LOCUS       Exported                6016 bp ds-DNA     linear   SYN 25-MAY-2021
DEFINITION  Bicistronic vector expressing mRuby2 and GCaMP6f from a single open 
            reading frame. .
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-hSyn1-mRuby2-GSG-P2A-GCaMP6f-WPRE-pA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6016)
  AUTHORS   Rose T, Jaepel J, Hubener M, Bonhoeffer T
  TITLE     Cell-specific restoration of stimulus preference after monocular 
            deprivation in the visual cortex.
  JOURNAL   Science. 2016 Jun 10;352(6291):1319-22. doi: 
            10.1126/science.aad3358.
  PUBMED    27284193
REFERENCE   2  (bases 1 to 6016)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 25, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6016
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      complement(70..658)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(150..169)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(829..1689)
                     /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     1452..1471
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1690..1794)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1820..2275)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1957..1978)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2169..2188
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2347..2476
                     /label=AAV2 ITR
     promoter        2558..3005
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      3041..3050
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3047..3757
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     CDS             3767..3823
                     /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             3824..5173
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="GSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNKT
                     GHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGPV
                     LLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEEL
                     FTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTY
                     GVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIEL
                     KGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRSL
                     GQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGNG
                     YISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      3917..3926
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     primer_bind     4207..4228
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4334..4355)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4595..4614)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    5198..5786
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5251..5271)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5669..5680)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5789..5805)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5790..5806)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    5835..5956
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5846..5865)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5900..5919
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
       61 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
      121 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
      181 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
      241 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
      301 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
      361 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
      421 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
      481 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
      541 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
      601 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
      661 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
      721 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
      781 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
      841 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
      901 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
      961 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     1021 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     1081 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     1141 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     1201 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     1261 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     1321 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     1381 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     1441 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     1501 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     1561 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     1621 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     1681 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     1741 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     1801 tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     1861 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     1921 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     1981 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     2041 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     2101 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     2161 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     2221 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat
     2281 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     2341 cgccagctgc gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg
     2401 acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc
     2461 atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca
     2521 tgctctagga agatctctgc agagggccct gcgtatgagt gcaagtgggt tttaggacca
     2581 ggatgaggcg gggtgggggt gcctacctga cgaccgaccc cgacccactg gacaagcacc
     2641 caacccccat tccccaaatt gcgcatcccc tatcagagag ggggagggga aacaggatgc
     2701 ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac agtgccttcg cccccgcctg
     2761 gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct gacgtcactc gccggtcccc
     2821 cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc cgccgccggc ccagccggac
     2881 cgcaccacgc gaggcgcgag ataggggggc acgggcgcga ccatctgcgc tgcggcgccg
     2941 gcgactcagc gctgcctcag tctgcggtgg gcagcggagg agtcgtgtcg tgcctgagag
     3001 cgcagtcgaa ttcaagctgc tagctagtac cggtcctagg gccaccatgg tgtctaaggg
     3061 cgaagagctg atcaaggaaa atatgcgtat gaaggtggtc atggaaggtt cggtcaacgg
     3121 ccaccaattc aaatgcacag gtgaaggaga aggcaatccg tacatgggaa ctcaaaccat
     3181 gaggatcaaa gtcatcgagg gaggacccct gccatttgcc tttgacattc ttgccacgtc
     3241 gttcatgtat ggcagccgta cttttatcaa gtacccgaaa ggcattcctg atttctttaa
     3301 acagtccttt cctgagggtt ttacttggga aagagttacg agatacgaag atggtggagt
     3361 cgtcaccgtc atgcaggaca ccagccttga ggatggctgt ctcgtttacc acgtccaagt
     3421 cagaggggta aactttccct ccaatggtcc cgtgatgcag aagaagacca agggttggga
     3481 gcctaataca gagatgatgt atccagcaga tggtggtctg aggggataca ctcatatggc
     3541 actgaaagtt gatggtggtg gccatctgtc ttgctctttc gtaacaactt acaggtcaaa
     3601 aaagaccgtc gggaacatca agatgcccgg tatccatgcc gttgatcacc gcctggaaag
     3661 gttagaggaa agtgacaatg aaatgttcgt agtacaacgc gaacacgcag ttgccaagtt
     3721 cgccgggctt ggtggtggga tggacgagct gtacaaggga tccggagcca cgaacttctc
     3781 tctgttaaag caagcaggag acgtggaaga aaaccccggt cctggttctc atcatcatca
     3841 tcatcatggt atggctagca tgactggtgg acagcaaatg ggtcgggatc tgtacgacga
     3901 tgacgataag gatctcgcca ccatggtcga ctcatcacgt cgtaagtgga ataagacagg
     3961 tcacgcagtc agagctatag gtcggctgag ctcactcgag aacgtctata tcaaggccga
     4021 caagcagaag aacggcatca aggcgaactt caagatccgc cacaacatcg aggacggcgg
     4081 cgtgcagctc gcctaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     4141 gcccgacaac cactacctga gcgtgcagtc caaactttcg aaagacccca acgagaagcg
     4201 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     4261 gctgtacaag ggcggtaccg gagggagcat ggtgagcaag ggcgaggagc tgttcaccgg
     4321 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     4381 cggcgagggt gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     4441 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     4501 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     4561 aggctacatc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     4621 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     4681 caaggaggac ggcaacatcc tggggcacaa gctggagtac aacctgccgg accaactgac
     4741 tgaagagcag atcgcagaat ttaaagagga attctcccta tttgacaagg acggggatgg
     4801 gacaataaca accaaggagc tggggacggt gatgcggtct ctggggcaga accccacaga
     4861 agcagagctg caggacatga tcaatgaagt agatgccgac ggtgacggca caatcgactt
     4921 ccctgagttc ctgacaatga tggcaagaaa aatgaaatac agggacacgg aagaagaaat
     4981 tagagaagcg ttcggtgtgt ttgataagga tggcaatggc tacatcagtg cagcagagct
     5041 tcgccacgtg atgacaaacc ttggagagaa gttaacagat gaagaggttg atgaaatgat
     5101 cagggaagca gacatcgatg gggatggtca ggtaaactac gaagagtttg tacaaatgat
     5161 gacagcgaag taagcggccg cactagagct tatcgataat caacctctgg attacaaaat
     5221 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     5281 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     5341 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     5401 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     5461 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     5521 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     5581 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct
     5641 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     5701 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     5761 gatctccctt tgggccgcct ccccgcatcg ataccgtcga cctcgacccg ggcggccgct
     5821 tcgagcagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg
     5881 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     5941 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     6001 gatgtgggag gttttt
//
