pAAV-GfaABC1D-GRAB_eCB2.0

LOCUS       pAAV-GfaABC1D-GRAB_eCB2.0                7327 bp DNA     circular SYN 21-APR-2023
DEFINITION  Express the endocannabinoid sensor GRAB_eCB2.0 in astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7327)
  AUTHORS   Dong A, He K, Dudok B, Farrell JS, Guan W, Liput DJ, Puhl HL, Cai R,
            Duan J, Albarran E, Ding J, Lovinger DM, Li B, Soltesz I, Li Y
  TITLE     A fluorescent sensor for spatiotemporally resolved endocannabinoid 
            dynamics in vitro and in vivo
  JOURNAL   bioRxiv October 20, 2020
REFERENCE   2  (bases 1 to 7327)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 28, 2026 from SnapGene 6.0.2
            https://www.biofeng.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            October 20, 2020"
FEATURES             Location/Qualifiers
     source          1..7327
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        205..706
                     /label=GfaABC1D
     primer_bind     complement(550..561)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     intron          995..1127
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1110..1129
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1172..1190
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1207..1216
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     1213..1275
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTGD"
     protein_bind    1276..1300
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1300..3429
                     /codon_start=1
                     /label=GRAB_eCB2.0
                     /translation="TKSILDGLADTTFRTITTDLLYVGSNDIQYEDIKGDMASKLGYFP
                     QKFPLTSFRGSPFQEKMTAGDNPQLVPADQVNITEFYNKSLSSFKENEENIQCGENFMD
                     IECFMVLNPSQQLAIAVLSLTLGTFTVLENLLVLCVILHSRSLRCRPSYHFIGSLAVAD
                     LLGSVIFVYSFIDFHVFHRKDSRNVFLFKLGGVTASFTASVGSLFLTAIDRYISIHRPL
                     AYKRIVTRPKAVVAFCLMWTIAIVIAVLPLLGWNCEKLQSVCSDIFPHIDETYLMFWIG
                     VTSVLLLFIVYAYMYILWKAHSHAVRMIQRTDALDLEEGGNVYIKADKQKNGIKANFCI
                     RHNIEDGGVQLAYHYQQNTPIGDGPVLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTA
                     AGITFGMDELYKGGTGGSMVRKGEELFTGVVPILVELDGDVNGHKFSVGGEGEGDATVG
                     KLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIF
                     FKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNTGAAARWRGRRMDI
                     RLAKTLVLILVVLIICWGPLLAIMVYDVFGKMNKLIKTVFAFCTMLCLLNSTVNPIIYA
                     LRSKDLRHAFRSMFPSCEGTAQPLDNSMGDSDCLHKHANNAASVHRAAESCIKSTVKIA
                     KVTMSVSTDTSAEAL"
     primer_bind     1796..1818
                     /label=WKG145-F
     primer_bind     2469..2490
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2596..2617)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2857..2876)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    3454..4042
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3507..3527)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3925..3936)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4074..4550
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4273..4292)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4590..4730
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4590..4719
                     /label=AAV2 ITR
     rep_origin      4805..5260
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4892..4911)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5102..5123
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5277..5296)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5396..5418
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5456..5474)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5542..5646
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5647..6507
                     /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(5865..5884)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6678..7266
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7167..7186
                     /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 gcgttgtagt taatgattaa cccgccatgc tacttatcta
      181 cgtagccatg ctctaggaag atctaacata tcctggtgtg gagtagggga cgctgctctg
      241 acagaggctc gggggcctga gctggctctg tgagctgggg aggaggcaga cagccaggcc
      301 ttgtctgcaa gcagacctgg cagcattggg ctggccgccc cccagggcct cctcttcatg
      361 cccagtgaat gactcacctt ggcacagaca caatgttcgg ggtgggcaca gtgcctgctt
      421 cccgccgcac cccagccccc ctcaaatgcc ttccgagaag cccattgagc agggggcttg
      481 cattgcaccc cagcctgaca gcctggcatc ttgggataaa agcagcacag ccccctaggg
      541 gctgcccttg ctgtgtggcg ccaccggcgg tggagaacaa ggctctattc agcctgtgcc
      601 caggaaaggg gatcagggga tgcccaggca tggacagtgg gtggcagggg gggagaggag
      661 ggctgtctgc ttcccagaag tccaaggaca caaatgggtg aggggagagc tctccccata
      721 gctgggctgc ggcccaaccc caccccctca ggctatgcca gggggtgttg ccaggggcac
      781 ccgggcatcg ccagtctagc ccactccttc ataaagccct cgcatcccag gagcgagcag
      841 agccagagca ggttggagag gagacgcatc acctccgctg ctcgcaagct ttattgcggt
      901 agtttatcac agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt
      961 aagctgcaga agttggtcgt gaggcactgg gcaggtaagt atcaaggtta caagacaggt
     1021 ttaaggagac caatagaaac tgggcttgtc gagacagaga agactcttgc gtttctgata
     1081 ggcacctatt ggtcttactg acatccactt tgcctttctc tccacaggtg tccactccca
     1141 gttcaattac agctcttaag gctagagtac ttaatacgac tcactatagg ctagcctcga
     1201 gaattcgcca ccatggagac agacacactc ctgctatggg tactgctgct ctgggttcca
     1261 ggttccactg gtgacacaag tttgtacaaa aaagttggca ccaagtcgat cctagatggc
     1321 cttgcagata ccaccttccg caccatcacc actgacctcc tgtacgtggg ctcaaatgac
     1381 attcagtacg aagacatcaa aggtgacatg gcatccaaat tagggtactt cccacagaaa
     1441 ttccctttaa cttcctttag gggaagtccc ttccaagaga agatgactgc gggagacaac
     1501 ccccagctag tcccagcaga ccaggtgaac attacagaat tttacaacaa gtctctctcg
     1561 tccttcaagg agaatgagga gaacatccag tgtggggaga acttcatgga catagagtgt
     1621 ttcatggtcc tgaaccccag ccagcagctg gccattgcag tcctgtccct cacgctgggc
     1681 accttcacgg tcctggagaa cctcctggtg ctgtgcgtca tcctccactc ccgcagcctc
     1741 cgctgcaggc cttcctacca cttcatcggc agcctggcgg tggcagacct cctggggagt
