LOCUS       sequence-434166-        6458 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses optimized PdCO in frame with mScarlet under control of 
            human synapsin1 promotor after Cre-dependent recombination..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6458)
  AUTHORS   Wietek J, Nozownik A, Pulin M, Saraf-Sinik I, Matosevich N, 
            Gowrishankar R, Gat A, Malan D, Brown BJ, Dine J, Imambocus BN, Levy
            R, Sauter K, Litvin A, Regev N, Subramaniam S, Abrera K, Summarli D,
            Goren EM, Mizrachi G, Bitton E, Benjamin A, Copits BA, Sasse P, Rost
            BR, Schmitz D, Bruchas MR, Soba P, Oren-Suissa M, Nir Y, Wiegert JS,
            Yizhar O
  TITLE     A bistable inhibitory optoGPCR for multiplexed optogenetic control 
            of neural circuits.
  JOURNAL   Nat Methods. 2024 May 29. doi: 10.1038/s41592-024-02285-8.
  PUBMED    38811857
REFERENCE   2  (bases 1 to 6458)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Methods. 2024 May 29. doi: 10.1038/s41592-024-02285-8."
COMMENT     Sequence Label: pAAV_hSyn-DIO-PdCO-mScarlet-WPRE
FEATURES             Location/Qualifiers
     source          1..6458
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   826..955
                     /label=AAV2 ITR
     polyA_signal    1117..1238
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1154..1173)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1208..1227
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1267..1283
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1268..1284
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1287..1875
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1393..1404
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1802..1822
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(1900..1933)
                     /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(1984..2017)
                     /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)."
     regulatory      2033..2042
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3215..3910
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(3348..3366)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3616..3635
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    3949..3982
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4033..4066
                     /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)."
     promoter        complement(4112..4559)
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     repeat_region   4641..4770
                     /label=AAV2 ITR
     rep_origin      4842..5297
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4929..4948)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5139..5160
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5323..5427
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5428..5496,5497..6288)
                     /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(5646..5665)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctgcg cgctcgctcg
      841 ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca
      901 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttccttgtag
      961 ttaatgatta acccgccatg ctacttatct acgtagccat gctctaggaa gatccttatc
     1021 gattttacca catttgtaga ggttttactt gctttaaaaa acctcccaca tctccccctg
     1081 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1141 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     1201 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgctc gaagcggccg
     1261 cccgggtcga ggtcgacggt atcgatgcgg ggaggcggcc caaagggaga tccgactcgt
     1321 ctgagggcga aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg
     1381 aaggtccgct ggattgaggg ccgaagggac gtagcagaag gacgtcccgc gcagaatcca
     1441 ggtggcaaca caggcgagca gccaaggaaa ggacgatgat ttccccgaca acaccacgga
     1501 attgtcagtg cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag
     1561 ttccgccgtg gcaataggga gggggaaagc gaaagtcccg gaaaggagct gacaggtggt
     1621 ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca gtgcacacca cgccacgttg
     1681 cctgacaacg ggccacaact cctcataaag agacagcaac caggatttat acaaggagga
     1741 gaaaatgaaa gccatacggg aagcaatagc atgatacaaa ggcattaaag cagcgtatcc
     1801 acatagcgta aaaggagcaa catagttaag aataccagtc aatctttcac aaattttgta
     1861 atccagaggt tgattatcga taagcttgat atcgaattca taacttcgta tagcatacat
     1921 tatacgaagt tattctttgc accattctaa agaataacag tgataatttc tgggttaagg
     1981 caaataactt cgtataggat actttatacg aagttatgct agcagcgctg ttgccaccat
     2041 ggacggggag aacttgacga ttccaaatcc tgtcaccgag ttgatggata ccccaatcaa
     2101 cagcacgtat ttccaaaacc ttaatgcgga aacggacggc ggaaatcatt acatctacaa
     2161 tgccttcaca gccaccgatt acaacatttg tgcagcttac cttttcttca ttgcttgtct
     2221 gggagtcagt ctcaacgttt tggtgttggt cctctttatc aaagacagaa agttgaggag
