pAAV-GfaABC1D-RGEPO2

LOCUS       pAAV-GfaABC1D-RGEPO2              6262 bp DNA     circular SYN 26-MAR-2024
DEFINITION  Express the adenosine sensor GRAB_Ado1.0 in astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6262)
  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 6262)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 4, 2026 from SnapGene 6.0.2
            https://www.biofeng.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"
FEATURES             Location/Qualifiers
     source          1..6262
                     /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"
     promoter        855..1356
                     /label=GfaABC1D
     misc_feature    931..936
                     /label=SDS
     misc_feature    998..1003
                     /label=SDS
     primer_bind     complement(1200..1211)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     misc_feature    1306..1311
                     /label=SDS
     primer_bind     1505..1526
                     /label=GfaABC1D-F
     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"
     primer_bind     1837..1875
                     /label=Gfa-NmA-F1
     primer_bind     complement(1839..1858)
                     /label=R1
     regulatory      1857..1862
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1863..2315
                     /codon_start=1
                     /label=N-mApple
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEA
                     FQTARLKVTKGGPLPFAWDILSPQFMYGSKAYIKHPADIPDYFKLSFPEGFRWERVMSF
                     EDGGIIHVNQDSSLQDGVFIYKVKLLGTNFPPDGPVMQKKTMGWEAS"
     primer_bind     2289..2336
                     /label=FR-1-3-F
     primer_bind     2289..2310
                     /label=FR-Y-Ra-F
     primer_bind     complement(2302..2315)
                     /label=N-mapple-R
     primer_bind     complement(2304..2322)
                     /label=FR-1-3-R
     primer_bind     complement(2311..2329)
                     /label=0905-R1
     primer_bind     2312..2351
                     /label=PA-S9A-F1
     primer_bind     2313..2345
                     /label=PA-F6A-F1
     primer_bind     2313..2336
                     /label=PA-K8A-F1
     CDS             2316..2318
                     /codon_start=1
                     /label=LINKER1
                     /translation="L"
     CDS             2319..2750
                     /codon_start=1
                     /label=YgaU_Hydrothermal
                     /translation="GIWDFVKSAGRKIGIGSEEAAPAADDLKKELDSYKLGTDDVKVEV
                     VGDKAVITGNVADQSILEKAIVAVGNTLGISKVESNVEVPDAKEPVFHTVKKGDTLWAI
                     ASKQYGNVSKYNAIFEANKPMLSHPDKIYPGQVLRIPDLG"
     primer_bind     2352..2369
                     /label=0811-F2
     primer_bind     2509..2541
                     /label=PA-T73A-F1
     primer_bind     2513..2547
                     /label=PA-G75A-F1
     primer_bind     complement(2513..2530)
                     /label=0905-R2
     primer_bind     2550..2568
                     /label=0811-F1
     CDS             2751..2759
                     /codon_start=1
                     /label=LINKER
                     /translation="SNP"
     CDS             2760..3017
                     /codon_start=1
                     /label=C-mApple
                     /translation="VSEWMYPEDGALKSEIRKGLRLKDGGHYAAEVKTTYKAKKPVQLP
                     GAYIVDIELDILSHNEDYTIVEQCERAVGRHPTGGTDALYK"
     primer_bind     complement(3001..3040)
                     /label=CmA-WPRE-R1
     primer_bind     3021..3040
                     /label=WPRE-F1
     misc_feature    3039..3627
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3092..3112)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     misc_feature    3175..3180
                     /label=SDS
     CDS             complement(3510..3521)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3659..4135
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3858..3877)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    3915..3920
                     /label=SDS
     repeat_region   4175..4315
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4175..4304
                     /label=AAV2 ITR
     rep_origin      4390..4845
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4477..4496)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4687..4708
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4862..4881)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4981..5003
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5041..5059)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5127..5231
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5232..6092
                     /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     complement(5450..5469)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     5624..5645
                     /label=Amp-F1
     primer_bind     complement(5626..5648)
                     /label=Amp-R1
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 ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc atgcgcttca
     1921 aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg
     1981 gccgccccta cgaggccttc cagaccgcta ggctgaaggt gaccaagggt ggccccctgc
     2041 ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacattaagc
     2101 acccagccga catccccgac tacttcaagc tgtccttccc cgagggcttc aggtgggagc
     2161 gcgtgatgag cttcgaggac ggcggcatta ttcacgttaa tcaggactcc tccctgcagg
     2221 acggcgtatt catctacaag gtgaagctgc tcggcaccaa cttccccccc gacggccccg
     2281 taatgcagaa gaagaccatg ggctgggagg cttccctggg catctgggac ttcgtgaaga
     2341 gcgccggcag gaagatcggc atcggcagcg aggaggccgc ccccgccgcc gacgacctga
     2401 agaaggagct ggacagctac aagctgggca ccgacgacgt gaaggtggag gtggtgggcg
     2461 acaaggccgt gatcaccggc aacgtggccg accagagcat cctggagaag gccatcgtgg
     2521 ccgtgggcaa caccctgggc atcagcaagg tggagagcaa cgtggaggtg cccgacgcca
     2581 aggagcccgt gttccacacc gtgaagaagg gcgacaccct gtgggccatc gccagcaagc
     2641 agtacggcaa cgtcagcaag tacaacgcca tcttcgaggc caacaagccc atgctgagcc
     2701 accccgacaa gatctacccc ggccaggtgc tgaggatccc cgacctgggc tctaacccgg
     2761 tttccgagtg gatgtacccc gaggacggcg ccctgaagag cgagatcagg aaggggctga
     2821 ggctgaagga cggcggccac tacgccgccg aggtcaagac cacctacaag gccaagaagc
     2881 ccgtgcagct gcccggcgcc tacatcgtcg acatcgagtt ggacatcttg tcccacaacg
     2941 aggactacac catcgtggaa cagtgcgaac gcgccgtggg ccgccacccc accggtggca
     3001 cggacgcgct gtacaagtag gatatcaagc ttatcgataa tcaacctctg gattacaaaa
     3061 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     3121 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     3181 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     3241 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     3301 gtcagctcct ttccggaact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     3361 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     3421 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc
     3481 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     3541 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     3601 ggatctccct ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctacgg
     3661 gtggcatccc tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt
     3721 gcccaccagc cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt
     3781 ctataatatt atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac
     3841 ctgtagggcc tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct
     3901 cactgcaatc tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt
     3961 gggattccag gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg
     4021 tttcaccata ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg
     4081 gcctcccaaa ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat
     4141 tttgtaggta accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc
     4201 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     4261 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc
     4321 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc
     4381 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4441 gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc tttcgctttc ttcccttcct
     4501 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4561 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac
     4621 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     4681 ttaatagtgg actcttgttc caaactggaa caacactcaa ctctatctcg ggctattctt
     4741 ttgatttata agggattttg ccgatttcgg tctattggtt aaaaaatgag ctgatttaac
     4801 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca
     4861 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     4921 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     4981 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     5041 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     5101 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     5161 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     5221 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     5281 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     5341 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     5401 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     5461 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     5521 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     5581 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     5641 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     5701 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     5761 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     5821 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     5881 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     5941 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     6001 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     6061 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     6121 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     6181 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     6241 tagaaaagat caaaggatct tc
//