LOCUS       sequence-465691-        6528 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of glutamate sensor with improved 
            sensitivity and fast deactivation kinetics (4f = fast); NGR vector 
            matches or outperforms PDGFR in all conditions tested..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6528)
  AUTHORS   Aggarwal A, Negrean A, Chen Y, Iyer R, Reep D, Liu A, Palutla A, Xie
            ME, MacLennan BJ, Hagihara KM, Kinsey LW, Sun JL, Yao P, Zheng J, 
            Tsang A, Tsegaye G, Zhang Y, Patel RH, Arthur BJ, Hiblot J, Leippe 
            P, Tarnawski M, Marvin JS, Vevea JD, Turaga SC, Tebo AG, Carandini 
            M, Rossi LF, Kleinfeld D, Konnerth A, Svoboda K, Turner GC, Hasseman
            J, Podgorski K
  TITLE     Glutamate indicators with increased sensitivity and tailored 
            deactivation rates
  JOURNAL   bioRxiv 2025.03.20.643984
REFERENCE   2  (bases 1 to 6528)
  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"; journalName: "bioRxiv 
            2025.03.20.643984"
COMMENT     Sequence Label: pAAV-syn-FLP-FRT-iGluSnFR4f-NGR-WPRE
FEATURES             Location/Qualifiers
     source          1..6528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   45..174
                     /label=AAV2 ITR
     promoter        256..703
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     intron          725..857
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     840..859
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    879..926
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             complement(1157..1186)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     1651..1672
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1775..1796)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     sig_peptide     complement(2765..2824)
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     regulatory      2821..2830
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2866..2913)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    3010..3598
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3063..3083)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3481..3492)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3601..3617)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3602..3618)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    3647..3768
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3658..3677)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3712..3731
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3930..4059
                     /label=AAV2 ITR
     rep_origin      complement(4296..4884)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4376..4395)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5055..5846,5847..5915))
                     /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     5678..5697
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5916..6020)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6046..6501)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6183..6204)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6395..6414
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctgcgc gctcgctcgc
       61 tcactgaggc cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag
      121 tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt tccttgtagt
      181 taatgattaa cccgccatgc tacttatcta cgtagccatg ctctaggaag atctctgcag
      241 agggccctgc gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc
      301 ctacctgacg accgaccccg acccactgga caagcaccca acccccattc cccaaattgc
      361 gcatccccta tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc
      421 agcttcagca ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct
      481 cagcactgaa ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca
      541 ccttggtcgc gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat
      601 aggggggcac gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc
      661 tgcggtgggc agcggaggag tcgtgtcgtg cctgagagcg cagtcgaatt caagctgcta
      721 gcaggtaagt atcaaggtta caagacaggt ttaaggagac caatagaaac tgggcttgtc
      781 gagacagaga agactcttgc gtttctgata ggcacctatt ggtcttactg acatccactt
      841 tgcctttctc tccacaggtg ataaggatcc ggcgcgccga agttcctatt ccgaagttcc
      901 tattctctag aaagtatagg aacttccctt tggccggagc ggccaaagcg ctctgaagtt
      961 cctattccga agttcctatt ctgaactaag tataggaact tcgtttaaac ttaattaaac
     1021 gcgtaccggt ttatcaacag gggcccaaca ccgtccacag aaccaatgcc agtcccagtg
     1081 gagccagact acatgcaagg gcaggaaggg cacctgatcc ttctgcatct cctgttccag
     1141 atgcaccact acctgccaga tcctcttctg agatgagttt ttgttcgtcg acctgcagtt
     1201 tcagtgcctt gtcattcggt tctttgaaca gtgctttcat ttcgtctgac agttcgaaat
     1261 tcatgaccag ggggttgtta aagttgtact ccagcttgtg ccccaggatg ttgccgtcct
     1321 ccttgaagtc gatgcccttt agctcgatgc ggttcaccag ggtgtcgccc tctaacttca
     1381 cctcggcgcg ggtcttgtag gttccgtcgt ccttgaagct gatggtgcgc tcctggacgt
     1441 agccttcggg catggcggac ttgaagaagt cgtgctgctt catgtggtcg gggtagcggg
     1501 cgaagcactg cacgccgtag ctcagggtgg tcacgagggt gggccagggc acgggcagct
     1561 tgccggtggt gcagatgaac ttcagggtta gcttgccgtt ggtggcatcg ccctcgccct
     1621 cgccgcgcac gctgaacttg tggccgttta cgtcgccgtc cagctcgacc aggatgggca
