LOCUS       sequence-457470-        7797 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Microglia-targeted transgene expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7797)
  AUTHORS   Okada Y, Hosoi N, Matsuzaki Y, Fukai Y, Hiraga A, Nakai J, Nitta K, 
            Shinohara Y, Konno A, Hirai H
  TITLE     Development of microglia-targeting adeno-associated viral vectors as
            tools to study microglial behavior in vivo.
  JOURNAL   Commun Biol. 2022 Nov 11;5(1):1224. doi: 10.1038/s42003-022-04200-3.
  PUBMED    36369525
REFERENCE   2  (bases 1 to 7797)
  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: "Commun 
            Biol."; date: "2022-11-11"; volume: "5(1):1224. doi"; pages: " 
            10.1038/s42003-022-04200-3"
COMMENT     Sequence Label: pAAV/mIba1.GFP.WPRE.miR-9.T.miR-129-2-3p.T.SV40pA
FEATURES             Location/Qualifiers
     source          1..7797
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(434..1022)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(514..533)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1193..1984,1985..2053))
                     /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     1816..1835
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2054..2158)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2226..2244
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2282..2304)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2404..2423
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(2905..3418)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3100..3121)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3312..3331
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   3789..3918
                     /label=AAV2 ITR
     primer_bind     complement(4014..4033)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4068..4087
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    4382..4970
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             4488..4499
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     4897..4917
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(4986..5699,5700..5702,5703..5705))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(4989..5699,5700..5702,5703..5705))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5031..5052)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5380..5399
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5639..5660
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      5702..5711
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 kttttccata ggctccsccs ysywkrsraa ttaaaggaga gcttgtagga gctggatcca
       61 accagttccc cagttaacag ctcttcatgg gagatttcct ccagcagcaa cctcagttcc
      121 tggtggaaac gaggttttca cgatgcatcc tatagtactc gagtctagag catggctacg
      181 tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg atggagttgg
      241 ccactccctc tctgcgcgct cgctcgctca ctgactcgct gcgctcggtc gttcggctgc
      301 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
      361 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
      421 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
      481 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
      541 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
      601 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
      661 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
      721 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
      781 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
      841 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
      901 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
      961 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     1021 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     1081 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     1141 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     1201 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1261 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     1321 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     1381 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1441 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1501 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     1561 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     1621 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     1681 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     1741 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     1801 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     1861 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     1921 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     1981 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     2041 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     2101 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     2161 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     2221 ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa
     2281 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     2341 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact
     2401 atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca
     2461 gatgcgtaag gagaaaatac cgcatcaggc gattccaaca tccaataaat catacaggca
     2521 aggcaaagaa ttagcaaaat taagcaataa agcctcagag cataaagcta aatcggttgt
     2581 accaaaaaca ttatgaccct gtaatacttt tgcgggagaa gcctttattt caacgcaagg
     2641 ataaaaattt ttagaaccct catatatttt aaatgcaatg cctgagtaat gtgtaggtaa
     2701 agattcaaac gggtgagaaa ggccggagac agtcaaatca ccatcaatat gatattcaac
     2761 cgttctagct gataaattca tgccggagag ggtagctatt tttgagaggt ctctacaaag
     2821 gctatcaggt cattgcctga gagtctggag caaacaagag aatcgatgaa cggtaatcgt
     2881 aaaactagca tgtcaatcat atgtaccccg gttgataatc agaaaagccc caaaaacagg
     2941 aagattgtat aagcaaatat ttaaattgta agcgttaata ttttgttaaa attcgcgtta
