LOCUS       sequence-393620-        7231 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses tKiMBI and caMEK in an AAV vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7231)
  AUTHORS   Wu Y, Walker JR, Westberg M, Ning L, Monje M, Kirkland TA, Lin MZ, 
            Su Y
  TITLE     Kinase-Modulated Bioluminescent Indicators Enable Noninvasive 
            Imaging of Drug Activity in the Brain.
  JOURNAL   ACS Cent Sci. 2023 Mar 20;9(4):719-732. doi: 
            10.1021/acscentsci.3c00074. eCollection 2023 Apr 26.
  PUBMED    37122464
REFERENCE   2  (bases 1 to 7231)
  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.3c00074"; journalName: "ACS Cent Sci"; date: 
            "2023-03-20- 20"; volume: "9"; issue: "4"; pages: "719-732"
COMMENT     Sequence Label: pAAV_tKiMBI-T2A-caMEK
FEATURES             Location/Qualifiers
     source          1..7231
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(1..69,70..861)
                     /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     complement(219..238)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1032..1620
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1521..1540
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1682..1811
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        1827..2206
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2207..2410
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2361..2381
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2455..2474
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2455..2473
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2509..2518
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4636..4689
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     complement(5922..5939)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5928..6152
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6175..6315
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6175..6304
                     /label=AAV2 ITR
     rep_origin      6390..6845
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6477..6496)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6687..6708
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6862..6881)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6981..7003
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7041..7059)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7127..7231
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
       61 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
      121 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
      181 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
      241 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
      301 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
      361 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
      421 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
      481 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
      541 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
      601 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
      661 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
      721 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
      781 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
      841 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
      901 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
      961 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     1021 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     1081 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     1141 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     1201 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     1261 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     1321 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     1381 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     1441 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     1501 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     1561 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     1621 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     1681 tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt
     1741 tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac
     1801 taggggttcc tgcggccgca cgcgttgaca ttgattattg actagttatt aatagtaatc
     1861 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1921 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1981 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     2041 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2101 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2161 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2221 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2281 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2341 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2401 aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac
     2461 gactcactat agggagaccc aagctggcta gcgtttaaac ttaagcttgc caccatggtg
     2521 agcaagggcg aggaggtcat caaagagttc atgcgcttca aggtgcgcat ggagggctcc
     2581 atgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc
     2641 cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg
     2701 tccccccagt tcatgtacgg ctccaaggcg tacgtgaagc accccgccga catccccgat
     2761 tacaagaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac
