aagccgcgag agcgccaaca accgcttctt ggtcgaaggc
     1501 agcaagcgcg atgaatgtct tactacggag caagttcccg aggtaatcgg agtccggctg
     1561 atgttgggag taggtggcta cgtctccgaa ctcacgaccg aaaagatcaa gagcagcccg
     1621 catggatttg acttggtcag ggccgagcct acatgtgcga atgatgccca tacttgagcc
     1681 acctaacttt gttttagggc gactgccctg ctgcgtaaca tcgttgctgc tgcgtaacat
     1741 cgttgctgct ccataacatc aaacatcgac ccacggcgta acgcgcttgc tgcttggatg
     1801 cccgaggcat agactgtaca aaaaaacagt cataacaagc catgaaaacc gccactgcgc
     1861 cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt gcgtgagcgc atacgctact
     1921 tgcattacag tttacgaacc gaacaggctt atgtcaactg ggttcgtgcc ttcatccgtt
     1981 tccacggtgt gcgtcacccg gcaaccttgg gcagcagcga agtcgaggca tttctgtcct
     2041 ggctggcgaa cgagcgcaag gtttcggtct ccacgcatcg tcaggcattg gcggccttgc
     2101 tgttcttcta cggcaaggtg ctgtgcacgg atctgccctt gcttcaggag atcggtagac
     2161 ctcggccgtc gcggcgcttg ccggtggtgc tgaccccgga tgaagtggtt cgcatcctcg
     2221 gttttctgga aggcgagcat cgtttgttcg cccaggactc tagctatagt tctagtggtt
     2281 ggctacgtac ccgtagtggc tatggcaggg cttgccgccc cgacgttggc tgcgagccct
     2341 gggccttcac ccgaacttgg gggttggggt ggggaaaagg aagaaacgcg ggcgtattgg
     2401 tcccaatggg gtctcggtgg ggtatcgaca gagtgccagc cctgggaccg aaccccgcgt
     2461 ttatgaacaa acgacccaac acccgtgcgt tttattctgt ctttttattg ccgtcatagc
     2521 gcgggttcct tccggtattg tctccttccg tgtttcagtt agcctccccc atctcccggt
     2581 acctcactgc acagcggcgt ccgggtggct ctggtatcgc tgcctccaaa cctcctggat
     2641 cttcctgcac cacttgtgtg cgttcccgct ggggtccatc agataatacg tcctgttagg
     2701 cgtgtggaca aagaaagttt taaaattctt ggcctctggt cgaagttctt gtgaccaagg
     2761 aatttcacct ttcagaactt tgttgacagg atccacataa tataaatgtg gtccttctgt
     2821 gagcaacagc tgtcgtcgtc ttgcaaataa acccttccgc ttatccactg ggcccatctt
     2881 tagtattaaa ttattttcta caaactggtg ccaagggttt ccgccagcct gcttctccaa
     2941 caacaacctc ttctcatctt cggaaaactg taagtccagt tcaaaggagt tcgagtccag
     3001 atcgtgaatg tactgctcta tgttgctgcc tgacctctgg ggcaggcccg tgtcggaggc
     3061 tgacagggag tgtgaggagg aggacgaaga cacctgcatg cagccaaact ggctcaggag
     3121 attgtcataa ttgccatagc agtcctcgtc gtcttccgac atagccggca ggtaagcggt
     3181 gagcttcgga ggcgtctgct ggtgcaggtt ctcccacgtg acggactcga agaacgggtg
     3241 tgctttaaga ggtccgtatc cttccatttc ctcacagcct aaccgctttg tggcatctaa
     3301 aaccaaaagt ttctccacga ggtctcttgc cttagggaag aatttttctg gaaagtcata
     3361 ttccaactta atgatcttct gaaatataag atactcgttt ccagctcgga atggtgggag
     3421 tcctgccaca agctggtata ttatgcatcc aagagcccaa aggtctgaac tcttacaggc
     3481 ggacttctcc gtgagcagct ctggagaaac gtactgcgct gttcccacga atgagttggc
     3541 cctggcttgt ttgctctctg gggataagac ttttgctgtt ccaaaatctg tgatctggat
     3601 gtgcatatct tcatttaaca aaatgttttc cggtttaagg tccctgtgaa tgatgccctt
     3661 gccgtgcaag tactctaaag cagacacaat ctcagccgtg taaaatcggg tacaggtctc
     3721 atcgaatgaa ccgattttgc gaatatattt aagtagttct ccatttttgg cataactaag
     3781 gccgaaatac agcttctcgt cgtcctgaaa tgtgaagtaa agcttaacaa agaaggggtg
     3841 atccaggcgc gacatgacat cccgctctct ggttacatag gggaccttgt tctcttttat
     3901 gatatgtcgc ttctccagaa ttttaatcgc atattctctg gaggttgcca gttctcgagc
     3961 caggacaacc gtggaaaaag agccttcccc aaggattttc ccaaacttga agtcctcagg
     4021 ccgcttcttc cgaggctgcg gcggaggctg ggcatgctgc agggagccgg cgccgggccg
     4081 aggctcggct gcagtgccgt ccatggcggg gccctgcctg ctgccaccag gaatgccagg
     4141 gggcgtgctg gactcagtct gggtcctcac cattgatggg gatgggcagg aacataacac
     4201 cacgctggac tggatgggca cggcgtcata cagctggctg gtggtcctgg ccatgctagc
     4261 accatggctc gagatcccgg gtgatcaagt cttcgtcgag tgattgtaaa taaaatgtaa
     4321 tttacagtat agtattttaa ttaatataca aatgatttga taataattct tatttaacta
     4381 taatatattg tgttgggttg aattaaaggt ccgtatactc cggaatatta atagatcatg
     4441 gagataatta aaatgataac catctcgcaa ataaataagt attttactgt tttcgtaaca
     4501 gttttgtaat aaaaaaacct ataaatattc cggattattc ataccgtccc accatcgggc
     4561 gcggatccat gtcgcaccat caccatcacc atcaccatca ccatgcaagc tggagccacc
     4621 cgcagttcga aaagggctcc ggtgaaaacc tgtatttcca ggcgcgcatg agcgacgtgg
     4681 ctattgtgaa ggagggttgg ctgcacaaac gaggggagta catcaagacc tggcggccac
     4741 gctacttcct cctcaagaat gatggcacct tcattggcta caaggagagg ccgcaggatg