     3001 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     3061 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     3121 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     3181 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     3241 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     3301 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     3361 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtac
     3421 tatggttgct ttgacgagca cgtataacgt gctttcctcg ttagaatcag agcgggagct
     3481 aaacaggagg ccgattaaag ggattttaga caggaacggt acgccagaat cctgagaagt
     3541 gtttttataa tcagtgaggc caccgagtaa aagagtctgt ccatcacgca aattaaccgt
     3601 tgtcgcaata cttctttgat tagtaataac atcacttgcc tgagtagaag aactcaaact
     3661 atcggccttg ctggtaatat ccagaacaat attaccgcca gccattgcaa cggaatcgcc
     3721 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     3781 tacgccagct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg
     3841 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
     3901 ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc
     3961 catgctctag agctccaacg cgttgggggt cgacagacat gataagatac attgatgagt
     4021 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     4081 ctattgcttt atttgtaacc attataagct gcaataaaca agtgaattcc tcgactctac
     4141 tagaggatcc aagcccttac cccaaaaagc atgtgaaagc ccttacccca aaaagcatct
     4201 taagaagccc ttaccccaaa aagcatatcg aagcccttac cccaaaaagc atgctagcct
     4261 ttggttatct agctgtatga gtgactttgg ttatctagct gtatgacaat tgctttggtt
     4321 atctagctgt atgaatcgct ttggttatct agctgtatga ggtaccgtat acggcatcga
     4381 tgcggggagg cggcccaaag ggagatccga ctcgtctgag ggcgaaggcg aagacgcgga
     4441 agaggccgca gagccggcag caggccgcgg gaaggaaggt ccgctggatt gagggccgaa
     4501 gggacgtagc agaaggacgt cccgcgcaga atccaggtgg caacacaggc gagcagccat
     4561 ggaaaggacg tcagcttccc cgacaacacc acggaattgt cagtgcccaa cagccgagcc
     4621 cctgtccagc agcgggcaag gcaggcggcg atgagttccg ccgtggcaat agggaggggg
     4681 aaagcgaaag tcccggaaag gagctgacag gtggtggcaa tgccccaacc agtgggggtt
     4741 gcgtcagcaa acacagtgca caccacgcca cgttgcctga caacgggcca caactcctca
     4801 taaagagaca gcaaccagga tttatacaag gaggagaaaa tgaaagccat acgggaagca
     4861 atagcatgat acaaaggcat taaagcagcg tatccacata gcgtaaaagg agcaacatag
     4921 ttaagaatac cagtcaatct ttcacaaatt ttgtaatcca gaggttgatt atcgatgcgg
     4981 ccgctttact tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc
     5041 agcaggacca tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga ctgggtgctc
     5101 aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag
     5161 tggtcggcga gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg
     5221 atgccgttct tctgcttgtc ggccatgata tagacgttgt ggctgttgta gttgtactcc
     5281 agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg
     5341 ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc
     5401 ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca tggcggactt gaagaagtcg
     5461 tgctgcttca tgtggtcggg gtagcggctg aagcactgca cgccgtaggt cagggtggtc
     5521 acgagggtgg gccagggcac gggcagcttg ccggtggtgc agatgaactt cagggtcagc
     5581 ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg
     5641 tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc
     5701 accatggtgg cgaccggtgg ctcctcagac gctggttgtc ttaggctggc agtctgtccg
     5761 ctgggtcctc tctccaccag ctccccagtc tcttctggga ccgttctcac acttcccctt
     5821 tccactcatt cccttgctaa ctcccaggca tcacttccac atcagcttct gctgacacca
     5881 gcaccccagg agaaggcaga acgggttgag agaggagatg aagaagtaat gtgctgaaaa
     5941 attactttgg gggtggggtg ggggcagggt ctcctatcca ggctcttgag aattgaagta
     6001 gccatcctgc ctcagcctcc cagatggctg gagctacagc cctgtaacac acctagctcc
     6061 acacagctcc tccaaagaag cagactgtac gtgggagaaa gacttagatc agctcccgga
     6121 ggtgtggaaa ggcaccagga tctgagggac cgagcgtgat accctaggcc tgcaatctag
     6181 cacctaagag aggctgaggc tagaggatcc tgggagttgg aacccagcca ggccaacaca
     6241 gtaagaacct gtctcaaacc aaggatgcgg tagacagtgg ctttccccag atcctgaggt
     6301 ggggcttggg agggtttact tgaaatctcc tcaggcttca tgatgcaggt taagacaggc
     6361 agctgaggag gactggctga ctttccccgg gatggaagag agaggatgaa gtctgcaggc
     6421 tgctccagga atggggtgag ctgagcctag cgccctggaa aggaaagaaa cctctctcag
     6481 gggctacaga ggagcactgg ggacacatgg agacatgtcc cagaaggact tgggtttgtg
     6541 ttcgtcaggc tcttgagtgc cagcctcctg cctcacctgc tttgagatct taggctgaat
     6601 ctacaaagtc atttgagagt ttgaagagac actttggagg aacagaaagt gagactgaaa
     6661 gagacagtag acagagctac ttcctgtccc caccatgggc tgtcaggggt ctaactcatg
     6721 caaatgccct cccctcatac cagggatagc cttggtcccc tcagtcactg ggtcaccagg
     6781 ggagggaggg ctgtgactcc aatgcactca ggccccttcc cttgtgcccc ccgccccagg
     6841 ctatgtcccc ttgtatggca ggcagtatct gtccactctg cagaagactg cagcctctcg
     6901 cctgcctttg aggcagccac cctctggccc tgtgagaggc tctcagtgtc tggtggggac
     6961 agttctctag ccacattcat gtgataaatg atgagcgaag gtttcctgtt tgcagggtac
     7021 actgctaggc ccagggagcc aacagatgaa aagtctgctc ccgtggtctc cattcagagc
     7081 tctgtactta gagggcaggt ttgaatagca tgattatgga agatggccaa gaaattgctc
     7141 tcagggatgc aggagatatg tgggtagaaa ggagaaagta tgctgttttt gttttaattt
     7201 tcatttcatt tctactgttt tatttgtact gtgtgtgcag gtgttcacag cttggcatgt
     7261 gtatggaaat taaaggagag cttgtaggag ctggatccaa ccagttcccc agttaacagc
     7321 tcttcatggg agatttcctc cagcagcaac ctcagttcct ggtggaaacg aggttttcac
     7381 gatgcatcct atagtactcg agtctagagc atggctacgt agataagtag catggcgggt
     7441 taatcattaa ctacaaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
     7501 gctcgctcac tgaggccgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg
     7561 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     7621 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     7681 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     7741 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcg
//
