LOCUS       sequence-344926-        7511 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cytosol-targeted AktAR2..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7511)
  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 7511)
  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-NES
FEATURES             Location/Qualifiers
     source          1..7511
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        187..566
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        567..770
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     721..741
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     815..834
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        815..833
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      853..862
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             874..891
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             895..927
                     /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(931..939,940..954)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(961..963,964..966,967..1677)
                     /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             2230..2964
                     /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             2971..2997
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     promoter        complement(3016..3034)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3017..3034)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3054..3071)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3060..3284
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3330..3758
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3417..3436)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3627..3648
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3767..3787)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3772..4101
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3952..4087
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4014..4033
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4168..4962
                     /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(4222..4241)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4832..4851
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5136..5257
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5173..5192)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5227..5246
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5306..5322)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5306..5322)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5319..5341)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5330..5346
                     /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(5354..5360,5361..5378,5379..5384))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5399..5420
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5537..5554)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5708..6296)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5788..5807)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6467..7258,7259..7327))
                     /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     7090..7109
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7328..7432)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(join(7507..7511,1..15))
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
       61 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
      121 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
      181 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
      241 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      301 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      361 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      421 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      481 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      541 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      601 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      661 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      721 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa
      781 ctagagaacc cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc
      841 aagcttgcgg ccgccaccat ggtgcggggt tctcatcatc atcatcatca tggtatggct
      901 agcatgactg gtggacagca aatgggtcgg gatctgtacg acgatgacga taaggatccc
      961 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     1021 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     1081 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     1141 ctcgtgacca ccctgagctg gggcgtgcag tgcttcgccc gctaccccga ccacatgaag
     1201 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     1261 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     1321 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     1381 aagctggagt acaacgccat ccacggcaac gtctatatca ccgccgacaa gcagaagaac
     1441 ggcatcaagg ccaacttcgg cctcaactgc aacatcgagg acggcagcgt gcagctcgcc
     1501 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     1561 tacctgagca cccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     1621 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagcgc
     1681 atgcataagt tttctcaaga acagatcggc gaaaacattg tgtgcagggt catttgtacc
     1741 acgggtcaaa ttcccatccg agatttgtca gctgatattt cacaagtgct taaggaaaaa
     1801 cgatccataa agaaagtttg gacatttggt agaaacccag cctgtgacta tcatttagga
     1861 aacatttcaa gactgtcaaa taagcatttc caaatactac taggagaaga cggtaacctt
     1921 ttattgaatg acatttccac taatgggacc tggttaaatg ggcaaaaagt cgagaagaac
     1981 agcaatcagt tactgtctca aggtgatgaa ataaccgttg gtgtaggcgt ggaatcagat
     2041 attttatctc tggtcatttt cataaacgac aaatttaagc agtgcctcga gcagaacaaa
     2101 gttgatcgct ctgcaggtaa gccaggcagc ggcgagggca gcaccaaggg cctggtcgac
     2161 cctcgtccgc gctcgtgcac ctggccggac cccaggccgg agtttggagg taccggcggc
     2221 agcgagctca tgggcggcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     2281 gacggccccg tgctgctgcc cgacaaccac tacctgagct accagtccaa gctgagcaaa
     2341 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     2401 actctcggca tggacgagct gtacaaggga ggtaccggtg gatctatggt gagcaagggc
     2461 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     2521 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     2581 aagctgatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     2641 ggctacggcc ttcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc
     2701 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     2761 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     2821 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     2881 tacaacagcc acaacgtcta tatcaccgcc gacaagcaga agaacggcat caaggccaac
     2941 ttcaagatcc gccacaacat cgaggaattc ctgccccccc tggagcgcct gaccctgtaa
     3001 tctagagggc cctattctat agtgtcacct aaatgctaga gctcgctgat cagcctcgac
     3061 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     3121 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     3181 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     3241 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     3301 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     3361 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     3421 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     3481 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     3541 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     3601 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     3661 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     3721 aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag
     3781 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     3841 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3901 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     3961 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     4021 gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt
     4081 ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat
     4141 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     4201 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     4261 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     4321 gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc
     4381 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     4441 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     4501 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     4561 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     4621 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     4681 gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     4741 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     4801 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     4861 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     4921 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     4981 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     5041 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     5101 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg
     5161 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     5221 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct
     5281 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     5341 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     5401 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     5461 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5521 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5581 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5641 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5701 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5761 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5821 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5881 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5941 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6001 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     6061 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6121 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     6181 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6241 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6301 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6361 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     6421 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     6481 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     6541 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     6601 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     6661 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     6721 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     6781 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     6841 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     6901 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     6961 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     7021 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     7081 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     7141 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     7201 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     7261 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     7321 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     7381 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     7441 cccgaaaagt gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac
     7501 tctcagtaca a
//