     2821 ggcggtctgg tgaccgtgac ccaggactcc tccctgcagg acggcacgct gatctacaag
     2881 gtgaagatgc gcggcaccaa cttccccccc gacggccccg taatgcagaa gaagaccatg
     2941 ggctgggagg cctccaccga gcgcctgtac ccccgcgacg gcgtgctgaa gggcgagatc
     3001 caccaggccc tgaagctgaa ggacggcggc cactacctgg tggagttcaa gaccatctac
     3061 atggccaaga agcccgtgca actgcccggc tactactacg tggacaccaa gctggacatc
     3121 acctcccaca acgaggacta caccatcgtg gaacagtacg agcgctccga gggccgccac
     3181 cacctgttcc tggggcatgg caccggcagc accggcagcg gcagctccgg caccgcctcc
     3241 tccgaggaca acaacatggc cgtcatcaaa gagttcatgc gcttcaaggt gcgcatggag
     3301 ggctccatga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag
     3361 ggcacccaga ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac
     3421 atcctgtccc cccagttcat gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc
     3481 cccgattaca agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc
     3541 gaggacggcg gtctggtgac cgtgacccag gactcctccc tgcaggacgg cacgctgatc
     3601 tacaaggtga agatgcgcgg caccaacttc ccccccgacg gccccgtaat gcagaagaag
     3661 accatgggct gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc
     3721 gagatccacc aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagacc
     3781 atctacatgg ccaagaagcc cgtgcaactg cccggctact actacgtgga caccaagctg
     3841 gacatcacct cccacaacga ggactacacc atcgtggaac agtacgagcg ctccgagggc
     3901 cgccaccacc tgttccggct ggaagatttc gttggggact gggaacagac agccgcctac
     3961 aacctggacc aagtccttga acagggaggt gtgtccagtt tgctgcagaa tctcgccgtg
     4021 tccgtaactc cgatccaaag gattgtccgg agcggtgaaa atgccctgaa gatcgacatc
     4081 catgtcatca tcccgtatga aggtctgagc gccgaccaaa tggcccagat cgaagaggtg
     4141 tttaaggtgg tgtaccctgt ggatgatcat cactttaagg tgatcctgcc ctatggcaca
     4201 ctggtaatcg acggggttac gccgaacatg ctgaactatt tcggacggcc gtatgaaggc
     4261 atcgccgtgt tcgacggcaa aaagatcact gtaacaggga ccctgtggaa cggcaacaaa
     4321 attatcgacg agcgcctgat cacccccgac ggctccatgc tgttccgagt aaccatcaac
     4381 agcatcatgg cggacgagga gaagctgccg cccggctggg agaagcgcat gagccgcagc
     4441 tcaggccgag tgtactactt caaccacatc actaacgcca gccagtggga gcggcccagc
     4501 ggcaacagca gcagtggtgg caaaaacggg cagggggagc ctgccagggt gaccggctac
     4561 cggctgttcg agccagatag ccctagaact ccagtggata aagcaaagct gtcattccaa
     4621 ttccccggat ccggtgaagg tcgaggctcc ctcctcactt gtggggatgt ggaagaaaac
     4681 ccagggcctg gctcaggtat gcccaagaag aagccgacgc ccatccagct gaacccgacc
     4741 cccgatggct cggcggttaa cgggaccagc tcggccgaga ccaacctgga ggccttgcag
     4801 aagaagctgg aggggctgga gcttgatgag cagcagcgga agcgccttga ggcctttctg
     4861 acccagaagc agaaggtggg agaactgaag gatgatgact ttgagaagat cagtgagctg
     4921 ggggccggca acggtggagt ggtattcaag gtctcccata agccgtctgg cctagttatg
     4981 gccaggaagc taattcatct ggagatcaaa cccgcaatcc ggaaccagat catccgggag
     5041 ctgcaggtgc tgcatgagtg caactcccca tacatcgtgg gcttctatgg ggctttctac
     5101 agtgacggcg agatcagcat ctgcatggaa cacatggatg gtgggtcctt ggatcaagtt
     5161 ctgaagaaag ctggaagaat tcctgagcaa attttaggaa aagttagcat tgctgtgata
     5221 aaaggcctga catatcttcg ggagaagcac aagattatgc acagagatgt caagccttcc
     5281 aatattctag taaactcccg tggagaaatc aaactctgtg attttggggt cagcgggcaa
     5341 ctaattgacg acatggccaa cgactttgta ggcacaagat catacatgtc gcccgagaga
     5401 ctccagggga ctcactactc cgtgcagtcg gacatctgga gcatggggct ctctctggtg
     5461 gagatggcag ttgggagata ccccatccct cctcctgatg ccaaggaact ggagctgctg
     5521 tttggatgcc aggtggaggg agatgcagcc gaaacgccac ccaggccgag gacccctggg
     5581 aggcccctta gctcatatgg aatggacagc cgacctccca tggcaatttt tgagttgttg
     5641 gattatatag tcaatgagcc tcctccaaaa ttgcccagtg gagtattcag tctggagttt
     5701 caagattttg tgaataaatg cttaataaag aaccctgcag agagagcaga tctaaagcag
     5761 cttctggttc acgctttcat caagagatct gatgccgagg aagtagactt cgcaggttgg
     5821 ctctgctcca ccattgggct taaccagccc agcacaccaa cccatgccgc tagcatctaa
     5881 tgaattctgc agatatccag cacagtggcg gccgctcgag gcctcgactg tgccttctag
     5941 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     6001 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     6061 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     6121 caggcatgct ggggatgcgg tgggctctat ggcacgtgcg gaccgagcgg ccgcaggaac
     6181 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     6241 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     6301 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     6361 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     6421 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
     6481 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     6541 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     6601 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     6661 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     6721 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     6781 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     6841 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     6901 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     6961 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     7021 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     7081 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     7141 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     7201 aatgcttcaa taatattgaa aaaggaagag t
//