     1681 ccaccccggt gaacagctcc tcgcccttgc tcatgctccc tccggtaccg cccttgtaca
     1741 gctcgtccat gccgagagag ttcccggtgg cggtcacgaa ctccagcagg accatgtgat
     1801 cgcgcttctc gttggggtct ttgctcagca cggactgata gctcaggtag tggttgtcgg
     1861 gcagcagcac ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg gcgagctgca
     1921 cgctgccgtc ctccacgttg tggtggatga agaagttcgc cttgatgccg ttcttctgct
     1981 tgtcggcggt gatatagacc gtgtggttca ccagaattgg atttttgaac cacttatcaa
     2041 accatttttc cgcttcaccg gaggtctgca cctgagcgat ggtgtcatcc atcagctttt
     2101 tgaactgcgg atcatcttta cgcaacatac aaccgtaggc ctcctgagac tgcggcttgc
     2161 cgacgatttc ccagtcgtct ggtttcttcg ctttcgcacg ttcaccggcc agcagtacgt
     2221 catccatcat aaaggcaacg gcacgaccgc tttccagggt gcggaaagag tcaccgtgat
     2281 ctttggcgct gatgatgcgc atattcattt tttgctcttc attcagtttg ttgagcaaaa
     2341 cttcagaggt agtgccggaa gtgacgacta cggctttgtc tttcaggttg gcaaaatctt
     2401 tgatatcgcc accctttttg gtcaacaggc gcgtaccgac cacgaaaata gtgtcagaga
     2461 aagccgcgaa tttttggcgt tcgacgttgt tggtggtaga accacattca aaatcgtaag
     2521 tgccgttttg cagtagtgga atacggtttt gtgaagtaat cggaatcagt tttacctgca
     2581 agtccggttt gttgagtttc tttttcactg cttcaacaat ggcgttggag tattcctgcg
     2641 agtaacccac cactttgtgc tgactgtctt gataagagaa aggcactgaa gagtcacggt
     2701 gaccgacgac aatcacaccg tttttggcga ttttgtccag cgtgctgcct gcggcggatc
     2761 cgtcaccagt ggaacctgga acccagagca gcagtaccca tagcaggagt gtgtctgtct
     2821 ccatggtggc gaattcagat ctactagtgg ccggccagtt taaacgaagt tcctatactt
     2881 tctagagaat aggaacttcg gaataggaac ttccattgcg atcgctccta ttttgatcca
     2941 agaagttcct atacttagtt cagaatagga acttcggaat aggaacttcc ctgcaggaag
     3001 cttatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     3061 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     3121 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     3181 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     3241 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     3301 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     3361 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct
     3421 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     3481 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     3541 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat
     3601 cgataccgtc gacctcgacc cgggcggccg cttcgagcag acatgataag atacattgat
     3661 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     3721 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     3781 tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggtttttta aagcaagtaa
     3841 aacctctaca aatgtggtaa aatcgataag gatcttccta gagcatggct acgtagataa
     3901 gtagcatggc gggttaatca ttaactacaa ggaaccccta gtgatggagt tggccactcc
     3961 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     4021 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcattaatg aatcggccaa
     4081 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4141 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4201 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4261 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4321 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4381 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4441 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4501 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4561 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4621 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4681 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4741 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4801 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4861 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4921 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4981 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5041 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5101 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5161 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5221 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5281 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5341 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5401 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5461 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5521 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5581 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5641 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5701 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5761 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5821 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5881 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5941 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6001 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta
     6061 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     6121 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     6181 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     6241 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     6301 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     6361 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     6421 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     6481 atgcgccgct acagggcgcg tcccattcgc cattcaggct gcgcaact
//
