LOCUS       sequence-344935-        6341 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasma membrane-targeted Akt-STOPS..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6341)
  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 6341)
  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-Lyn-Akt-STOPS
FEATURES             Location/Qualifiers
     source          1..6341
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      28..37
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             73..780
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(218..236)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     486..505
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     promoter        complement(1021..1039)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1022..1039)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1059..1076)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1065..1289
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1335..1763
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1422..1441)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1632..1653
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1772..1792)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1777..2106
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1957..2092
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2019..2038
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2173..2967
                     /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(2227..2246)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2837..2856
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3141..3262
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3178..3197)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3232..3251
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3311..3327)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3311..3327)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3324..3346)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3335..3351
                     /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(3359..3365,3366..3383,3384..3389))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3404..3425
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3542..3559)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3713..4301)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3793..3812)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4472..5263,5264..5332))
                     /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     5095..5114
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5333..5437)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5512..5531)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5703..6082
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6083..6286
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6237..6257
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     join(6331..6341,1..9)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(6331..6341,1..8)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 actataggga gacccaagct tgcggccgcc accatgggct gcatcaagag caagcgcaag
       61 gacaaggatc ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc
      121 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
      181 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt
      241 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc
      301 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
      361 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
      421 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
      481 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac
      541 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc
      601 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
      661 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg
      721 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
      781 cgcatgcatc ctcgtccgcg ctcgtgcacc tggccggacc ccaggccgga gtttggagtc
      841 gaccctcgtc cgcgctcgtg cacctggccg gaccccaggc cggagtttgg aggtaccggc
      901 ggcagcgagc tccctcgtcc gcgctcgtgc acctggccgg accccaggcc ggagttttaa
      961 gaattctgca gatatccatc acactggcgg ccgctcgagc atgcatctag agggccctat
     1021 tctatagtgt cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
     1081 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     1141 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     1201 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     1261 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     1321 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1381 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1441 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1501 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1561 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1621 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     1681 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     1741 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     1801 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     1861 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     1921 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     1981 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     2041 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     2101 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga
     2161 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     2221 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     2281 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     2341 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     2401 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     2461 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     2521 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     2581 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     2641 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     2701 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     2761 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     2821 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     2881 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     2941 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     3001 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     3061 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     3121 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
     3181 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     3241 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     3301 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     3361 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     3421 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     3481 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     3541 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     3601 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     3661 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3721 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3781 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     3841 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3901 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3961 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     4021 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     4081 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     4141 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     4201 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     4261 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     4321 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     4381 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     4441 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     4501 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     4561 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     4621 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     4681 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     4741 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     4801 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     4861 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     4921 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     4981 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     5041 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     5101 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     5161 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     5221 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     5281 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     5341 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     5401 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     5461 ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg
     5521 ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga
     5581 gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa
     5641 gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg
     5701 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     5761 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     5821 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     5881 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     5941 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     6001 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     6061 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
     6121 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     6181 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
     6241 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag
     6301 agaacccact gcttactggc ttatcgaaat taatacgact c
//
