LOCUS       sequence-181814-        5034 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses eGFP-tagged PH(WT) domain of human Akt1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5034)
  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 5034)
  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-PH1(WT)
FEATURES             Location/Qualifiers
     source          1..5034
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      4..592
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     493..512
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        767..1070
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1071..1274
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1225..1245
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1313..1322
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1319..1321,1322..1324,1325..2035)
                     /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(1364..1385)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1625..1644)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1972..1993
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2528..2649
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2565..2584)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2619..2638
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2656..3111)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2793..2814)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3005..3024
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3138..3242
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3244..3601
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3244..3435
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3267..3287)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3452..3587
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3636..4430
                     /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             3636..4430
                     /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(3690..3709)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4300..4319
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4618..4637)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4662..4709
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
       61 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
      121 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
      181 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
      241 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
      301 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
      361 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
      421 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
      481 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
      541 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
      601 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
      661 tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcattagt tattaatagt
      721 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
      781 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
      841 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
      901 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
      961 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     1021 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
     1081 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     1141 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     1201 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     1261 atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg tcgccaccat
     1321 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     1381 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     1441 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     1501 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     1561 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     1621 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     1681 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     1741 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     1801 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     1861 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     1921 cctgagcacc cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     1981 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtccgg
     2041 actcagatct cgagctatga gcgacgtggc tattgtgaag gagggttggc tgcacaaacg
     2101 aggggagtac atcaagacct ggcggccacg ctacttcctc ctcaagaatg atggcacctt
     2161 cattggctac aaggagcggc cgcaggatgt ggaccaacgt gaggctcccc tcaacaactt
     2221 ctctgtggcg cagtgccagc tgatgaagac ggagcggccc cggcccaaca ccttcatcat
     2281 ccgctgcctg cagtggacca ctgtcatcga acgcaccttc catgtggaga ctcctgagga
     2341 gcgggaggag tggacaaccg ccatccagac tgtggctgac ggcctcaaga agcaggagga
     2401 ggaggagtaa tctagataac tgatcataat cagccatacc acatttgtag aggttttact
     2461 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     2521 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     2581 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     2641 tgtatcttaa cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     2701 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     2761 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     2821 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     2881 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     2941 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     3001 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     3061 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     3121 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     3181 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     3241 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     3301 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     3361 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3421 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     3481 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     3541 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3601 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     3661 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     3721 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     3781 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     3841 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     3901 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     3961 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     4021 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     4081 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     4141 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     4201 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4261 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     4321 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     4381 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     4441 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     4501 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4561 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     4621 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     4681 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     4741 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     4801 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     4861 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     4921 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     4981 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatc
//
