LOCUS       sequence-301932-        7369 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express the adenosine-insensitive control sensor GRAB_Ado1.0mut in 
            astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7369)
  AUTHORS   Peng W, Wu Z, Song K, Zhang S, Li Y, Xu M
  TITLE     Regulation of sleep homeostasis mediator adenosine by basal 
            forebrain glutamatergic neurons.
  JOURNAL   Science. 2020 Sep 4;369(6508). pii: 369/6508/eabb0556. doi: 
            10.1126/science.abb0556.
  PUBMED    32883833
REFERENCE   2  (bases 1 to 7369)
  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: "Science."; 
            date: "2020-09-4"; volume: "369(6508). pii: 369/6508/eabb0556. doi";
            pages: " 10.1126/science.abb0556"
COMMENT     Sequence Label: pAAV-GfaABC1D-GRAB_Ado1.0mut
FEATURES             Location/Qualifiers
     source          1..7369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          1645..1777
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1760..1779
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1822..1840
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1857..1866
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     1863..1922
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     protein_bind    1926..1950
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     3008..3029
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3135..3156)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3396..3415)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    4146..4734
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4199..4219)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4617..4628)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4766..5242
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4965..4984)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5282..5422
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5282..5411
                     /label=AAV2 ITR
     rep_origin      5497..5952
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5584..5603)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5794..5815
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5969..5988)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6088..6110
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6148..6166)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6234..6338
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6339..6407,6408..7199)
                     /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"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggccgcac gcgttgtagt taatgattaa cccgccatgc tacttatcta cgtagccatg
      841 ctctaggaag atctaacata tcctggtgtg gagtagggga cgctgctctg acagaggctc
      901 gggggcctga gctggctctg tgagctgggg aggaggcaga cagccaggcc ttgtctgcaa
      961 gcagacctgg cagcattggg ctggccgccc cccagggcct cctcttcatg cccagtgaat
     1021 gactcacctt ggcacagaca caatgttcgg ggtgggcaca gtgcctgctt cccgccgcac
     1081 cccagccccc ctcaaatgcc ttccgagaag cccattgagc agggggcttg cattgcaccc
     1141 cagcctgaca gcctggcatc ttgggataaa agcagcacag ccccctaggg gctgcccttg
     1201 ctgtgtggcg ccaccggcgg tggagaacaa ggctctattc agcctgtgcc caggaaaggg
     1261 gatcagggga tgcccaggca tggacagtgg gtggcagggg gggagaggag ggctgtctgc
     1321 ttcccagaag tccaaggaca caaatgggtg aggggagagc tctccccata gctgggctgc
     1381 ggcccaaccc caccccctca ggctatgcca gggggtgttg ccaggggcac ccgggcatcg
     1441 ccagtctagc ccactccttc ataaagccct cgcatcccag gagcgagcag agccagagca
     1501 ggttggagag gagacgcatc acctccgctg ctcgcaagct ttattgcggt agtttatcac
     1561 agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt aagctgcaga
     1621 agttggtcgt gaggcactgg gcaggtaagt atcaaggtta caagacaggt ttaaggagac
     1681 caatagaaac tgggcttgtc gagacagaga agactcttgc gtttctgata ggcacctatt
     1741 ggtcttactg acatccactt tgcctttctc tccacaggtg tccactccca gttcaattac
     1801 agctcttaag gctagagtac ttaatacgac tcactatagg ctagcctcga gaattcgcca
     1861 ccatggagac agacacactc ctgctatggg tactgctgct ctgggttcca ggttccactg
     1921 gtgacacaag tttgtacaaa aaagttggca ccaccggtcc catcatgggc tcctcggtgt
     1981 acatcacggt ggagctggcc attgctgtgc tggccatcct gggcaatgtg ctggtgtgct
     2041 gggccgtgtg gctcaacagc aacctgcaga acgtcaccaa ctactttgtg gtgtcactgg
     2101 cggcggccga catcgcagtg ggtgtgctcg ccatcccctt tgccatcacc atcagcaccg
     2161 ggttctgcgc tgcctgccac ggctgcctct tcattgcctg cttcgtcctg gtcctcacgc
     2221 agagctccat cttcagtctc ctggccatcg ccattgaccg ctacattgcc atccgcatcc
     2281 cgctccggta caatggcttg gtgaccggca cgagggctaa gggcatcatt gccatctgct
     2341 gggtgctgtc gtttgccatc ggcctgactc ccatgctagg ttggaacaac tgcggtcagc
     2401 caaaggaggg caagaaccac tcccagggct gcggggaggg ccaagtggcc tgtctcgctg
     2461 aggatgtggt ccccatgaac tacatggtgt acttcaactt ctttgcctgt gtgctggtgc
     2521 ccctgctgct catgctgggt gtctatttgc ggatcttcct ggcggcgcga cgacagctga
     2581 agcagatgga gagccagcct cccagccgcc ggggtccgga cgccgtcgcc gcgccgccgg
     2641 ggggcaccga gcgcaggccc aacggtctgg gccccgagcg cagcgcgggc ccggggggcg
     2701 cagaggccga accgctgccc acccagctca acggcgcccc tggcgagccc gcgccggccg
     2761 ggccgcgcga caccgacgcg ctggacctgg aggagggagg aaacgtctat atcaaggccg
