LOCUS Exported 7369 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Express the adenosine-insensitive control sensor GRAB_Ado1.0mut in astrocytes. ACCESSION . VERSION . KEYWORDS pAAV-GfaABC1D-GRAB_Ado1.0mut 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 May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7369 /organism="synthetic DNA construct" /mol_type="other DNA" 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 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" sig_peptide 1863..1925 /product="leader sequence from mouse immunoglobulin kappa light chain" /label=Ig-kappa leader 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 6339..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" primer_bind complement(6557..6576) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" 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 //