LOCUS       sequence-444170-        7511 bp    DNA     circular SYN 15-APR-2025
DEFINITION  AAV expression of mGrm3 from GfaABC1D promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7511)
  TITLE     Nagai Lab plasmids for expression in astrocytes and neurons
REFERENCE   2  (bases 1 to 7511)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: GfaABC1D-mGrm3-WPRE
FEATURES             Location/Qualifiers
     source          1..7511
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     protein_bind    137..157
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     intron          1008..1140
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1123..1142
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1185..1203
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1219..1243
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3890..3914)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3924..4512
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3977..3997)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4395..4406)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4542..4559)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4548..4755
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4763..4903
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4763..4892
                     /label=AAV2 ITR
     rep_origin      4978..5433
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5065..5084)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5275..5296
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5450..5469)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5569..5591
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5629..5647)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5715..5819
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5820..5888,5889..6680)
                     /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"
     rep_origin      6851..7439
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7340..7359
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   join(7501..7511,1..130)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tctagacaac tttgtataga aaagttgtgt agttaatgat taacccgcca
      181 tgctacttat ctacgtagcc atgctctagg aagatctaac atatcctggt gtggagtagg
      241 ggacgctgct ctgacagagg ctcgggggcc tgagctggct ctgtgagctg gggaggaggc
      301 agacagccag gccttgtctg caagcagacc tggcagcatt gggctggccg ccccccaggg
      361 cctcctcttc atgcccagtg aatgactcac cttggcacag acacaatgtt cggggtgggc
      421 acagtgcctg cttcccgccg caccccagcc cccctcaaat gccttccgag aagcccattg
      481 agcagggggc ttgcattgca ccccagcctg acagcctggc atcttgggat aaaagcagca
      541 cagcccccta ggggctgccc ttgctgtgtg gcgccaccgg cggtggagaa caaggctcta
      601 ttcagcctgt gcccaggaaa ggggatcagg ggatgcccag gcatggacag tgggtggcag
      661 ggggggagag gagggctgtc tgcttcccag aagtccaagg acacaaatgg gtgaggggag
      721 agctctcccc atagctgggc tgcggcccaa ccccaccccc tcaggctatg ccagggggtg
      781 ttgccagggg cacccgggca tcgccagtct agcccactcc ttcataaagc cctcgcatcc
      841 caggagcgag cagagccaga gcaggttgga gaggagacgc atcacctccg ctgctcgcaa
      901 gctttattgc ggtagtttat cacagttaaa ttgctaacgc agtcagtgct tctgacacaa
      961 cagtctcgaa cttaagctgc agaagttggt cgtgaggcac tgggcaggta agtatcaagg
     1021 ttacaagaca ggtttaagga gaccaataga aactgggctt gtcgagacag agaagactct
     1081 tgcgtttctg ataggcacct attggtctta ctgacatcca ctttgccttt ctctccacag
     1141 gtgtccactc ccagttcaat tacagctctt aaggctagag tacttaatac gactcactat
     1201 aggctagcct cgagaattcc aagtttgtac aaaaaagcag gctgccacca tgaagatgtt
     1261 gacaagacta caagttctta tgttagcttt gttttcaaag ggatttttag tctctttagg
     1321 agatcacaac tttatgagga gagaaattaa aatagaagga gaccttgttt tagggggctt
     1381 atttcctatt aatgaaaaag gcactggaac tgaagagtgt ggacgaatca atgaagacag
     1441 aggtatccaa cgcctggagg ccatgttgtt tgccattgat gaaatcaaca aagacaatta
     1501 cttgcttcca ggagtgaagc tgggggttca cattttggat acatgttcaa gagacaccta
     1561 tgcattagag cagtcactgg agtttgtcag agcatcgttg actaaagtgg atgaagctga
     1621 atatatgtgt cctgatggat catatgctat tcaagaaaac atcccactac tcattgcagg
     1681 agtcattggc ggttcgtaca gcagtgtttc catacaggtg gcaaacctgc tgaggctctt
     1741 ccagatccct cagataagct acgcctccac cagtgccaaa ctcagcgaca aatcgcgcta
     1801 tgattatttt gccaggaccg tgccccctga cttctaccag gccaaagcca tggccgagat
     1861 cttgcgctac tttaactgga cctatgtgtc cactgttgcc tctgaaggtg actatgggga
     1921 gacagggatt gaggccttcg agcaggaagc aaggctacgc aacatctgca tcgccactgc
     1981 tgaaaaggtg gggcgctcca acatccgcaa gtcctacgac agcgtgatcc gtgagctcct
     2041 gcagaaacct aacgcgcgag ttgtggtcct gttcatgcgc agtgatgact cacgagagtt
     2101 gatcgctgca gccagccgcg tgaatgcttc cttcacctgg gtggccagcg atggctgggg
     2161 tgcacaggag agcattgtca agggcagtga gcacgtcgcc tatggagcca tcaccctgga
     2221 gctggcgtcc caccctgttc gtcagtttga tcgctacttc cagagcctca acccctacaa
     2281 caatcatcgt aacccctggt tccgagactt ctgggagcag aagttccagt gcagcctcca
     2341 gaacaagaga aaccacagac agatttgtga caagcacctg gccattgaca gcagcaacta
     2401 tgaacaagaa tccaagatca tgtttgtggt gaatgcagtg tatgccatgg cgcatgcgct
     2461 gcacaaaatg caacgcaccc tctgtcccaa caccaccaag ctctgtgatg caatgaagat
     2521 cctggatgga aagaagttgt acaaagatta tttgctgaaa atcaacttca cagctccatt
     2581 caacccaaat aaaggagcag acagcattgt gaagtttgac acttacggag acgggatggg
     2641 aagatacaac gtgttcaact tccagcatat aggtggaaag tattcctact taaaagttgg
