LOCUS       sequence-484526-        5897 bp    DNA     circular SYN 25-AUG-2025
DEFINITION  pAAV vector for Cre-dependent OCaMP (orange calcium indicator) 
            expression under the control of hSyn promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5897)
  AUTHORS   Aggarwal A, Baker HA, Durst CD, Chen I-W, de Chambrier P, Gonzales 
            JM, Marvin JS, Vandal M, Lundberg T, Sakoi K, Patel RH, Wang C-Y, 
            Visser F, Fouad Y, Sunil S, Wiens M, Terai T, Takahashi-Yamashiro K,
            Thompson RJ, Brown TA, Nasu Y, Nguyen MD, Gordon GRJ, McFarlane S, 
            Podgorski K, Holtmaat A, Campbell RE, Lohman AW
  TITLE     A sensitive orange fluorescent calcium ion indicator for imaging 
            neural activity
  JOURNAL   bioRxiv 2025.07.28.667269; doi: 
            https://doi.org/10.1101/2025.07.28.667269
REFERENCE   2  (bases 1 to 5897)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 25, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2025.07.28.667269; doi: https://doi.org/10.1101/2025.07.28.667269"
COMMENT     Sequence Label: pAAV-hSyn-FLEX(cre)-OCaMP-WPRE
FEATURES             Location/Qualifiers
     source          1..5897
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     protein_bind    137..157
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     promoter        179..626
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    626..650
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(651..684)
                     /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(717..750)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(1230..1249)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             complement(1945..2001)
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     CDS             complement(join(2017..2031,2032..2040))
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             complement(2044..2076)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             complement(2080..2097)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    2176..2209
                     /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    2242..2275
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(2276..2300)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2310..2898
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2363..2383)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2781..2792)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2928..2945)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2934..3141
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3149..3289
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3149..3278
                     /label=AAV2 ITR
     rep_origin      3364..3819
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3451..3470)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3661..3682
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3836..3855)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3955..3977
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4015..4033)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4101..4205
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4206..4274,4275..5066)
                     /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(4424..4443)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5237..5825
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5726..5745
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   join(5887..5897,1..130)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tctagacaac tttgtataga aaagttgctg cagagggccc tgcgtatgag
      181 tgcaagtggg ttttaggacc aggatgaggc ggggtggggg tgcctacctg acgaccgacc
      241 ccgacccact ggacaagcac ccaaccccca ttccccaaat tgcgcatccc ctatcagaga
      301 gggggagggg aaacaggatg cggcgaggcg cgtgcgcact gccagcttca gcaccgcgga
      361 cagtgccttc gcccccgcct ggcggcgcgc gccaccgccg cctcagcact gaaggcgcgc
      421 tgacgtcact cgccggtccc ccgcaaactc cccttcccgg ccaccttggt cgcgtccgcg
      481 ccgccgccgg cccagccgga ccgcaccacg cgaggcgcga gatagggggg cacgggcgcg
      541 accatctgcg ctgcggcgcc ggcgactcag cgctgcctca gtctgcggtg ggcagcggag
      601 gagtcgtgtc gtgcctgaga gcgcagcaag tttgtacaaa aaagcaggct ataacttcgt
      661 ataggatact ttatacgaag ttatcctcga gtaccggtca cgcgtacgca agcttgataa
      721 cttcgtatag catacattat acgaagttat cctagggcca ccttacttcg ctgtcatcat
      781 ttgtacaaac tcttcgtagt ttacctgacc atccccatcg atgtctgcta ccctgatcat
      841 ttcatcaacc tcctcatctg ttaacttctc tccaaggtct gtcatcacgt ggcgaagctc
      901 tgctgcgccg atgtagccat tgccgtcctt atcaaacacg cggaacgctt ctctaatttc
      961 ttcttcactg tctgtgtcat tcatctttct tgccatcatc gtcaggaact cagggaagtc
     1021 gaatgtgccg tcaccgtcgg catctacttc attgatcatg tcctgcagct ctgcttctgt
     1081 ggggttctgc cccagagacc ggaacaccgt ccccagctcc ttggttgtta tcgtcccatc
     1141 cccgtccttg tcaaataggg agaaagcctc tttaaattct gcgatctgct cttcagtcag
     1201 ttggtcacgc gtagcctccc agcccatggt cttcttctgc attacggggc cgtcgggggg
     1261 gaagttggtg ccgcgcagct tcaccttgta gatgaatacg ccgtcctgca gggaggagtc
     1321 ctggttaacg tgaataatgc cgccgtcctc gaagttcatc acgcgctccc acctgaagcc
     1381 ctcggggaag gacagcttaa agtagtcggg gatgtcggct gggtgcttaa tgtaggcctt
     1441 ggagccgtat gtgaactgag gggacaggat gtcccaggcg aagggcaggg ggccaccctt
     1501 ggtcaccttc agcttagcgg tctgaaaggc ctcgtagggg cggccctcgc cctcgccctc
     1561 gatctcgaac tcgtggccgt tcacggagcc ctccatgtgc accttgaagc gcatgaactc
     1621 cttgatgatg gccatgttat cctcctcgcc cttgctcacc agactccctc ctgtacctcc
     1681 cttgtacagc tcgtccctgc cgccggtggg gtggcggccc tcggcgcgtt cgcactgttc
     1741 cacgatggtg tagtcctcgt tgtgggacac gatgtccaac tttatgtcga cgatgtaggc
     1801 gccgggcagc tgcacgggct tcttggcctt gtaggtggtc ttgacctcgg cggcgtagtg
     1861 gccgccgtcc ttcagcctca gccccttctt gatctcgctc ttcaggacgc cgtcctcggg
     1921 gtacatccgc tcggaaacca cgggtgagct cagccgacct atagctctga ctgcgtgacc
     1981 tgccttgttc cacttacgac gggatcccca tcgatcctta tcgtcatcgt cgtacagatc
     2041 ccgacccatt tgctgtccac cagtcatgct agccatacca tgatgatgat gatgatgaga
     2101 acctcctcca gtcttgttaa tatcaagtcc agccaactta agagcaagct cgttctgcag
     2161 catggtggct ctagaataac ttcgtataaa gtatcctata cgaagttata ggtacctgag
     2221 ctcaacatcg atagcactag tataacttcg tataatgtat gctatacgaa gttataccca
     2281 gctttcttgt acaaagtggg aattccgata atcaacctct ggattacaaa atttgtgaaa
     2341 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     2401 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     2461 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     2521 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     2581 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     2641 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     2701 ggaagctgac gtcctttcca tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     2761 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     2821 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     2881 tttgggccgc ctccccgcat cgggaattcc tagagctcgc tgatcagcct cgactgtgcc
     2941 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     3001 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     3061 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     3121 gaatagcagg catgctgggg agggccgcag gaacccctag tgatggagtt ggccactccc
     3181 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     3241 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     3301 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     3361 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     3421 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     3481 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     3541 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     3601 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3661 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     3721 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     3781 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     3841 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     3901 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     3961 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     4021 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     4081 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     4141 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4201 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     4261 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     4321 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     4381 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     4441 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     4501 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     4561 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     4621 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     4681 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     4741 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     4801 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     4861 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     4921 gaagccgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     4981 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     5041 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     5101 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     5161 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     5221 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     5281 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     5341 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     5401 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     5461 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     5521 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     5581 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     5641 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     5701 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     5761 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     5821 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     5881 acatgtcctg caggcag
//
