LOCUS       sequence-173549-        8381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human full-length Akt1(E17K) mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8381)
  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 8381)
  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-Akt1(E17K)
FEATURES             Location/Qualifiers
     source          1..8381
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      18..606
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     507..526
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     760..777
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  911..1135
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(1202..1735)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1924..1929,1930..1946,1947..1952))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     polyA_signal    2460..2508
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     2533..2552
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     promoter        complement(4297..4406)
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     promoter        4430..4521
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     4446..4463
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4570..4599
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4606..4629
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             4639..4659
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQA"
     CDS             4663..6102
                     /codon_start=1
                     /gene="AKT1"
                     /product="human serine/threonine protein kinase activated 
                     by growth factors"
                     /label=Akt1
                     /translation="MSDVAIVKEGWLHKRGKYIKTWRPRYFLLKNDGTFIGYKERPQDV
                     DQREAPLNNFSVAQCQLMKTERPRPNTFIIRCLQWTTVIERTFHVETPEEREEWTTAIQ
                     TVADGLKKQEEEEMDFRSGSPSDNSGAEEMEVSLAKPKHRVTMNEFEYLKLLGKGTFGK
                     VILVKEKATGRYYAMKILKKEVIVAKDEVAHTLTENRVLQNSRHPFLTALKYSFQTHDR
                     LCFVMEYANGGELFFHLSRERVFSEDRARFYGAEIVSALDYLHSEKNVVYRDLKLENLM
                     LDKDGHIKITDFGLCKEGIKDGATMKTFCGTPEYLAPEVLEDNDYGRAVDWWGLGVVMY
                     EMMCGRLPFYNQDHEKLFELILMEEIRFPRTLGPEAKSLLSGLLKKDPKQRLGGGSEDA
                     KEIMQHRFFAGIVWQHVYEKKLSPPFKPQVTSETDTRYFDEEFTAQMITITPPDQDDSM
                     ECVDSERRPHFPQFSYSASSTA"
     polyA_signal    6271..6405
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6308..6327)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6362..6381
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6434..6599
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      6783..7237
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6870..6889)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7079..7100
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        7263..7367
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7368..7436,7437..8228)
                     /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(7586..7605)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
       61 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
      121 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
      181 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
      241 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
      301 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
      361 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
      421 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
      481 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
      541 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
      601 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
      661 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
      721 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
      781 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
      841 atagaccagc cgcgtaacct ggcaaaatcg gttacggttg agtaataaat ggatgccctg
      901 cgtaagcggg tgtgggcgga caataaagtc ttaaactgaa caaaatagat ctaaactatg
      961 acaataaagt cttaaactag acagaatagt tgtaaactga aatcagtcca gttatgctgt
     1021 gaaaaagcat actggacttt tgttatggct aaagcaaact cttcattttc tgaagtgcaa
     1081 attgcccgtc gtattaaaga ggggcgtggc caagggcatg gtaaagacta tattcgcggc
     1141 gttgtgacaa tttaccgaac aactccgcgg ccgggaagcc gatctcggct tgaacgaatt