     2701 ccactgggca gaaactttat atctagatgt ggactctatt cattggtccc ggaactcagt
     2761 ccccacttcc cagtgcagtg atccctgtgc ccccaatgaa atgaaaaaca tgcagccagg
     2821 agatgtttgc tgctggatct gcatcccatg tgagccctat gaatacctgg ttgatgagtt
     2881 cacctgcatg gattgtggcc ctggccagtg gcccactgca gacctatctg gatgctacaa
     2941 ccttccagag gattacatca ggtgggaaga tgcctgggca ataggcccag tcactattgc
     3001 ctgcctgggt tttatgtgta catgcatagt cataactgtt tttatcaagc acaacaacac
     3061 acccttggtc aaagcatcag gccgagaact ctgctacatc ttgttatttg gagttagcct
     3121 gtcctattgc atgacattct tcttcattgc taagccatcg cctgtcatct gtgcattgcg
     3181 ccgacttggg cttgggacct cctttgccat ctgttattca gctctcctga ccaagacaaa
     3241 ctgcatcgct cgcatctttg atggggtcaa gaatggcgct cagaggccaa aattcatcag
     3301 ccccagttct caggttttta tctgcctggg tttgatactg gtgcaaattg tgatggtgtc
     3361 tgtgtggctt atcttggaga ctccaggtac tagaagatac accctgccag agaagcggga
     3421 aacagtcatc ctaaaatgca atgtcaaaga ttccagcatg ttgatctctc tgacctatga
     3481 cgtggttctg gtgattctat gcactgtgta tgccttcaaa acaaggaagt gtcctgaaaa
     3541 cttcaatgaa gccaagttca taggcttcac catgtacacc acctgcatca tctggttggc
     3601 attcctccct atattttatg tgacatcaag tgactacaga gtgcagacga caacaatgtg
     3661 catctccgtt agcttgagtg gtttcgtggt cttgggctgt ttgtttgccc ccaaggtgca
     3721 cattgtcctg ttccaacccc agaagaatgt ggtcacacac agacttcacc tcaacaggtt
     3781 cagtgtcagt ggaactgcga ccacatattc tcagtcctct gcaagcacat atgtcccgac
     3841 agtgtgcaac ggaagggaag tcctggactc caccacctca tctctgtaga cccagctttc
     3901 ttgtacaaag tgggaattcc gataatcaac ctctggatta caaaatttgt gaaagattga
     3961 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     4021 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     4081 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg
     4141 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg
     4201 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc
     4261 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc
     4321 tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct
     4381 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg
     4441 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg
     4501 ccgcctcccc gcatcgggaa ttcctagagc tcgctgatca gcctcgactg tgccttctag
     4561 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     4621 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     4681 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagagaatag
     4741 caggcatgct ggggagggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     4801 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4861 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4921 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4981 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     5041 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     5101 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     5161 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     5221 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     5281 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     5341 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5401 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     5461 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     5521 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     5581 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     5641 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     5701 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     5761 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5821 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5881 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5941 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     6001 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     6061 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     6121 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     6181 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     6241 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     6301 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     6361 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     6421 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6481 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtggaagcc
     6541 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6601 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6661 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     6721 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6781 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6841 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6901 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6961 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     7021 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     7081 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     7141 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     7201 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     7261 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     7321 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     7381 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     7441 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     7501 cctgcaggca g
//
