LOCUS       sequence-363291-        6695 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasma membrane-targeted ExRai-AktAR2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6695)
  AUTHORS   Chen M, Sun T, Zhong Y, Zhou X, Zhang J
  TITLE     A Highly Sensitive Fluorescent Akt Biosensor Reveals 
            Lysosome-Selective Regulation of Lipid Second Messengers and Kinase 
            Activity.
  JOURNAL   ACS Cent Sci. 2021 Dec 22;7(12):2009-2020. doi: 
            10.1021/acscentsci.1c00919. Epub 2021 Dec 3.
  PUBMED    34963894
REFERENCE   2  (bases 1 to 6695)
  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: 
            "10.1021/acscentsci.1c00919"; journalName: "ACS Cent Sci"; date: 
            "2021-12-22- 22"; volume: "7"; issue: "12"; pages: "2009-2020"
COMMENT     Sequence Label: pcDNA3-PM-ExRai-AktAR2
FEATURES             Location/Qualifiers
     source          1..6695
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1236..1257
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1363..1384)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1624..1643)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     promoter        complement(2248..2266)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2249..2266)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2286..2303)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2292..2516
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2562..2990
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2649..2668)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2859..2880
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2999..3019)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3004..3333
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3184..3319
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3246..3265
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3400..4194
                     /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(3454..3473)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4064..4083
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4368..4489
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4405..4424)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4459..4478
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4538..4554)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4538..4554)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4551..4573)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4562..4578
                     /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(4586..4592,4593..4610,4611..4616))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4631..4652
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4769..4786)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4940..5528)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5020..5039)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5699..6490,6491..6559))
                     /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     6322..6341
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6560..6664)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgg gctgcatcaa gagcaagcgc aaggacaagg atcccgtcga ccctcgtccg
      961 cgctcgtgca cctggccgga ccccaggccg gagtttggag gtaccggcgg cagcgagctc
     1021 agcttgcaga acgtctatat caaggccgac aagcagaaga acggcatcaa ggcgaacttc
     1081 aagatccgcc acaacatcga ggacggcggc gtgcagctcg cctaccacta ccagcagaac
     1141 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cgtgcagtcc
     1201 aaactttcga aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     1261 gccgccggga tcactctcgg catggacgag ctgtacaagg gcggtaccgg agggagcatg
     1321 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1381 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggtg agggcgatgc cacctacggc
     1441 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1501 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     1561 cacgacttct tcaagtccgc catgcccgaa ggctacatcc aggagcgcac catcttcttc
     1621 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1681 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1741 ctggagtaca acttgctgca taagttttct caagaacaga tcggcgaaaa cattgtgtgc
     1801 agggtcattt gtaccacggg tcaaattccc atccgagatt tgtcagctga tatttcacaa
     1861 gtgcttaagg aaaaacgatc cataaagaaa gtttggacat ttggtagaaa cccagcctgt
     1921 gactatcatt taggaaacat ttcaagactg tcaaataagc atttccaaat actactagga
     1981 gaagacggta accttttatt gaatgacatt tccactaatg ggacctggtt aaatgggcaa
     2041 aaagtcgaga agaacagcaa tcagttactg tctcaaggtg atgaaataac cgttggtgta
     2101 ggcgtggaat cagatatttt atctctggtc attttcataa acgacaaatt taagcagtgc
     2161 ctcgagcaga acaaagttga tcgctaagaa ttctgcagat atccatcaca ctggcggccg
     2221 ctcgagcatg catctagagg gccctattct atagtgtcac ctaaatgcta gagctcgctg
     2281 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     2341 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     2401 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     2461 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     2521 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     2581 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     2641 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     2701 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     2761 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     2821 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     2881 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     2941 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     3001 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3061 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3121 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3181 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3241 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     3301 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     3361 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     3421 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     3481 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     3541 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     3601 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     3661 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     3721 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     3781 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     3841 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     3901 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     3961 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     4021 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     4081 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     4141 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     4201 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     4261 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     4321 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     4381 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4441 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     4501 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     4561 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     4621 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     4681 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     4741 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     4801 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     4861 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4921 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     4981 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5041 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5101 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5161 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5221 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5281 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5341 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5401 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5461 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5521 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5581 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5641 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     5701 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     5761 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     5821 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5881 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     5941 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6001 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6061 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6121 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6181 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6241 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6301 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6361 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6421 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     6481 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     6541 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6601 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     6661 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
