LOCUS       Exported                7047 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV vector to drive the expression of dChr2-GFP-P2A-GFP in PV 
            cortical interneuronsunder the control of the E14 regulatory 
            element.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-E14-ChR2GFP2x
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7047)
  AUTHORS   Vormstein-Schneider DC, Lin JD, Pelkey KA, Chittajallu R, Guo B, 
            Arias Garcia M, Sakopoulos S, Stevenson O, Schneider G, Zhang Q, 
            Sharma J, Franken TP, Smith J, Vogel I, Sanchez V, Ibrahim LA, 
            Burbridge T, Favuzzi E, Saldi GA, Xu Q, Guo L, Yuan X, Zaghloul KA, 
            Sabri E, Goldberg EM, Devinsky O, Batista-Brito R, Reynolds J, Feng 
            G, Fu Z, McBain CJ, Fishell GJ, Dimidschstein J
  TITLE     Viral manipulation of functionally distinct interneurons in mice, 
            non-human primates and humans
  JOURNAL   Nature Neuroscience (2020)
REFERENCE   2  (bases 1 to 7047)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7047
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          816..948
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     931..950
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      989..998
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             995..1921
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             1937..2650
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             1937..2650
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             2672..2728
                     /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             2741..2761
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2762..3478
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             2762..3475
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     misc_feature    3488..4076
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3541..3561)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3959..3970)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4084..4308
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4310..4450
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4310..4439
                     /label=AAV2 ITR
     rep_origin      4525..4980
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4612..4631)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4822..4843
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4997..5016)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5116..5138
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5176..5194)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5262..5366
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5367..6227
                     /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(5585..5604)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6398..6986
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6887..6906
                     /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 gctagactgc agagggcccg cggccgcacg cgttcggttc tctctgtagt
      181 ctgcaaagaa cccacttctt gcccagtgag agattcggga gatttccact gcactgcctg
      241 aggccactgt gctttaatga gaagaacacc cgcttcctct caaggccagc agcggtctga
      301 atccccagcc cccaaacaca ctgcagcttg cccagctagc tgctcctgtg aggggtgacc
      361 ttgacttgaa gttagcccca acttgctgga caactccacc actttctgag ccccatggta
      421 tatgtgagca ttgttgggag gtgagaggta gtgttgtaat atttaactta ttggactcgt
      481 tagagaaatt aaaatcactg tatgctgcaa ctagcttata gaagggatcc aaagactctt
      541 agatttttct ttttccggga aacaaattta ttaaagagtg ttgtagaggc caggccgtag
      601 cgaattcctg cttgagatgt ccttggtcta tgttgcgttt tgaagacacc cctccccgcg
      661 tttaaacttc ctgcagcccg ggctgggcat aaaagtcagg gcagagccat ctattgctta
      721 catttgcttc tagcctgcaa gtcgaggagc gcagccttcc agaagcagag cgcggcgcct
      781 taagctgcag aagttggtcg tgaggcactg ggcacgtaag tatcaaggtt acaagacagg
      841 tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg cgtttctgat
      901 aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt gtccactccc
      961 agttcaatta cagctcttaa gaaagctagc caccatggac tatggcggcg ctttgtctgc
     1021 cgtcggacgc gaacttttgt tcgttactaa tcctgtggtg gtgaacgggt ccgtcctggt
     1081 ccctgaggat caatgttact gtgccggatg gattgaatct cgcggcacga acggcgctca
     1141 gaccgcgtca aatgtcctgc agtggcttgc agcaggattc agcattttgc tgctgatgtt
     1201 ctatgcctac caaacctgga aatctacatg cggctgggag gagatctatg tgtgcgccat
     1261 tgaaatggtt aaggtgattc tcgagttctt ttttgagttt aagaatccct ctatgctcta
     1321 ccttgccaca ggacaccggg tgcagtggct gcgctatgca gagtggctgc tcacttgtcc
     1381 tgtcatcctt atccgcctga gcaacctcac cggcctgagc aacgactaca gcaggagaac
     1441 catgggactc cttgtctcag acatcgggac tatcgtgtgg ggggctacca gcgccatggc
     1501 aaccggctat gttaaagtca tcttcttttg tcttggattg tgctatggcg cgaacacatt
     1561 ttttcacgcc gccaaagcat atatcgaggg ttatcatact gtgccaaagg gtcggtgccg
     1621 ccaggtcgtg accggcatgg catggctgtt tttcgtgagc tggggtatgt tcccaattct
     1681 cttcattttg gggcccgaag gttttggcgt cctgagcgtc tatggctcca ccgtaggtca
     1741 cacgattatt gatctgatga gtaaaaattg ttgggggttg ttgggacact acctgcgcgt
     1801 cctgatccac gagcacatat tgattcacgg agatatccgc aaaaccacca aactgaacat
     1861 cggcggaacg gagatcgagg tcgagactct cgtcgaagac gaagccgagg ccggagccgt
     1921 gccagctgca gccacagtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     1981 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     2041 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     2101 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     2161 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     2221 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     2281 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     2341 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     2401 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     2461 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     2521 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     2581 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     2641 gctgtacaag tccggagata acggaagcgg agccactaac ttctccctgt tgaaacaagc
     2701 aggggatgtc gaagagaatc ccgggccatc tagaatggcc cccaagaaga agaggaaggt
     2761 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     2821 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     2881 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     2941 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     3001 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     3061 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     3121 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     3181 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     3241 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     3301 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     3361 cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     3421 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaatg
     3481 gcgcgccaat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     3541 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     3601 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     3661 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     3721 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     3781 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     3841 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt cctttccatg
     3901 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     3961 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     4021 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcctgg
     4081 atcctgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     4141 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     4201 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     4261 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggca ggaaccccta
     4321 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
     4381 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
     4441 tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     4501 cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     4561 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     4621 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     4681 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     4741 tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     4801 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     4861 tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     4921 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     4981 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     5041 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     5101 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     5161 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     5221 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     5281 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     5341 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     5401 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     5461 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     5521 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     5581 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     5641 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     5701 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     5761 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     5821 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     5881 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     5941 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     6001 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     6061 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     6121 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     6181 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     6241 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     6301 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     6361 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     6421 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     6481 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     6541 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     6601 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6661 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     6721 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     6781 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     6841 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6901 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     6961 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     7021 ccttttgctg gccttttgct cacatgt
//