     1201 gttaggtggc ggtacttggg tcgatatcaa agtgcatcac ttcttcccgt atgcccaact
     1261 ttgtatagag agccactgcg ggatcgtcac cgtaatctgc ttgcacgtag atcacataag
     1321 caccaagcgc gttggcctca tgcttgagga gattgatgag cgcggtggca atgccctgcc
     1381 tccggtgctc gccggagact gcgagatcat agatatagat ctcactacgc ggctgctcaa
     1441 acttgggcag aacgtaagcc gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa
     1501 gcgcgatgaa tgtcttacta cggagcaagt tcccgaggta atcggagtcc ggctgatgtt
     1561 gggagtaggt ggctacgtct ccgaactcac gaccgaaaag atcaagagca gcccgcatgg
     1621 atttgacttg gtcagggccg agcctacatg tgcgaatgat gcccatactt gagccaccta
     1681 actttgtttt agggcgactg ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg
     1741 ctgctccata acatcaaaca tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga
     1801 ggcatagact gtacaaaaaa acagtcataa caagccatga aaaccgccac tgcgccgtta
     1861 ccaccgctgc gttcggtcaa ggttctggac cagttgcgtg agcgcatacg ctacttgcat
     1921 tacagtttac gaaccgaaca ggcttatgtc aactgggttc gtgccttcat ccgtttccac
     1981 ggtgtgcgtc acccggcaac cttgggcagc agcgaagtcg aggcatttct gtcctggctg
     2041 gcgaacgagc gcaaggtttc ggtctccacg catcgtcagg cattggcggc cttgctgttc
     2101 ttctacggca aggtgctgtg cacggatctg cccttgcttc aggagatcgg tagacctcgg
     2161 ccgtcgcggc gcttgccggt ggtgctgacc ccggatgaag tggttcgcat cctcggtttt
     2221 ctggaaggcg agcatcgttt gttcgcccag gactctagct atagttctag tggttggcta
     2281 cgtacccgta gtggctatgg cagggcttgc cgccccgacg ttggctgcga gccctgggcc
     2341 ttcacccgaa cttgggggtt ggggtgggga aaaggaagaa acgcgggcgt attggtccca
     2401 atggggtctc ggtggggtat cgacagagtg ccagccctgg gaccgaaccc cgcgtttatg
     2461 aacaaacgac ccaacacccg tgcgttttat tctgtctttt tattgccgtc atagcgcggg
     2521 ttccttccgg tattgtctcc ttccgtgttt cagttagcct cccccatctc ccggtacctc
     2581 actgcacagc ggcgtccggg tggctctggt atcgctgcct ccaaacctcc tggatcttcc
     2641 tgcaccactt gtgtgcgttc ccgctggggt ccatcagata atacgtcctg ttaggcgtgt
     2701 ggacaaagaa agttttaaaa ttcttggcct ctggtcgaag ttcttgtgac caaggaattt
     2761 cacctttcag aactttgttg acaggatcca cataatataa atgtggtcct tctgtgagca
     2821 acagctgtcg tcgtcttgca aataaaccct tccgcttatc cactgggccc atctttagta
     2881 ttaaattatt ttctacaaac tggtgccaag ggtttccgcc agcctgcttc tccaacaaca
     2941 acctcttctc atcttcggaa aactgtaagt ccagttcaaa ggagttcgag tccagatcgt
     3001 gaatgtactg ctctatgttg ctgcctgacc tctggggcag gcccgtgtcg gaggctgaca
     3061 gggagtgtga ggaggaggac gaagacacct gcatgcagcc aaactggctc aggagattgt
     3121 cataattgcc atagcagtcc tcgtcgtctt ccgacatagc cggcaggtaa gcggtgagct
     3181 tcggaggcgt ctgctggtgc aggttctccc acgtgacgga ctcgaagaac gggtgtgctt
     3241 taagaggtcc gtatccttcc atttcctcac agcctaaccg ctttgtggca tctaaaacca
     3301 aaagtttctc cacgaggtct cttgccttag ggaagaattt ttctggaaag tcatattcca
     3361 acttaatgat cttctgaaat ataagatact cgtttccagc tcggaatggt gggagtcctg
     3421 ccacaagctg gtatattatg catccaagag cccaaaggtc tgaactctta caggcggact
     3481 tctccgtgag cagctctgga gaaacgtact gcgctgttcc cacgaatgag ttggccctgg
     3541 cttgtttgct ctctggggat aagacttttg ctgttccaaa atctgtgatc tggatgtgca
     3601 tatcttcatt taacaaaatg ttttccggtt taaggtccct gtgaatgatg cccttgccgt
     3661 gcaagtactc taaagcagac acaatctcag ccgtgtaaaa tcgggtacag gtctcatcga
     3721 atgaaccgat tttgcgaata tatttaagta gttctccatt tttggcataa ctaaggccga
     3781 aatacagctt ctcgtcgtcc tgaaatgtga agtaaagctt aacaaagaag gggtgatcca
     3841 ggcgcgacat gacatcccgc tctctggtta cataggggac cttgttctct tttatgatat
     3901 gtcgcttctc cagaatttta atcgcatatt ctctggaggt tgccagttct cgagccagga
     3961 caaccgtgga aaaagagcct tccccaagga ttttcccaaa cttgaagtcc tcaggccgct
     4021 tcttccgagg ctgcggcgga ggctgggcat gctgcaggga gccggcgccg ggccgaggct
     4081 cggctgcagt gccgtccatg gcggggccct gcctgctgcc accaggaatg ccagggggcg
     4141 tgctggactc agtctgggtc ctcaccattg atggggatgg gcaggaacat aacaccacgc
     4201 tggactggat gggcacggcg tcatacagct ggctggtggt cctggccatg ctagcaccat
     4261 ggctcgagat cccgggtgat caagtcttcg tcgagtgatt gtaaataaaa tgtaatttac
     4321 agtatagtat tttaattaat atacaaatga tttgataata attcttattt aactataata
     4381 tattgtgttg ggttgaatta aaggtccgta tactccggaa tattaataga tcatggagat
     4441 aattaaaatg ataaccatct cgcaaataaa taagtatttt actgttttcg taacagtttt
     4501 gtaataaaaa aacctataaa tattccggat tattcatacc gtcccaccat cgggcgcgga
     4561 tccatgtcgc accatcacca tcaccatcac catcaccatg caagctggag ccacccgcag
