LOCUS       sequence-465730-        6847 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of glutamate sensor with improved 
            sensitivity and slow deactivation kinetics (4s = slow); 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 6847)
  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 6847)
  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-CAG-flex-iGluSnFR4s-PDGFR-WPRE
FEATURES             Location/Qualifiers
     source          1..6847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   45..174
                     /label=AAV2 ITR
     promoter        485..762
                     /label=chicken beta-actin promoter
     primer_bind     1041..1063
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1125..1144
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1179..1212
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    1275..1308
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1322..1468)
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     CDS             complement(1472..1501)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     1966..1987
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2090..2111)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     sig_peptide     complement(3080..3139)
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     regulatory      3136..3145
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3156..3189)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(3260..3293)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3335..3923
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3388..3408)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3806..3817)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3966..4087
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3977..3996)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4031..4050
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4249..4378
                     /label=AAV2 ITR
     rep_origin      complement(4615..5203)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4695..4714)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5374..6165,6166..6234))
                     /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     5997..6016
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6235..6339)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6365..6820)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6502..6523)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6714..6733
                     /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 agtaccattg
      241 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
      301 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
      361 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
      421 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
      481 atggtcgagg tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc
      541 ccaattttgt atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg
      601 ggggggcgcg cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga
      661 ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg
      721 cggcggcggc ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc
      781 cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc
      841 gcgttactcc cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagcgc
      901 ttggtttaat gacggcttgt ttcttttctg tggctgcgtg aaagccttga ggggctccgg
      961 gagggccctt tgtgcggggg gagcggctcg gggctgtccg cggggggacg gctgccttcg
     1021 ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg accggcggct ctagagcctc
     1081 tgctaaccat gttcatgcct tcttcttttt cctacagctc ctgggcaacg tgctggttat
     1141 tgtgctgtct catcattttg gcaaagaatt ggatccccat aacttcgtat aaagtatcct
     1201 atacgaagtt atatcaaaat aggaagacca atgcttcacc atcgacccga attgccaagc
     1261 atcaccatcg acccataact tcgtataatg tatgctatac gaagttatac tagagctttc
     1321 aacgtggctt cttctgccaa agcatgatga ggatgataag ggagatgatg gtgagcacca
     1381 ccagggccag gatggctgag atcaccacca ccttaaaggg caaggagtgt ggcaccacga
     1441 tgacctcctg cgtgtcctgg cccacagcat tcagatcctc ttctgagatg agtttttgtt
     1501 cgtcgacctg cagtttcagt gccttgtcat tcggttcttt gaacagtgct ttcatttcgt
     1561 ctgacagttc gaaattcatg accagggggt tgttaaagtt gtactccagc ttgtgcccca
     1621 ggatgttgcc gtcctccttg aagtcgatgc cctttagctc gatgcggttc accagggtgt
     1681 cgccctctaa cttcacctcg gcgcgggtct tgtaggttcc gtcgtccttg aagctgatgg
     1741 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt
     1801 ggtcggggta gcgggcgaag cactgcacgc cgtagctcag ggtggtcacg agggtgggcc
     1861 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggttagcttg ccgttggtgg
     1921 catcgccctc gccctcgccg cgcacgctga acttgtggcc gtttacgtcg ccgtccagct
     1981 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcatg ctccctccgg
     2041 taccgccctt gtacagctcg tccatgccga gagagttccc ggtggcggtc acgaactcca
     2101 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagcacggac tgatagctca
     2161 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt
     2221 ggtcggcgag ctgcacgctg ccgtcctcca cgttgtggtg gatgaagaag ttcgccttga
     2281 tgccgttctt ctgcttgtcg gcggtgatat agaccgtgtg gttcaccaga attggatttt
     2341 tgaaccactt atcaaaccat ttttccgctt caccggaggt ctgcacctga gcgatggtgt
     2401 catccatcag ctttttgaac tgcggatcat ctttacgcaa catacaaccg taggcctcct
     2461 gagactgcgg cttgccgacg atttcccagt cgtctggttt cttcgctttc gcacgttcac
     2521 cggccagcag cgagtcatcc atcataaagg caacggcacg accgctttcc agggtgcgga
     2581 aagagtcacc gtgatctttg gcgctgatga tgcgcatatt cattttttgc tcttcattca
     2641 gtttgttgag caaaacttca gaggtagtgc cggaagtgac gactacggct ttgtctttca
     2701 ggttggcaaa atctttgata tcgccaccct ttttggtcaa caggcgcgta ccgaccacga
     2761 aaatagtgtc agagaaagcc gcgaattttt ggcgttcgac gttgttggtg gtagaaccac
     2821 attcaaaatc gtaagtgccg ttttgcagta gtggaatacg gttttgtgaa gtaatcggaa
     2881 tcagttttac ctgcaagtcc ggtttgttga gtttcttttt cactgcttca acaatggcgt
     2941 tggagtattc ctgcgagtaa cccaccactt tgtgctgact gtcttgataa gagaaaggca
     3001 ctgaagagtc acggtgaccg acgacaatca caccgttttt ggcgattttg tccagcgtgc
     3061 tgcctgcggc ggatccgtca ccagtggaac ctggaaccca gagcagcagt acccatagca
     3121 ggagtgtgtc tgtctccatg gtggcggatc ctagtataac ttcgtatagg atactttata
     3181 cgaagttatc attgggattc ttcctatttt gatccaagca tcaccatcga ccctctagtc
     3241 cagatctcac catcgaccca taacttcgta tagcatacat tatacgaagt tatgtccctc
     3301 gaagaggttc gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg
     3361 tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc
     3421 tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta
     3481 taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt
     3541 ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca
     3601 gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc
     3661 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     3721 gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg
     3781 cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg
     3841 cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat
     3901 ctccctttgg gccgcctccc cgcatcgata ccgtcgaccc gggcggccgc ttcgagcaga
     3961 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     4021 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     4081 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg agatgtggga
     4141 ggttttttaa agcaagtaaa acctctacaa atgtggtaaa atcgataagg atcttcctag
     4201 agcatggcta cgtagataag tagcatggcg ggttaatcat taactacaag gaacccctag
     4261 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     4321 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
     4381 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     4441 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     4501 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     4561 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     4621 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     4681 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     4741 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     4801 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     4861 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     4921 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     4981 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5041 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5101 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5161 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5221 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5281 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5341 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     5401 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     5461 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     5521 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     5581 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     5641 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     5701 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     5761 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     5821 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     5881 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     5941 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6001 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6061 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6121 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6181 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6241 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6301 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6361 acctaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     6421 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     6481 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     6541 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     6601 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     6661 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     6721 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     6781 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg
     6841 cgcaact
//