     2281 ccccaataac ttcctgtatg taagtctcgc tctcggggat ctgttggttg cagttttcgg
     2341 caccgcattt aagttcatta ttacagctcg caaaaccctg ttgcgagaag aagacggttt
     2401 ctgcaaatgg tacggcttca taacatacct cggcgggctc gcggcactga tgaccctgag
     2461 cgttattgct tttgttcgtt gtctggccgt gctgcgtctc ggctccttca caggtttgac
     2521 aacgcggatg ggcgtggcgg cgatggcttt catttggatc tactccctgg cattcacact
     2581 cgcgcctctc cttgggtgga atcactacat accggaaggt ttggccactt ggtgtagtat
     2641 cgactggctt tccgatgaaa catccgataa gtcctacgtg tttgctattt tcatcttctg
     2701 ctttcttgtc cctgttctca ttatcgtcgt gtcatacggg ctgatctacg ataaggttcg
     2761 gaaggtagcg aaaacaggcg gttctgttgc caaagcagaa cgcgaagtac tgcgcatgac
     2821 gctgctgatg gttagcctgt tcatgcttgc ctggagtccc tacgctgtta tctgcatgct
     2881 cgccagcttc ggccctaaag acctgctcca tccagtggcc acagtgattc ctgccatgtt
     2941 tgcaaagtct tctacgatgt ataaccctct tatctatgtg ttcatgaaca aacaattcag
     3001 gcggagcctt aaagttcttc tcggaatggg agtagaggac ctgaactccg agagtgagag
     3061 agcaactggt ggtacagcca cgaatcaagt cgccgcgacg accgagactt cccaggttgc
     3121 tcctgctccg cgagcccgag atccaaccgg taagagcagg atcaccagcg agggcgagta
     3181 catccccctg gaccagatcg acatcaacgt ggtgatggtg agcaagggcg aggcagtgat
     3241 caaggagttc atgcggttca aggtgcacat ggagggctcc atgaacggcc acgagttcga
     3301 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
     3361 gaccaagggt ggccccctgc ccttctcctg ggacatcctg tcccctcagt tcatgtacgg
     3421 ctccagggcc ttcaccaagc accccgccga catccccgac tactataagc agtccttccc
     3481 cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgccg tgaccgtgac
     3541 ccaggacacc tccctggagg acggcaccct gatctacaag gtgaagctcc gcggcaccaa
     3601 cttccctcct gacggccccg taatgcagaa gaagacaatg ggctgggaag cgtccaccga
     3661 gcggttgtac cccgaggacg gcgtgctgaa gggcgacatt aagatggccc tgcgcctgaa
     3721 ggacggcgga cgctacctgg cggacttcaa gaccacctac aaggccaaga agcccgtgca
     3781 gatgcccggc gcctacaacg tcgaccgcaa gttggacatc acctcccaca acgaggacta
     3841 caccgtggtg gaacagtacg aacgctccga gggccgccac tccaccggcg gcatggacga
     3901 gctgtacaag ttctgctacg agaacgaggt gtaagtcgac ggcgcgccat aacttcgtat
     3961 aatgtatgct atacgaagtt attaagaggt ttcatattgc taatagcagc tacaatccag
     4021 ctaccattct gcataacttc gtataaagta tcctatacga agttattccg gagtcgactc
     4081 tagaggatcc ttgctatcag cttgaattcg actgcgctct caggcacgac acgactcctc
     4141 cgctgcccac cgcagactga ggcagcgctg agtcgccggc gccgcagcgc agatggtcgc
     4201 gcccgtgccc ccctatctcg cgcctcgcgt ggtgcggtcc ggctgggccg gcggcggcgc
     4261 ggacgcgacc aaggtggccg ggaaggggag tttgcggggg accggcgagt gacgtcagcg
     4321 cgccttcagt gctgaggcgg cggtggcgcg cgccgccagg cgggggcgaa ggcactgtcc
     4381 gcggtgctga agctggcagt gcgcacgcgc ctcgccgcat cctgtttccc ctccccctct
     4441 ctgatagggg atgcgcaatt tggggaatgg gggttgggtg cttgtccagt gggtcggggt
     4501 cggtcgtcag gtaggcaccc ccaccccgcc tcatcctggt cctaaaaccc acttgcactc
     4561 atacgcaggg ccctctgcag agatcttcct agagcatggc tacgtagata agtagcatgg
     4621 cgggttaatc attaactaca aggaacccct agtgatggag ttggccactc cctctctgcg
     4681 cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg
     4741 ggcggcctca gtgagcgagc gagcgcgcag ctggcgtaat agcgaagagg cccgcaccga
     4801 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
     4861 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     4921 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     4981 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     5041 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     5101 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     5161 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     5221 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     5281 attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     5341 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     5401 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     5461 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     5521 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     5581 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     5641 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     5701 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     5761 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     5821 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     5881 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     5941 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     6001 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     6061 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     6121 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     6181 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     6241 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     6301 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6361 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6421 tgagcgtcag accccgtaga aaagatcaaa ggatcttc
//