     2821 acaagcagaa gaacggcatc aaggcgaact tccacatccg ccacaacatc gaggacggcg
     2881 gcgtgcagct cgcctaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     2941 tgcccgacaa ccactacctg agcgtgcagt ccaaactttc gaaagacccc aacgagaagc
     3001 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     3061 agctgtacaa gggcggtacc ggagggagca tggtgagaaa gggcgaggag ctgttcaccg
     3121 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
     3181 ccggcgaggg tgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
     3241 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
     3301 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     3361 aaggctacat ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     3421 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     3481 tcaaggagga cggcaacatc ctggggcaca agctggagta caacaccgga gcagcagcac
     3541 gctggcgcgg gcggcagaac cgcgagctcc atgctgccaa gtcactggcc atcattgtgg
     3601 ggctctttgc cctctgctgg ctgcccctac acatcatcaa ctgcttcact ttcttctgcc
     3661 ccgactgcag ccacgcccct ctctggctca tgtacctggc catcgtcctc tcccacacca
     3721 attcggttgt gaatcccttc atctacgcct accgtatccg cgagttccgc cagaccttcc
     3781 gcaagatcat tcgcagccac gtcctgaggc agcaagaacc tttcaaggca gctggcacca
     3841 gtgcccgggt cttggcagct catggcagtg acggagagca ggtcagcctc cgtctcaacg
     3901 gccacccgcc aggagtgtgg gccaacggca gtgctcccca ccctgagcgg aggcccaatg
     3961 gctacgccct ggggctggtg agtggaggga gtgcccaaga gtcccagggg aacacgggcc
     4021 tcccagacgt ggagctcctt agccatgagc tcaagggagt gtgcccagag ccccctggcc
     4081 tagatgaccc cctggcccag gatggagcag gagtgtcctg agaattcgat atcaagctta
     4141 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     4201 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     4261 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     4321 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     4381 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     4441 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     4501 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     4561 tgctcgccta tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     4621 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
     4681 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat
     4741 accgagcgct gctcgagaga tctacgggtg gcatccctgt gacccctccc cagtgcctct
     4801 cctggccctg gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg
     4861 catcattttg tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg
     4921 gagcaagggg caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag
     4981 ctggagtgca gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat
     5041 tctcctgcct cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta
     5101 atttttgttt ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc
     5161 ctaatctcag gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac
     5221 cactgctccc ttccctgtcc ttctgatttt gtaggtaacc acgtgcggac cgagcggccg
     5281 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     5341 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     5401 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     5461 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     5521 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     5581 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     5641 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     5701 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     5761 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     5821 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     5881 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     5941 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     6001 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     6061 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     6121 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     6181 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     6241 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     6301 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     6361 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     6421 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     6481 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     6541 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     6601 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     6661 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     6721 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     6781 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     6841 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     6901 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6961 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     7021 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     7081 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     7141 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     7201 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7261 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7321 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttc
//