     1801 gtcatttttg tctacagctt cattgacttc cacgtgttcc accgcaaaga tagccgcaac
     1861 gtgtttctgt tcaaactggg tggggtcacg gcctccttca ctgcctccgt gggcagcctg
     1921 ttcctcacag ccatcgacag gtacatatcc attcacaggc ccctggccta taagaggatt
     1981 gtcaccaggc ccaaggccgt ggtggcgttt tgcctgatgt ggaccatagc cattgtgatc
     2041 gccgtgctgc ctctcctggg ctggaactgc gagaaactgc aatctgtttg ctcagacatt
     2101 ttcccacaca ttgatgaaac ctacctgatg ttctggatcg gggtcaccag cgtactgctt
     2161 ctgttcatcg tgtatgcgta catgtatatt ctctggaagg ctcacagcca cgccgtccgc
     2221 atgattcagc gtaccgacgc gctggacctg gaggagggag gaaacgtcta tatcaaggcc
     2281 gacaagcaga agaacggcat caaggcgaac ttctgcatcc gccacaacat cgaggacggc
     2341 ggcgtgcagc tcgcctacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     2401 ctgcccgaca accactacct gagcgtgcag tccaaacttt cgaaagaccc caacgagaag
     2461 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcacttt cggcatggac
     2521 gagctgtaca agggcggtac cggagggagc atggtgagaa agggcgagga gctgttcacc
     2581 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     2641 ggcggcgagg gtgagggcga tgccaccgtt ggcaagctga ccctgaagtt catctgcacc
     2701 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     2761 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     2821 gaaggctaca tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     2881 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     2941 ttcaaggagg acggcaacat cctggggcac aagctggagt acaacaccgg agcagcagca
     3001 cgctggcgcg ggcggcgcat ggacattagg ttagccaaga ccctggtcct gatcctggtg
     3061 gtgttgatca tctgctgggg ccctctgctt gcaatcatgg tgtatgatgt ctttgggaag
     3121 atgaacaagc tcattaagac ggtgtttgca ttctgcacca tgctctgcct gctgaactcc
     3181 accgtgaacc ccatcatcta tgctctgagg agtaaggacc tgcgacacgc tttccggagc
     3241 atgtttccct cttgtgaagg cactgcgcag cctctggata acagcatggg ggactcggac
     3301 tgcctgcaca aacacgcaaa caatgcagcc agtgttcaca gggccgcaga aagctgcatc
     3361 aagagcacgg tcaagattgc caaggtaacc atgtctgtgt ccacagacac gtctgccgag
     3421 gctctgtgag aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     3481 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     3541 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     3601 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     3661 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     3721 ctcctttccg gaactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     3781 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     3841 tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg gattctgcgc
     3901 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     3961 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     4021 tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc tacgggtggc
     4081 atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact ccagtgccca
     4141 ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg tccttctata
     4201 atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag acaacctgta
     4261 gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct tggctcactg
     4321 caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag ttgttgggat
     4381 tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga cggggtttca
     4441 ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca ccttggcctc
     4501 ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt ctgattttgt
     4561 aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc
     4621 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     4681 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     4741 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     4801 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4861 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     4921 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4981 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     5041 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     5101 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     5161 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     5221 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     5281 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     5341 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     5401 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     5461 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5521 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5581 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     5641 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     5701 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5761 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5821 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5881 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5941 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     6001 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     6061 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     6121 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     6181 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     6241 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     6301 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6361 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6421 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6481 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6541 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6601 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     6661 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6721 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6781 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6841 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6901 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6961 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     7021 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     7081 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     7141 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     7201 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     7261 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     7321 cacatgt
//