LOCUS       sequence-304894-        7366 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express the medium-affinity adenosine sensor GRAB_Ado1.0med in 
            astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7366)
  AUTHORS   Wu Z, Cui Y, Wang H, Wu H, Wan Y, Li B, Wang L, Pan S, Peng W, Dong 
            A, Yuan Z, Jing M, Xu M, Luo M, Li Y
  TITLE     Neuronal activity-induced, equilibrative nucleoside 
            transporter-dependent, somatodendritic adenosine release revealed by
            a GRAB sensor.
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Apr 4;120(14):e2212387120. doi: 
            10.1073/pnas.2212387120. Epub 2023 Mar 30.
  PUBMED    36996110
REFERENCE   2  (bases 1 to 7366)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2023-04-4"; volume: "120(14):e2212387120. 
            doi"; pages: " 10.1073/pnas.2212387120. Epub 2023 Mar 30"
COMMENT     Sequence Label: pAAV-GfaABC1D-GRAB_Ado1.0med
FEATURES             Location/Qualifiers
     source          1..7366
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          838..970
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     953..972
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1015..1033
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1050..1059
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     1056..1115
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     protein_bind    1119..1143
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     2198..2219
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2325..2346)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2586..2605)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    3336..3924
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3389..3409)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3807..3818)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3956..4432
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4155..4174)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4472..4612
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4472..4601
                     /label=AAV2 ITR
     rep_origin      4687..5142
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4774..4793)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4984..5005
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5159..5178)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5278..5300
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5338..5356)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5424..5528
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5529..5597,5598..6389)
                     /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      6560..7148
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7049..7068
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   7210..7339
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 taacccgcca tgctacttat ctacgtagcc atgctctagg aagatctaac atatcctggt
       61 gtggagtagg ggacgctgct ctgacagagg ctcgggggcc tgagctggct ctgtgagctg
      121 gggaggaggc agacagccag gccttgtctg caagcagacc tggcagcatt gggctggccg
      181 ccccccaggg cctcctcttc atgcccagtg aatgactcac cttggcacag acacaatgtt
      241 cggggtgggc acagtgcctg cttcccgccg caccccagcc cccctcaaat gccttccgag
      301 aagcccattg agcagggggc ttgcattgca ccccagcctg acagcctggc atcttgggat
      361 aaaagcagca cagcccccta ggggctgccc ttgctgtgtg gcgccaccgg cggtggagaa
      421 caaggctcta ttcagcctgt gcccaggaaa ggggatcagg ggatgcccag gcatggacag
      481 tgggtggcag ggggggagag gagggctgtc tgcttcccag aagtccaagg acacaaatgg
      541 gtgaggggag agctctcccc atagctgggc tgcggcccaa ccccaccccc tcaggctatg
      601 ccagggggtg ttgccagggg cacccgggca tcgccagtct agcccactcc ttcataaagc
      661 cctcgcatcc caggagcgag cagagccaga gcaggttgga gaggagacgc atcacctccg
      721 ctgctcgcaa gctttattgc ggtagtttat cacagttaaa ttgctaacgc agtcagtgct
      781 tctgacacaa cagtctcgaa cttaagctgc agaagttggt cgtgaggcac tgggcaggta
      841 agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggctt gtcgagacag
      901 agaagactct tgcgtttctg ataggcacct attggtctta ctgacatcca ctttgccttt
      961 ctctccacag gtgtccactc ccagttcaat tacagctctt aaggctagag tacttaatac
     1021 gactcactat aggctagcct cgagaattcg ccaccatgga gacagacaca ctcctgctat
     1081 gggtactgct gctctgggtt ccaggttcca ctggtgacac aagtttgtac aaaaaagttg
     1141 gcaccggtcc catcatgggc tcctcggtgt acatcacggt ggagctggcc attgctgtgc
     1201 tggccatcct gggcaatgtg ctggtgtgct gggccgtgtg gctcaacagc aacctgcaga
     1261 acgtcaccaa ctactttgtg gtgtcactgg cggcggccga catcgcagtg ggtgtgctcg
     1321 ccatcccctt tgccatcacc atcagcaccg ggttctgcgc tgcctgccac ggctgcctct
     1381 tcattgcctg cttcgtcctg gtcctcacgc agagctccat cttcagtctc ctggccatcg
     1441 ccattgaccg ctacattgcc atccgcatcc cgctccggta caatggcttg gtgaccggca
     1501 cgagggctaa gggcatcatt gccatctgct gggtgctgtc gtttgccatc ggcctgactc
     1561 ccatgctagg ttggaacaac tgcggtcagc caaaggaggg caagaaccac tcccagggct
     1621 gcggggaggg ccaagtggcc tgtctctttg aggatgtggt ccccatgaac tacatggtgt
     1681 acttcaactt ctttgcctgt gtgctggtgc ccctgctgct catgctgggt gtctatttgc
     1741 ggatcttcct ggcggcgcga cgacagctga agcagatgga gagccagcct cccagccgcc
     1801 ggggtccgga cgccgtcgcc gcgccgccgg ggggcaccga gcgcaggccc aacggtctgg
     1861 gccccgagcg cagcgcgggc ccggggggcg cagaggccga accgctgccc acccagctca
     1921 acggcgcccc tggcgagccc gcgccggccg ggccgcgcga caccgacgcg ctggacctgg
     1981 aggagggagg aaacgtctat atcaaggccg acaagcagaa gaacggcatc aaggcgaact
     2041 tccacatccg ccacaacatc gaggacggcg gcgtgcagct cgcctaccac taccagcaga
     2101 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcgtgcagt
     2161 ccaaactttc gaaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     2221 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gggcggtacc ggagggagca
