LOCUS       sequence-307196-        6341 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the 2nd generation medium-affinity DA sensor GRAB_DA2m in 
            neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6341)
  AUTHORS   Sun F, Zhou J, Dai B, Qian T, Zeng J, Li X, Zhuo Y, Zhang Y, Wang Y,
            Qian C, Tan K, Feng J, Dong H, Lin D, Cui G, Li Y
  TITLE     Next-generation GRAB sensors for monitoring dopaminergic activity in
            vivo.
  JOURNAL   Nat Methods. 2020 Nov;17(11):1156-1166. doi: 
            10.1038/s41592-020-00981-9. Epub 2020 Oct 21.
  PUBMED    33087905
REFERENCE   2  (bases 1 to 6341)
  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"; doi: 
            "10.1038/s41592-020-00981-9"; journalName: "Nat Methods"; date: 
            "2020-11"; volume: "17"; issue: "11"; pages: "1156-1166"
COMMENT     Sequence Label: pAAV-hsyn-GRAB_DA2m
FEATURES             Location/Qualifiers
     source          1..6341
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      581..590
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     587..646
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     protein_bind    650..674
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     1648..1669
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1778..1799)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2039..2058)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    2468..3056
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2521..2541)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2939..2950)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3088..3564
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3287..3306)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3604..3744
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3604..3733
                     /label=AAV2 ITR
     rep_origin      3819..4274
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3906..3925)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4116..4137
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4291..4310)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4410..4432
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4470..4488)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4556..4660
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4661..4729,4730..5521)
                     /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(4879..4898)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5692..6280
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6181..6200
                     /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 gcgtatgagg cggggtgggg gtgcctacct gacgaccgac
      181 cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag
      241 agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg
      301 acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg
      361 ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc
      421 gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc
      481 gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga
      541 ggagtcgtgt cgtgcctgag agcgcagtcg agaaaccggt gccaccatgg agacagacac
      601 actcctgcta tgggtactgc tgctctgggt tccaggttcc actggtgaca caagtttgta
      661 caaaaaagtt ggcaccaccg gtgatccact gaatctgtcc tggtatgatg atgatctgga
      721 gaggcagaac tggagccggc ccttcaacgg gtcagacggg aaggcggaca gaccccacta
      781 caactactat gccacactgc tcaccctgct catcgctgtc atcgtcttcg gcaacgtgct
      841 ggtgtgcatg gctgtgtccc gcgagaaggc gctgcagacc accaccaact acctgatcgt
      901 cagcctcgca gtggccgacc tcctcgtcgc cacactggtc atgccctggg ttgtctacct
      961 ggaggtggta ggtgagtgga aattcagcag gattcactgt gacatcttcg tcactctgga
     1021 cgtcatgatg tgcacggcga gcatcctgaa cttgtgtgcc atcagcatcg acaggtacac
     1081 agctgtggcc atgcccatgc tgtacaatac gcgctacagc tccaagcgcc gggtcaccgt
     1141 catgatctcc atcgtctggg tcctgtcctt caccatctcc tgcccactcc tcttcggact
     1201 caataacgca gaccagaacg agtgcatcat tgccaacccg gccttcgtgg tctactcctc
     1261 catcgtctcc ttctacgtgc ccttcattgt caccctgctg gtctacatca agatctacat
     1321 tgtcctccgc agacgccgca agcgagtcaa caccaaacgc agcagccgag ctttcagggc
     1381 ccacctgagg gctccactaa agggcaactg tactcacccc gaggacatga aactcggagg
     1441 aactgtctat atcaaggccg acaagcagaa gaacggcatc aaggcgaact tccacatccg
     1501 ccacaacatc gaggacggcg gcgtgcagct cgcctaccac taccagcaga acacccccat
     1561 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcgtgcagt ccaaactttc
     1621 gaaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     1681 gatcactctc ggcatggacg agctgtacaa gggcggtacc ggagggaaat ccatggtgag
     1741 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     1801 aaacggccac aagttcagcg tgtccggcga gggtgagggc gatgccacct acggcaagct
     1861 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     1921 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
     1981 cttcttcaag tccgccatgc ccgaaggcta catccaggag cgcaccatct tcttcaagga
     2041 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     2101 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     2161 gtacaacact ggagcagcag caatgagccg taggaagctc tcccagcaga aggagaagaa
     2221 agccactcag atgctcgcca ttgttctcgg cgtgttcatc atctgctggc tgcccttctt
     2281 catcacacac atcctgaaca tacactgtga ctgcaacatc ccgcctgtcc tgtacagcgc
     2341 cttcacgtgg ctgggctatg tcaacagcgc cgtgaacccc atcatctaca ccaccttcaa
     2401 cattgagttc cgcaaggcct tcctgaagat cctccactgc tgagaattcg atatcaagct
     2461 tatcgataat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     2521 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     2581 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     2641 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     2701 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     2761 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     2821 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg
     2881 gctgctcgcc tatgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     2941 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     3001 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg
     3061 ataccgagcg ctgctcgaga gatctacggg tggcatccct gtgacccctc cccagtgcct
     3121 ctcctggccc tggaagttgc cactccagtg cccaccagcc ttgtcctaat aaaattaagt
     3181 tgcatcattt tgtctgacta ggtgtccttc tataatatta tggggtggag gggggtggta
     3241 tggagcaagg ggcaagttgg gaagacaacc tgtagggcct gcggggtcta ttgggaacca
     3301 agctggagtg cagtggcaca atcttggctc actgcaatct ccgcctcctg ggttcaagcg
     3361 attctcctgc ctcagcctcc cgagttgttg ggattccagg catgcatgac caggctcagc
     3421 taatttttgt ttttttggta gagacggggt ttcaccatat tggccaggct ggtctccaac
     3481 tcctaatctc aggtgatcta cccaccttgg cctcccaaat tgctgggatt acaggcgtga
     3541 accactgctc ccttccctgt ccttctgatt ttgtaggtaa ccacgtgcgg accgagcggc
     3601 cgcaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc tcgctcactg
     3661 aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg
     3721 agcgagcgcg cagctgcctg caggggcgcc tgatgcggta ttttctcctt acgcatctgt
     3781 gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta gcggcgcatt
     3841 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccttagc
     3901 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3961 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     4021 caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     4081 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     4141 aacactcaac tctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggt
     4201 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     4261 aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     4321 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     4381 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     4441 gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     4501 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     4561 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     4621 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     4681 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     4741 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     4801 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     4861 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     4921 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     4981 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     5041 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     5101 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     5161 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     5221 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     5281 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     5341 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5401 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5461 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5521 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     5581 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     5641 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5701 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5761 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     5821 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     5881 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     5941 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     6001 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     6061 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6121 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     6181 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     6241 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     6301 tttacggttc ctggcctttt gctggccttt tgctcacatg t
//
