LOCUS       sequence-421886-        6339 bp    DNA     circular SYN 14-APR-2025
DEFINITION  AAV construct expressing dTomato driven by SST interneuron-targeting
            enhancer. Alias: AAV_WDS0004_dTomato_nlsdTomato.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6339)
  AUTHORS   Furlanis E, Dai M, Leyva Garcia B, Vergara J, Pereira A, Pelkey K, 
            Tran T, Gorissen BL, Vlacho A, Hairston A, Huang S, Dwivedi D, Du S,
            Wills S, McMahon J, Lee AT, Chang EF, Razzaq T, Qazi A, Vargish G, 
            Yuan X, Caccavano A, Hunt S, Chittajallu R, McLean N, Hewit L, 
            Paranzino E, Rice H, Cummins AC, Plotnikova A, Mohanty A, Tangen AC,
            Shin J, Azadi R, Eldridge MAG, Alvarez VA, Averbeck BB, Alyahyay M, 
            Reyes Vallejo T, Soheib M, Vattino LG, MacGregor CP, Banks E, Olah 
            V, Naskar S, Hill S, Liebergall S, Badiani R, Hyde L, Xu Q, Allaway 
            KC, Goldberg EM, Nowakowski TJ, Lee S, Takesian AE, Ibrahim LA, 
            Iqbal A, McBain CJ, Dimidschstein J, Fishell G, Wang Y
  TITLE     An enhancer-AAV toolbox to target and manipulate distinct 
            interneuron subtypes
  JOURNAL   bioRxiv 2024.07.17.603924
REFERENCE   2  (bases 1 to 6339)
  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: "bioRxiv 
            2024.07.17.603924"
COMMENT     Sequence Label: pAAV_BiSSTe4_dTomato_nlsdTomato
FEATURES             Location/Qualifiers
     source          1..6339
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          958..1090
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1113..1122
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1119..1820
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     CDS             1842..1898
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1911..1931
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1932..2630
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGL
                     VTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQ
                     ALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHH
                     LFLYGMDELYK"
     misc_feature    2685..3273
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2738..2758)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3156..3167)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3442..3523
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3479..3498)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3533..3552
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   3602..3742
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3602..3731
                     /label=AAV2 ITR
     rep_origin      3817..4272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3904..3923)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4114..4135
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4289..4308)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4408..4430
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4468..4486)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4554..4658
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4659..4727,4728..5519)
                     /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(4877..4896)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5690..6278
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6179..6198
                     /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 gcgtatcgat tttcagcata agcaagatgg aatggacaat
      181 ctcttatttc atcctaagtt ttacaggagg tccttgtttt atgtaaattt ccaaccattt
      241 caaaagctct ttatgtgtgt attttttttt taaccatttt tgacagtttc aactctttct
      301 tctcccatga tttctttttg ctctcaactg ttatgggaga gacaccaggt tctgtatgca
      361 tttctgcttt gggttgaaaa ttttatgcac ttcaaatcaa gcttcattca catagctaaa
      421 ttcactgagg atacttcttc cctttgaaaa tttgacaaga ttcttaacaa cccttccccg
      481 ttctcctcat atttgttgag tttcctagtg catgtccttc tccctgcccc cagaataact
      541 agcaggtgtg acacaggcat gcattttgca ttttccaagt ctgtgatgct aactttaggc
      601 ctagaaacat tagcagattt cctcttttta tcatgcatac atttgctgct tctgttatcc
      661 aatctagtca ctttaaagaa ttttcttatt tcttttggga cctgtgcaat ctgttgcaag
      721 ctagaaatgt ccatggctag accaataaat tcactagata aatttacctg tggccacatt
      781 atttaaaagt ctgtgatcga tgtttaaact tcctgcagcc cgggctgggc ataaaagtca
      841 gggcagagcc atctattgct tacatttgct tctagcctgc aagtcgagga gcgcagcctt
      901 ccagaagcag agcgcggcgc cttaagctgc agaagttggt cgtgaggcac tgggcacgta
      961 agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggctt gtcgagacag
     1021 agaagactct tgcgtttctg ataggcacct attggtctta ctgacatcca ctttgccttt
     1081 ctctccacag cgagctcgct agcgcgatcg ctgccaccat ggtgagcaag ggcgaggagg
     1141 tcatcaaaga gttcatgcgc ttcaaggtgc gcatggaggg ctccatgaac ggccacgagt
     1201 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
     1261 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtccccc cagttcatgt
     1321 acggctccaa ggcgtacgtg aagcaccccg ccgacatccc cgattacaag aagctgtcct
     1381 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggt ctggtgaccg
     1441 tgacccagga ctcctccctg caggacggca cgctgatcta caaggtgaag atgcgcggca
     1501 ccaacttccc ccccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcca
     1561 ccgagcgcct gtacccccgc gacggcgtgc tgaagggcga gatccaccag gccctgaagc
     1621 tgaaggacgg cggccactac ctggtggagt tcaagaccat ctacatggcc aagaagcccg
     1681 tgcaactgcc cggctactac tacgtggaca ccaagctgga catcacctcc cacaacgagg
     1741 actacaccat cgtggaacag tacgagcgct ccgagggccg ccaccacctg ttcctgtacg
     1801 gcatggacga gctgtacaag tccggagata acggaagcgg agccactaac ttctccctgt
     1861 tgaaacaagc aggggatgtc gaagagaatc ccgggccatc tagaatggcc cccaagaaga
     1921 agaggaaggt ggtgagcaag ggcgaggagg tcatcaaaga gttcatgcgc ttcaaggtgc
     1981 gcatggaggg ctccatgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
     2041 cctacgaggg cacccagacc gccaagctga aggtgaccaa gggcggcccc ctgcccttcg
     2101 cctgggacat cctgtccccc cagttcatgt acggctccaa ggcgtacgtg aagcaccccg
     2161 ccgacatccc cgattacaag aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga
     2221 tgaacttcga ggacggcggt ctggtgaccg tgacccagga ctcctccctg caggacggca
     2281 cgctgatcta caaggtgaag atgcgcggca ccaacttccc ccccgacggc cccgtaatgc
     2341 agaagaagac catgggctgg gaggcctcca ccgagcgcct gtacccccgc gacggcgtgc
     2401 tgaagggcga gatccaccag gccctgaagc tgaaggacgg cggccactac ctggtggagt
     2461 tcaagaccat ctacatggcc aagaagcccg tgcaactgcc cggctactac tacgtggaca
     2521 ccaagctgga catcacctcc cacaacgagg actacaccat cgtggaacag tacgagcgct
     2581 ccgagggccg ccaccacctg ttcctgtacg gcatggacga gctgtacaag taagtcgacg
     2641 gcgcgcccca cccctgcagg gaattcgata tcaagcttat cgataatcaa cctctggatt
     2701 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     2761 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     2821 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     2881 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     2941 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     3001 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     3061 ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctat gttgccacct
     3121 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     3181 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     3241 cgagtcggat ctccctttgg gccgcctccc cgcatcgata ccgagcgctg ctcgagagat
     3301 ctgtgatagc ggccatcaag ctggccgcga ctctagatca taatcagcca taccacattt
     3361 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     3421 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     3481 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3541 tccaaactca tcaatgtatc agcttatcga taccgcatgc acgtgcggac cgagcggccg
     3601 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     3661 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     3721 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     3781 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     3841 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     3901 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3961 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4021 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4081 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4141 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     4201 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     4261 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     4321 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     4381 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     4441 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     4501 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4561 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4621 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4681 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4741 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4801 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4861 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     4921 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4981 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     5041 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5101 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5161 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5221 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5281 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5341 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5401 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5461 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5521 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5581 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5641 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     5701 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     5761 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     5821 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5881 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     5941 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     6001 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     6061 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     6121 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     6181 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     6241 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     6301 tacggttcct ggccttttgc tggccttttg ctcacatgt
//