     2281 tggtgagaaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     2341 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg tgagggcgat gccacctacg
     2401 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     2461 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     2521 agcacgactt cttcaagtcc gccatgcccg aaggctacat ccaggagcgc accatcttct
     2581 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     2641 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     2701 agctggagta caacaccgga gcagcagcac gctggcgcgg gcggcagaac cgcgagctcc
     2761 atgctgccaa gtcactggcc atcattgtgg ggctctttgc cctctgctgg ctgcccctac
     2821 acatcatcaa ctgcttcact ttcttctgcc ccgactgcag ccacgcccct ctctggctcc
     2881 actacctggc catcgtcctc tcccacacca attcggttgt gaatcccttc atctacgcct
     2941 accgtatccg cgagttccgc cagaccttcc gcaagatcat tcgcagccac gtcctgaggc
     3001 agcaagaacc tttcaaggca gctggcacca gtgcccgggt cttggcagct catggcagtg
     3061 acggagagca ggtcagcctc cgtctcaacg gccacccgcc aggagtgtgg gccaacggca
     3121 gtgctcccca ccctgagcgg aggcccaatg gctacgccct ggggctggtg agtggaggga
     3181 gtgcccaaga gtcccagggg aacacgggcc tcccagacgt ggagctcctt agccatgagc
     3241 tcaagggagt gtgcccagag ccccctggcc tagatgaccc cctggcccag gatggagcag
     3301 gagtgtcctg agaattcgat atcaagctta tcgataatca acctctggat tacaaaattt
     3361 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     3421 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     3481 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     3541 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     3601 agctcctttc cggaactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     3661 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     3721 tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc
     3781 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     3841 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     3901 tctccctttg ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctacgggtg
     3961 gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc
     4021 caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta
     4081 taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg
     4141 tagggcctgc ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac
     4201 tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg
     4261 attccaggca tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt
     4321 caccatattg gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc
     4381 tcccaaattg ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt
     4441 gtaggtaacc acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact
     4501 ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg
     4561 ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg
     4621 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac
     4681 catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     4741 tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc
     4801 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     4861 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     4921 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     4981 atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg
     5041 atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa
     5101 aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta
     5161 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     5221 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     5281 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     5341 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     5401 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     5461 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     5521 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     5581 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     5641 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     5701 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     5761 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     5821 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     5881 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     5941 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     6001 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     6061 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     6121 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     6181 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     6241 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     6301 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     6361 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     6421 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     6481 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     6541 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     6601 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     6661 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     6721 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     6781 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     6841 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     6901 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     6961 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     7021 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     7081 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     7141 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     7201 ctcacatgtc ctgcaggcag ctgcgcgctc gctcgctcac tgaggccgcc cgggcgtcgg
     7261 gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac
     7321 tccatcacta ggggttcctg cggccgcacg cgttgtagtt aatgat
//
