LOCUS       sequence-343414-        7514 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nuclear-targeted AktAR2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7514)
  AUTHORS   Zhou X, Zhong Y, Molinar-Inglis O, Kunkel MT, Chen M, Sun T, Zhang 
            J, Shyy JY, Trejo J, Newton AC, Zhang J
  TITLE     Location-specific inhibition of Akt reveals regulation of mTORC1 
            activity in the nucleus.
  JOURNAL   Nat Commun. 2020 Nov 30;11(1):6088. doi: 10.1038/s41467-020-19937-w.
  PUBMED    33257668
REFERENCE   2  (bases 1 to 7514)
  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"; journalName: "Nat 
            Commun."; date: "2020-11-30"; volume: "11(1):6088. doi"; pages: " 
            10.1038/s41467-020-19937-w"
COMMENT     Sequence Label: pcDNA3-AktAR2-NLS
FEATURES             Location/Qualifiers
     source          1..7514
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(112..131)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        303..682
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        683..886
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     837..857
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     931..950
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        931..949
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      969..978
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             990..1007
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1011..1043
                     /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(1047..1055,1056..1070)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(1077..1079,1080..1082,1083..1793)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2346..3080
                     /codon_start=1
                     /product="circularly permuted form of the Venus yellow 
                     fluorescent protein (Nagai et al., 2004)"
                     /label=cp173Venus
                     /note="efficient FRET acceptor for CFP"
                     /translation="MGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDH
                     MVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEELFTGVVPILVELDGDVNGHKFSVSG
                     EGEGDATYGKLTLKLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPE
                     GYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHN
                     VYITADKQKNGIKANFKIRHNIE"
     CDS             3087..3107
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     promoter        complement(3135..3153)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3136..3153)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3173..3190)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3179..3403
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3449..3877
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3536..3555)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3746..3767
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3886..3906)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3891..4220
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4071..4206
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4133..4152
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4287..5081
                     /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(4341..4360)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4951..4970
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5255..5376
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5292..5311)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5346..5365
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5425..5441)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5425..5441)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5438..5460)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5449..5465
                     /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(5473..5479,5480..5497,5498..5503))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5518..5539
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5656..5673)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5827..6415)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5907..5926)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6586..7377,7378..7446))
                     /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     7209..7228
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7447..7514,1..37))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
       61 ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg
      121 ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga
      181 gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa
      241 gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg
      301 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
      361 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
      421 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
      481 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
      541 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
      601 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
      661 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
      721 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
      781 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
      841 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag
      901 agaacccact gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc
      961 ttgcggccgc caccatggtg cggggttctc atcatcatca tcatcatggt atggctagca
     1021 tgactggtgg acagcaaatg ggtcgggatc tgtacgacga tgacgataag gatcccatgg
     1081 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
     1141 acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca
     1201 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
     1261 tgaccaccct gagctggggc gtgcagtgct tcgcccgcta ccccgaccac atgaagcagc
     1321 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
     1381 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     1441 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     1501 tggagtacaa cgccatccac ggcaacgtct atatcaccgc cgacaagcag aagaacggca
     1561 tcaaggccaa cttcggcctc aactgcaaca tcgaggacgg cagcgtgcag ctcgccgacc
     1621 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     1681 tgagcaccca gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     1741 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagcgcatgc
     1801 ataagttttc tcaagaacag atcggcgaaa acattgtgtg cagggtcatt tgtaccacgg
     1861 gtcaaattcc catccgagat ttgtcagctg atatttcaca agtgcttaag gaaaaacgat
     1921 ccataaagaa agtttggaca tttggtagaa acccagcctg tgactatcat ttaggaaaca
     1981 tttcaagact gtcaaataag catttccaaa tactactagg agaagacggt aaccttttat
     2041 tgaatgacat ttccactaat gggacctggt taaatgggca aaaagtcgag aagaacagca
     2101 atcagttact gtctcaaggt gatgaaataa ccgttggtgt aggcgtggaa tcagatattt
     2161 tatctctggt cattttcata aacgacaaat ttaagcagtg cctcgagcag aacaaagttg
     2221 atcgctctgc aggtaagcca ggcagcggcg agggcagcac caagggcctg gtcgaccctc
     2281 gtccgcgctc gtgcacctgg ccggacccca ggccggagtt tggaggtacc ggcggcagcg
     2341 agctcatggg cggcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     2401 gccccgtgct gctgcccgac aaccactacc tgagctacca gtccaagctg agcaaagacc
     2461 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     2521 tcggcatgga cgagctgtac aagggaggta ccggtggatc tatggtgagc aagggcgagg
     2581 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     2641 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagc
     2701 tgatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgggct
     2761 acggccttca gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     2821 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     2881 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     2941 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     3001 acagccacaa cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttca
     3061 agatccgcca caacatcgag gaattcccca aaaagaagag aaaggtggaa gacgcataat
     3121 ctagagggcc ctattctata gtgtcaccta aatgctagag ctcgctgatc agcctcgact
     3181 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     3241 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     3301 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     3361 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     3421 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     3481 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     3541 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     3601 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     3661 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     3721 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     3781 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     3841 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     3901 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3961 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4021 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     4081 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     4141 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg
     4201 gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc
     4261 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     4321 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     4381 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     4441 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca
     4501 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     4561 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     4621 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     4681 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     4741 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     4801 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     4861 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     4921 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     4981 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     5041 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     5101 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     5161 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     5221 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     5281 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     5341 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     5401 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     5461 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     5521 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     5581 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     5641 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     5701 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     5761 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     5821 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5881 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5941 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     6001 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     6061 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     6121 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     6181 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     6241 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     6301 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     6361 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6421 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     6481 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     6541 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     6601 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     6661 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6721 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6781 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6841 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6901 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6961 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     7021 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     7081 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     7141 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     7201 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     7261 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     7321 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     7381 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     7441 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     7501 cggatacata tttg
//
