LOCUS       sequence-174214-        6123 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses eGFP-tagged human Akt1(WT).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6123)
  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 6123)
  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: pEGFP-Akt1(WT)
FEATURES             Location/Qualifiers
     source          1..6123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      16..604
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     505..524
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        779..1082
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1083..1286
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1237..1257
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1325..1334
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1331..1333,1334..1336,1337..2047)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1376..1397)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1637..1656)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1984..2005
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2069..3508
                     /codon_start=1
                     /gene="AKT1"
                     /product="human serine/threonine protein kinase activated 
                     by growth factors"
                     /label=Akt1
                     /translation="MSDVAIVKEGWLHKRGEYIKTWRPRYFLLKNDGTFIGYKERPQDV
                     DQREAPLNNFSVAQCQLMKTERPRPNTFIIRCLQWTTVIERTFHVETPEEREEWTTAIQ
                     TVADGLKKQEEEEMDFRSGSPSDNSGAEEMEVSLAKPKHRVTMNEFEYLKLLGKGTFGK
                     VILVKEKATGRYYAMKILKKEVIVAKDEVAHTLTENRVLQNSRHPFLTALKYSFQTHDR
                     LCFVMEYANGGELFFHLSRERVFSEDRARFYGAEIVSALDYLHSEKNVVYRDLKLENLM
                     LDKDGHIKITDFGLCKEGIKDGATMKTFCGTPEYLAPEVLEDNDYGRAVDWWGLGVVMY
                     EMMCGRLPFYNQDHEKLFELILMEEIRFPRTLGPEAKSLLSGLLKKDPKQRLGGGSEDA
                     KEIMQHRFFAGIVWQHVYEKKLSPPFKPQVTSETDTRYFDEEFTAQMITITPPDQDDSM
                     ECVDSERRPHFPQFSYSASGTA"
     polyA_signal    3629..3750
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3666..3685)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3720..3739
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3757..4212)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3894..3915)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4106..4125
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4239..4343
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4345..4702
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4345..4536
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4368..4388)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4553..4688
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4737..5531
                     /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             4737..5531
                     /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(4791..4810)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5401..5420
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5719..5738)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5763..5810
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
       61 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
      121 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
      181 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
      241 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
      301 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
      361 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
      421 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
      481 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
      541 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
      601 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
      661 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcatta
      721 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
      781 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      841 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      901 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      961 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
     1021 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     1081 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
     1141 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
     1201 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac
     1261 ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc tagcgctacc
     1321 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     1381 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     1441 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     1501 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     1561 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     1621 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     1681 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     1741 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     1801 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     1861 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     1921 cgacaaccac tacctgagca cccagtccaa gctgagcaaa gaccccaacg agaagcgcga
     1981 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     2041 gtacaagtcc ggactcagat ctcgagctat gagcgacgtg gctattgtga aggagggttg
     2101 gctgcacaaa cgaggggagt acatcaagac ctggcggcca cgctacttcc tcctcaagaa
     2161 tgatggcacc ttcattggct acaaggagcg gccgcaggat gtggaccaac gtgaggctcc
     2221 cctcaacaac ttctctgtgg cgcagtgcca gctgatgaag acggagcggc cccggcccaa
     2281 caccttcatc atccgctgcc tgcagtggac cactgtcatc gaacgcacct tccatgtgga
     2341 gactcctgag gagcgggagg agtggacaac cgccatccag actgtggctg acggcctcaa
     2401 gaagcaggag gaggaggaga tggacttccg gtcgggctca cccagtgaca actcaggggc
     2461 tgaagagatg gaggtgtccc tggccaagcc caagcaccgc gtgaccatga acgagtttga
     2521 gtacctgaag ctgctgggca agggcacttt cggcaaggtg atcctggtga aggagaaggc
     2581 cacaggccgc tactacgcca tgaagatcct caagaaggaa gtcatcgtgg ccaaggacga
     2641 ggtggcccac acactcaccg agaaccgcgt cctgcagaac tccaggcacc ccttcctcac
     2701 agccctgaag tactctttcc agacccacga ccgcctctgc tttgtcatgg agtacgccaa
     2761 cgggggcgag ctgttcttcc acctgtcccg ggagcgtgtg ttctccgagg accgggcccg
     2821 cttctatggc gctgagattg tgtcagccct ggactacctg cactcggaga agaacgtggt
     2881 gtaccgggac ctcaagctgg agaacctcat gctggacaag gacgggcaca ttaagatcac
     2941 agacttcggg ctgtgcaagg aggggatcaa ggacggtgcc accatgaaga ccttttgcgg
     3001 cacacctgag tacctggccc ccgaggtgct ggaggacaat gactacggcc gtgcagtgga
     3061 ctggtggggg ctgggcgtgg tcatgtacga gatgatgtgc ggtcgcctgc ccttctacaa
     3121 ccaggaccat gagaagcttt ttgagctcat cctcatggag gagatccgct tcccgcgcac
     3181 gcttggtccc gaggccaagt ccttgctttc agggctgctc aagaaggacc ccaagcagag
     3241 gcttggcggg ggctccgagg acgccaagga gatcatgcag catcgcttct ttgccggtat
     3301 cgtgtggcag cacgtgtacg agaagaagct cagcccaccc ttcaagcccc aggtcacgtc
     3361 ggagactgac accaggtatt ttgatgagga gttcacggcc cagatgatca ccatcacacc
     3421 acctgaccaa gatgacagca tggagtgtgt ggacagcgag cgcaggcccc acttccccca
     3481 gttctcctac tcggccagcg gcacggcctg atctagataa ctgatcataa tcagccatac
     3541 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     3601 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     3661 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     3721 tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt
     3781 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     3841 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     3901 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     3961 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     4021 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     4081 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     4141 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     4201 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4261 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4321 aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca
     4381 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4441 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4501 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4561 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4621 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4681 tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga
     4741 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4801 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4861 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag
     4921 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4981 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     5041 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     5101 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     5161 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     5221 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg
     5281 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     5341 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     5401 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     5461 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     5521 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     5581 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     5641 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     5701 ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat
     5761 gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg
     5821 ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata
     5881 cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct
     5941 cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta
     6001 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     6061 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     6121 aaa
//
