LOCUS       sequence-442752-        6296 bp    DNA     circular SYN 15-APR-2025
DEFINITION  AAV expression of flag-mAdrb1 from GfaABC1D promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6296)
  TITLE     Nagai Lab plasmids for expression in astrocytes and neurons
REFERENCE   2  (bases 1 to 6296)
  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-flag-mAdrb1-WPRE
FEATURES             Location/Qualifiers
     source          1..6296
                     /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"
     regulatory      1244..1253
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1253..1276
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    complement(2675..2699)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2709..3297
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2762..2782)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3180..3191)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3327..3344)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3333..3540
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3548..3688
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3548..3677
                     /label=AAV2 ITR
     rep_origin      3763..4218
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3850..3869)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4060..4081
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4235..4254)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4354..4376
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4414..4432)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4500..4604
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4605..4673,4674..5465)
                     /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      5636..6224
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6125..6144
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   join(6286..6296,1..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 tggactacaa
     1261 ggacgacgat gacaagggcg cgggggcgct cgccctgggc gcctccgaac cctgcaacct
     1321 gtcgtccgcc gcgccgctgc ccgacggtgc ggccaccgcg gcgcggctgc tggtgctcgc
     1381 gtcgcctccc gcctcgctgc tgcctccagc cagcgagggc tcagcgccgc tgtcgcagca
     1441 gtggaccgcg ggtatgggcc tactgctggc gctcatcgtt ctgctcatcg tggtgggtaa
     1501 cgtgctggtg atcgtggcca tcgccaagac cccgcggctg cagacgctca ccaacctctt
     1561 catcatgtcc ctggccagcg ctgatctggt catgggattg ctggtggtgc ctttcggggc
     1621 caccatcgtg gtgtggggcc gctgggagta cggctccttc ttctgcgagc tctggacttc
     1681 ggtagatgtg ctgtgtgtga cggccagcat tgagaccctg tgtgtcatcg ccctggaccg
     1741 ctacctcgcc atcacgtcgc cctttcgcta ccagagtttg ctgacgcgcg cgcgagcgcg
     1801 ggccctcgtg tgcacagtgt gggccatctc ggcgttggtg tccttcctgc ccatcctcat
     1861 gcactggtgg cgggccgaga gcgacgaagc gcgccgctgc tacaacgacc ccaagtgctg
     1921 cgatttcgtc accaacaggg cctacgccat cgcctcgtcc gtcgtctcct tctacgtgcc
     1981 cctgtgcatc atggccttcg tgtacctgcg ggtgttccgc gaggcccaaa aacaggtaaa
     2041 gaagatcgac agctgcgagc gccgcttcct cggcggccca gcccggccgc cctcgcctga
     2101 gccctcgccg tcacctgggc caccgcgccc cgcagactcg ctggccaacg ggcgctccag
     2161 caagcggcgg ccgtcgcgcc tcgtggctct gcgcgagcag aaggcgctca agacactggg
     2221 catcatcatg ggtgtgttca cgctctgctg gctgcccttc ttcctggcca acgtggtgaa
     2281 ggctttccac cgcgacctgg tgccggatcg cctcttcgtc ttcttcaact ggctgggcta
     2341 cgccaactcg gccttcaacc ccatcatcta ctgccgcagc cccgacttcc gcaaggcttt
     2401 ccagcgcctg ctctgctgcg cgcgccgggc cgcctgcaga cgccgcgcag cccacgggga
     2461 ccggccgcgc gcctccggct gcctggcgag agctgggccg ccgccgtccc ccggagctcc
     2521 ctcggacgac gacgacgacg acgccgggac caccccaccg gcgcgcctgc tggagccctg
     2581 gaccggctgc aacggcggga caaccactgt ggacagcgat tcgagcctgg acgagccggg
     2641 gcgccagggc ttctcctcgg agtccaaggt gtagacccag ctttcttgta caaagtggga
     2701 attccgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     2761 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     2821 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     2881 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     2941 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     3001 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     3061 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaagctgacg tcctttccat
     3121 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     3181 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     3241 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
     3301 gggaattcct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     3361 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     3421 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     3481 tggggtgggg caggacagca agggggagga ttgggaagag aatagcaggc atgctgggga
     3541 gggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     3601 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     3661 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     3721 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     3781 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     3841 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     3901 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     3961 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     4021 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     4081 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     4141 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     4201 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     4261 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4321 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4381 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     4441 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     4501 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     4561 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     4621 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     4681 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     4741 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     4801 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     4861 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     4921 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     4981 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     5041 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     5101 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5161 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     5221 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5281 gctggtttat tgctgataaa tctggagccg gtgagcgtgg aagccgcggt atcattgcag
     5341 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5401 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5461 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     5521 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     5581 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     5641 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     5701 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     5761 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     5821 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     5881 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     5941 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     6001 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     6061 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     6121 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     6181 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     6241 cctttttacg gttcctggcc ttttgctggc cttttgctca catgtcctgc aggcag
//
