LOCUS       Exported                6792 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Express the medium-affinity adenosine sensor GRAB_Ado1.0med in Cre 
            positive cells.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-EFS-DIO-GRAB_Ado1.0med
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6792)
  AUTHORS   Zhaofa Wu, Yuting Cui, Huan Wang, Kun Song, Zhengwei Yuan, Ao Dong, 
            Hao Wu, Yi Wan, Sunlei Pan, Wanling Peng, Miao Jing, Min Xu, Minmin 
            Luo, Yulong Li
  TITLE     A GRAB sensor reveals activity-dependent non-vesicular 
            somatodendritic adenosine release
  JOURNAL   bioRxiv
REFERENCE   2  (bases 1 to 6792)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6792
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        151..362
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    381..414
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    465..498
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     1215..1234
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1474..1495
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1601..1622)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(2677..2701)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     sig_peptide     complement(2702..2764)
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
     regulatory      2761..2770
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    complement(2777..2810)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(2861..2894)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    2919..3507
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2972..2992)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3390..3401)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3539..4015
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3738..3757)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4055..4195
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4055..4184
                     /label=AAV2 ITR
     rep_origin      4270..4725
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4357..4376)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4567..4588
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4742..4761)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4861..4883
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4921..4939)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5007..5111
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5112..5972
                     /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(5330..5349)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6143..6731
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6632..6651
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaagctt gggcagagcg cacatcgccc acagtccccg
      181 agaagttggg gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa
      241 actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt
      301 atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac
      361 agtctagagt cgactccgga ataacttcgt ataggatact ttatacgaag ttatgcagaa
      421 tggtagctgg attgtagctg ctattagcaa tatgaaacct cttaataact tcgtatagca
      481 tacattatac gaagttatgg cgcatatgtc aggacactcc tgctccatcc tgggccaggg
      541 ggtcatctag gccagggggc tctgggcaca ctcccttgag ctcatggcta aggagctcca
      601 cgtctgggag gcccgtgttc ccctgggact cttgggcact ccctccactc accagcccca
      661 gggcgtagcc attgggcctc cgctcagggt ggggagcact gccgttggcc cacactcctg
      721 gcgggtggcc gttgagacgg aggctgacct gctctccgtc actgccatga gctgccaaga
      781 cccgggcact ggtgccagct gccttgaaag gttcttgctg cctcaggacg tggctgcgaa
      841 tgatcttgcg gaaggtctgg cggaactcgc ggatacggta ggcgtagatg aagggattca
      901 caaccgaatt ggtgtgggag aggacgatgg ccaggtagtg gagccagaga ggggcgtggc
      961 tgcagtcggg gcagaagaaa gtgaagcagt tgatgatgtg taggggcagc cagcagaggg
     1021 caaagagccc cacaatgatg gccagtgact tggcagcatg gagctcgcgg ttctgccgcc
     1081 cgcgccagcg tgctgctgct ccggtgttgt actccagctt gtgccccagg atgttgccgt
     1141 cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact
     1201 tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg cgctcctgga
     1261 tgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc
     1321 ggctgaagca ctgcacgccg taggtcaggg tggtcacgag ggtgggccag ggcacgggca
     1381 gcttgccggt ggtgcagatg aacttcaggg tcagcttgcc gtaggtggca tcgccctcac
     1441 cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg
     1501 gcaccacccc ggtgaacagc tcctcgccct ttctcaccat gctccctccg gtaccgccct
     1561 tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca
     1621 tgtgatcgcg cttctcgttg gggtctttcg aaagtttgga ctgcacgctc aggtagtggt
     1681 tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtaggcga
     1741 gctgcacgcc gccgtcctcg atgttgtggc ggatgtggaa gttcgccttg atgccgttct
     1801 tctgcttgtc ggccttgata tagacgtttc ctccctcctc caggtccagc gcgtcggtgt
     1861 cgcgcggccc ggccggcgcg ggctcgccag gggcgccgtt gagctgggtg ggcagcggtt
     1921 cggcctctgc gccccccggg cccgcgctgc gctcggggcc cagaccgttg ggcctgcgct
     1981 cggtgccccc cggcggcgcg gcgacggcgt ccggaccccg gcggctggga ggctggctct
     2041 ccatctgctt cagctgtcgt cgcgccgcca ggaagatccg caaatagaca cccagcatga
     2101 gcagcagggg caccagcaca caggcaaaga agttgaagta caccatgtag ttcatgggga
     2161 ccacatcctc aaagagacag gccacttggc cctccccgca gccctgggag tggttcttgc
     2221 cctcctttgg ctgaccgcag ttgttccaac ctagcatggg agtcaggccg atggcaaacg
     2281 acagcaccca gcagatggca atgatgccct tagccctcgt gccggtcacc aagccattgt
     2341 accggagcgg gatgcggatg gcaatgtagc ggtcaatggc gatggccagg agactgaaga
     2401 tggagctctg cgtgaggacc aggacgaagc aggcaatgaa gaggcagccg tggcaggcag
     2461 cgcagaaccc ggtgctgatg gtgatggcaa aggggatggc gagcacaccc actgcgatgt
     2521 cggccgccgc cagtgacacc acaaagtagt tggtgacgtt ctgcaggttg ctgttgagcc
     2581 acacggccca gcacaccagc acattgccca ggatggccag cacagcaatg gccagctcca
     2641 ccgtgatgta caccgaggag cccatgatgg gaccggtgcc aacttttttg tacaaacttg
     2701 tgtcaccagt ggaacctgga acccagagca gcagtaccca tagcaggagt gtgtctgtct
     2761 ccatggtggc gctagcataa cttcgtataa agtatcctat acgaagttat ttgccttaac
     2821 ccagaaatta tcactgttat tctttagaat ggtgcaaaga ataacttcgt ataatgtatg
     2881 ctatacgaag ttatgaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa
     2941 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     3001 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     3061 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     3121 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     3181 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     3241 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     3301 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc
     3361 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     3421 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     3481 ggatctccct ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctacgg
     3541 gtggcatccc tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt
     3601 gcccaccagc cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt
     3661 ctataatatt atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac
     3721 ctgtagggcc tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct
     3781 cactgcaatc tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt
     3841 gggattccag gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg
     3901 tttcaccata ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg
     3961 gcctcccaaa ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat
     4021 tttgtaggta accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc
     4081 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     4141 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc
     4201 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc
     4261 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4321 gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc tttcgctttc ttcccttcct
     4381 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4441 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac
     4501 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     4561 ttaatagtgg actcttgttc caaactggaa caacactcaa ctctatctcg ggctattctt
     4621 ttgatttata agggattttg ccgatttcgg tctattggtt aaaaaatgag ctgatttaac
     4681 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca
     4741 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     4801 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     4861 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     4921 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     4981 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     5041 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     5101 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     5161 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     5221 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     5281 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     5341 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     5401 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     5461 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     5521 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     5581 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     5641 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     5701 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     5761 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     5821 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     5881 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     5941 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     6001 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     6061 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     6121 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     6181 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     6241 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     6301 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     6361 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     6421 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6481 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6541 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6601 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     6661 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     6721 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     6781 ttgctcacat gt
//
