LOCUS       Exported                7059 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 7059)
  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 7059)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7059
                     /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          828..960
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     943..962
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1001..1010
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1007..1933
                     /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             1949..2662
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             1949..2662
                     /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             2684..2740
                     /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             2753..2773
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2774..3490
                     /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             2774..3487
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     misc_feature    3500..4088
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3553..3573)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3971..3982)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4096..4320
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4322..4462
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4322..4451
                     /label=AAV2 ITR
     rep_origin      4537..4992
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4624..4643)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4834..4855
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5009..5028)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5128..5150
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5188..5206)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5274..5378
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5379..6239
                     /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(5597..5616)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6410..6998
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6899..6918
                     /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 cgtatcgatt cggttctctc
      181 tgtagtctgc aaagaaccca cttcttgccc agtgagagat tcgggagatt tccactgcac
      241 tgcctgaggc cactgtgctt taatgagaag aacacccgct tcctctcaag gccagcagcg
      301 gtctgaatcc ccagccccca aacacactgc agcttgccca gctagctgct cctgtgaggg
      361 gtgaccttga cttgaagtta gccccaactt gctggacaac tccaccactt tctgagcccc
      421 atggtatatg tgagcattgt tgggaggtga gaggtagtgt tgtaatattt aacttattgg
      481 actcgttaga gaaattaaaa tcactgtatg ctgcaactag cttatagaag ggatccaaag
      541 actcttagat ttttcttttt ccgggaaaca aatttattaa agagtgttgt agaggccagg
      601 ccgtagcgaa ttcctgcttg agatgtcctt ggtctatgtt gcgttttgaa gacacccctc
      661 cccgcatcga tgtttaaact tcctgcagcc cgggctgggc ataaaagtca gggcagagcc
      721 atctattgct tacatttgct tctagcctgc aagtcgagga gcgcagcctt ccagaagcag
      781 agcgcggcgc cttaagctgc agaagttggt cgtgaggcac tgggcacgta agtatcaagg
      841 ttacaagaca ggtttaagga gaccaataga aactgggctt gtcgagacag agaagactct
      901 tgcgtttctg ataggcacct attggtctta ctgacatcca ctttgccttt ctctccacag
      961 gtgtccactc ccagttcaat tacagctctt aagaaagcta gccaccatgg actatggcgg
     1021 cgctttgtct gccgtcggac gcgaactttt gttcgttact aatcctgtgg tggtgaacgg
     1081 gtccgtcctg gtccctgagg atcaatgtta ctgtgccgga tggattgaat ctcgcggcac
     1141 gaacggcgct cagaccgcgt caaatgtcct gcagtggctt gcagcaggat tcagcatttt
     1201 gctgctgatg ttctatgcct accaaacctg gaaatctaca tgcggctggg aggagatcta
     1261 tgtgtgcgcc attgaaatgg ttaaggtgat tctcgagttc ttttttgagt ttaagaatcc
     1321 ctctatgctc taccttgcca caggacaccg ggtgcagtgg ctgcgctatg cagagtggct
     1381 gctcacttgt cctgtcatcc ttatccgcct gagcaacctc accggcctga gcaacgacta
     1441 cagcaggaga accatgggac tccttgtctc agacatcggg actatcgtgt ggggggctac
     1501 cagcgccatg gcaaccggct atgttaaagt catcttcttt tgtcttggat tgtgctatgg
     1561 cgcgaacaca ttttttcacg ccgccaaagc atatatcgag ggttatcata ctgtgccaaa
     1621 gggtcggtgc cgccaggtcg tgaccggcat ggcatggctg tttttcgtga gctggggtat
     1681 gttcccaatt ctcttcattt tggggcccga aggttttggc gtcctgagcg tctatggctc
     1741 caccgtaggt cacacgatta ttgatctgat gagtaaaaat tgttgggggt tgttgggaca
     1801 ctacctgcgc gtcctgatcc acgagcacat attgattcac ggagatatcc gcaaaaccac
     1861 caaactgaac atcggcggaa cggagatcga ggtcgagact ctcgtcgaag acgaagccga
     1921 ggccggagcc gtgccagctg cagccacagt gagcaagggc gaggagctgt tcaccggggt
     1981 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
     2041 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
     2101 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
     2161 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
     2221 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
     2281 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     2341 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     2401 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     2461 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     2521 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     2581 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     2641 cggcatggac gagctgtaca agtccggaga taacggaagc ggagccacta acttctccct
     2701 gttgaaacaa gcaggggatg tcgaagagaa tcccgggcca tctagaatgg cccccaagaa
     2761 gaagaggaag gtggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     2821 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     2881 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     2941 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     3001 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     3061 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     3121 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     3181 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     3241 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     3301 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     3361 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     3421 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     3481 gtacaagtaa tggcgcgcca atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3541 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     3601 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     3661 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     3721 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     3781 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     3841 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaagctgac
     3901 gtcctttcca tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
     3961 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     4021 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     4081 ctccccgcct ggatcctgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     4141 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     4201 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     4261 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     4321 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     4381 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     4441 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     4501 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     4561 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     4621 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     4681 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4741 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4801 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4861 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     4921 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     4981 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     5041 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     5101 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     5161 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     5221 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     5281 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     5341 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     5401 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     5461 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     5521 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     5581 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     5641 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     5701 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     5761 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5821 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5881 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5941 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6001 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     6061 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     6121 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     6181 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     6241 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     6301 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     6361 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     6421 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     6481 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     6541 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     6601 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     6661 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     6721 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     6781 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     6841 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     6901 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     6961 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     7021 tacggttcct ggccttttgc tggccttttg ctcacatgt
//