     4621 ttcgaaaagg gctccggtga aaacctgtat ttccaggcgc gcatgagcga cgtggctatt
     4681 gtgaaggagg gttggctgca caaacgaggg aagtacatca agacctggcg gccacgctac
     4741 ttcctcctca agaatgatgg caccttcatt ggctacaagg agcggccgca ggatgtggac
     4801 caacgtgagg ctcccctcaa caacttctct gtggcgcagt gccagctgat gaagacggag
     4861 cggccccggc ccaacacctt catcatccgc tgcctgcagt ggaccactgt catcgaacgc
     4921 accttccatg tggagactcc tgaggagcgg gaggagtgga caaccgccat ccagactgtg
     4981 gctgacggcc tcaagaagca ggaggaggag gagatggact tccggtcggg ctcacccagt
     5041 gacaactcag gggctgaaga gatggaggtg tccctggcca agcccaagca ccgcgtgacc
     5101 atgaacgagt ttgagtacct gaagctgctg ggcaagggca ctttcggcaa ggtgatcctg
     5161 gtgaaggaga aggccacagg ccgctactac gccatgaaga tcctcaagaa ggaagtcatc
     5221 gtggccaagg acgaggtggc ccacacactc accgagaacc gcgtcctgca gaactccagg
     5281 caccccttcc tcacagccct gaagtactct ttccagaccc acgaccgcct ctgctttgtc
     5341 atggagtacg ccaacggggg cgagctgttc ttccacctgt cccgggagcg tgtgttctcc
     5401 gaggaccggg cccgcttcta tggcgctgag attgtgtcag ccctggacta cctgcactcg
     5461 gagaagaacg tggtgtaccg ggacctcaag ctggagaacc tcatgctgga caaggacggg
     5521 cacattaaga tcacagactt cgggctgtgc aaggagggga tcaaggacgg tgccaccatg
     5581 aagacctttt gcggcacacc tgagtacctg gcccccgagg tgctggagga caatgactac
     5641 ggccgtgcag tggactggtg ggggctgggc gtggtcatgt acgagatgat gtgcggtcgc
     5701 ctgcccttct acaaccagga ccatgagaag ctttttgagc tcatcctcat ggaggagatc
     5761 cgcttcccgc gcacgcttgg tcccgaggcc aagtccttgc tttcagggct gctcaagaag
     5821 gaccccaagc agaggcttgg cgggggctcc gaggacgcca aggagatcat gcagcatcgc
     5881 ttctttgccg gtatcgtgtg gcagcacgtg tacgagaaga agctcagccc acccttcaag
     5941 ccccaggtca cgtcggagac tgacaccagg tattttgatg aggagttcac ggcccagatg
     6001 atcaccatca caccacctga ccaagatgac agcatggagt gtgtggacag cgagcgcagg
     6061 ccccacttcc cccagttctc ctactcggcc agcagcacgg cctgagcggc cgctttcgaa
     6121 tctagagcct gcagtctcga caagcttgtc gagaagtact agaggatcat aatcagccat
     6181 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
     6241 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
     6301 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     6361 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatctgatc actgcttgag
     6421 cctaggagat ccgaaccaga taagtgaaat ctagttccaa actattttgt catttttaat
     6481 tttcgtatta gcttacgacg ctacacccag ttcccatcta ttttgtcact cttccctaaa
     6541 taatccttaa aaactccatt tccacccctc ccagttccca actattttgt ccgcccacag
     6601 cggggcattt ttcttcctgt tatgttttta atcaaacatc ctgccaactc catgtgacaa
     6661 accgtcatct tcggctactt tttctctgtc acagaatgaa aatttttctg tcatctcttc
     6721 gttattaatg tttgtaattg actgaatatc aacgcttatt tgcagcctga atggcgaatg
     6781 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     6841 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     6901 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     6961 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     7021 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tcttaatagt
     7081 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     7141 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     7201 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     7261 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     7321 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     7381 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     7441 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     7501 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     7561 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     7621 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     7681 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     7741 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     7801 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     7861 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     7921 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     7981 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     8041 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     8101 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     8161 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     8221 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     8281 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     8341 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga a
//