     4801 tggaccaacg tgaggctccc ctcaacaact tctctgtggc gcagtgccag ctgatgaaga
     4861 cggagcggcc ccggcccaac accttcatca tccgctgcct gcagtggacc actgtcatcg
     4921 aacgcacctt ccatgtggag actcctgagg agcgggagga gtggacaacc gccatccaga
     4981 ctgtggctga cggcctcaag aagcaggagg aggaggagat ggacttccgg tcgggctcac
     5041 ccagtgacaa ctcactagaa gttctattcc aaggacctgg ggctgaagag atggaggtgt
     5101 ccctggccaa gcccaagcac cgcgtgacca tgaacgagtt tgagtacctg aagctgctgg
     5161 gcaagggcac tttcggcaag gtgatcctgg tgaaggagaa ggccacaggc cgctactacg
     5221 ccatgaagat cctcaagaag gaagtcatcg tggccaagga cgaggtggcc cacacactca
     5281 ccgagaaccg cgtcctgcag aactccaggc accccttcct cacagccctg aagtactctt
     5341 tccagaccca cgaccgcctc tgctttgtca tggagtacgc caacgggggc gagctgttct
     5401 tccacctgtc ccgggagcgt gtgttctccg aggaccgggc ccgcttctat ggcgctgaga
     5461 ttgtgtcagc cctggactac ctgcactcgg agaagaacgt ggtgtaccgg gacctcaagc
     5521 tggagaacct catgctggac aaggacgggc acattaagat cacagacttc gggctgtgca
     5581 aggaggggat caaggacggt gccaccatga agaccttttg cggcacacct gagtacctgg
     5641 cccccgaggt gctggaggac aatgactacg gccgtgcagt ggactggtgg gggctgggcg
     5701 tggtcatgta cgagatgatg tgcggtcgcc tgcccttcta caaccaggac catgagaagc
     5761 tttttgagct catcctcatg gaggagatcc gcttcccgcg cacgcttggt cccgaggcca
     5821 agtccttgct ttcagggctg ctcaagaagg accccaagca gaggcttggc gggggctccg
     5881 aggacgccaa ggagatcatg cagcatcgct tctttgccgg tatcgtgtgg cagcacgtgt
     5941 acgagaagaa gctcagccca cccttcaagc cccaggtcac gtcggagact gacaccaggt
     6001 attttgatga ggagttcacg gcccagatga tcaccatcac accacctgac caagatgaca
     6061 gcatggagtg tgtggacagc gagcgcaggc cccacttccc ccagttcgac tactcggcca
     6121 gcagcacggc ctgagcggcc gctttcgaat ctagagcctg cagtctcgac aagcttgtcg
     6181 agaagtacta gaggatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     6241 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     6301 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     6361 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     6421 atcatgtctg gatctgatca ctgcttgagc ctaggagatc cgaaccagat aagtgaaatc
     6481 tagttccaaa ctattttgtc atttttaatt ttcgtattag cttacgacgc tacacccagt
     6541 tcccatctat tttgtcactc ttccctaaat aatccttaaa aactccattt ccacccctcc
     6601 cagttcccaa ctattttgtc cgcccacagc ggggcatttt tcttcctgtt atgtttttaa
     6661 tcaaacatcc tgccaactcc atgtgacaaa ccgtcatctt cggctacttt ttctctgtca
     6721 cagaatgaaa atttttctgt catctcttcg ttattaatgt ttgtaattga ctgaatatca
     6781 acgcttattt gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg
     6841 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     6901 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     6961 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     7021 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     7081 ttgacgttgg agtccacgtt cttaatagtg gactcttgtt ccaaactgga acaacactca
     7141 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     7201 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     7261 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     7321 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     7381 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     7441 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     7501 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     7561 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     7621 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     7681 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     7741 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     7801 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     7861 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     7921 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     7981 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     8041 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     8101 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     8161 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     8221 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     8281 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     8341 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     8401 ccccg
//
