LOCUS       sequence-317686-        6725 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ExRai-AKAR2 targeted to the outer mitochondrial membrane..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6725)
  AUTHORS   Zhang JF, Liu B, Hong I, Mo A, Roth RH, Tenner B, Lin W, Zhang JZ, 
            Molina RS, Drobizhev M, Hughes TE, Tian L, Huganir RL, Mehta S, 
            Zhang J
  TITLE     An ultrasensitive biosensor for high-resolution kinase activity 
            imaging in awake mice.
  JOURNAL   Nat Chem Biol. 2020 Sep 28. pii: 10.1038/s41589-020-00660-y. doi: 
            10.1038/s41589-020-00660-y.
  PUBMED    32989297
REFERENCE   2  (bases 1 to 6725)
  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 Chem 
            Biol. 2020 Sep 28. pii: 10.1038/s41589-020-00660-y. doi: 
            10.1038/s41589-020-00660-y."
COMMENT     Sequence Label: pcDNA3-mitoExRai-AKAR2
FEATURES             Location/Qualifiers
     source          1..6725
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(50..69)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        241..620
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        621..824
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     775..795
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     869..888
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        869..887
                     /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     1272..1293
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1399..1420)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1660..1679)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     promoter        complement(2284..2302)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2285..2302)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2322..2339)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2328..2552
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2598..3026
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2685..2704)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2895..2916
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3035..3055)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3040..3369
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3220..3355
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3282..3301
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3436..4230
                     /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(3490..3509)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4100..4119
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4404..4525
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4441..4460)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4495..4514
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4574..4590)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4574..4590)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4587..4609)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4598..4614
                     /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(4622..4628,4629..4646,4647..4652))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4667..4688
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4805..4822)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4976..5564)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5056..5075)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5735..6526,6527..6595))
                     /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     6358..6377
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6596..6700)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct
       61 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt
      121 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
      181 atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt
      241 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
      301 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      361 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      421 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      481 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      541 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      601 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      661 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      721 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      781 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      841 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      901 gccaccatgg caatccagtt gcgttcgctc ttccccttgg cgttgcccgg aatgctggcc
      961 ctccttggct ggtggtggtt tttctctcgt aaaaaagcgg atcccatgct gcgtcgcgcc
     1021 accctggtgg acggcggcac cggcggcagc gagctcagct tctgcaacgt ctatatcaag
     1081 gccgacaagc agaagaacgg catcaaggcg aacttcaaga tccgccacaa catcgaggac
     1141 ggcggcgtgc agctcgccta ccactaccag cagaacaccc ccatcggcga cggccccgtg
     1201 ctgctgcccg acaaccacta cctgagcgtg cagtccaaac tttcgaaaga ccccaacgag
     1261 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
     1321 gacgagctgt acaagggcgg taccggaggg agcatggtga gcaagggcga ggagctgttc
     1381 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     1441 gtgtccggcg agggtgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     1501 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     1561 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     1621 cccgaaggct acatccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     1681 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     1741 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaactt gctgcataag
     1801 ttttctcaag aacagatcgg cgaaaacatt gtgtgcaggg tcatttgtac cacgggtcaa
     1861 attcccatcc gagatttgtc agctgatatt tcacaagtgc ttaaggaaaa acgatccata
     1921 aagaaagttt ggacatttgg tagaaaccca gcctgtgact atcatttagg aaacatttca
     1981 agactgtcaa ataagcattt ccaaatacta ctaggagaag acggtaacct tttattgaat
     2041 gacatttcca ctaatgggac ctggttaaat gggcaaaaag tcgagaagaa cagcaatcag
     2101 ttactgtctc aaggtgatga aataaccgtt ggtgtaggcg tggaatcaga tattttatct
     2161 ctggtcattt tcataaacga caaatttaag cagtgcctcg agcagaacaa agttgatcgc
     2221 taagaattct gcagatatcc atcacactgg cggccgctcg agcatgcatc tagagggccc
     2281 tattctatag tgtcacctaa atgctagagc tcgctgatca gcctcgactg tgccttctag
     2341 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     2401 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     2461 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     2521 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     2581 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     2641 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     2701 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     2761 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     2821 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     2881 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     2941 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     3001 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     3061 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3121 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     3181 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     3241 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     3301 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     3361 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     3421 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     3481 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     3541 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     3601 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     3661 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     3721 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     3781 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     3841 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     3901 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     3961 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     4021 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     4081 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     4141 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     4201 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     4261 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     4321 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     4381 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     4441 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     4501 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     4561 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4621 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4681 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4741 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     4801 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4861 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4921 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4981 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5041 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5101 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5161 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5221 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5281 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5341 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5401 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5461 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5521 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5581 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5641 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5701 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5761 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5821 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5881 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5941 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6001 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6061 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6121 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6181 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6241 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6301 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6361 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6421 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6481 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6541 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6601 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6661 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6721 cacct
//
