LOCUS       pOTTC1642_-_pAAV        7197 bp DNA     circular SYN 13-NOV-2023
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7197)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 13, 2023 from SnapGene 7.0.2
            https://www.snapgene.com
COMMENT     pOTTC1642 was constructed by Chris Richie (CR2005_C).
            
            Deposited to Addgene as 195018.
            
            backbone was pOTTC1553 - pAAV EF1a NucEnv-EGFP.ape from 1 to 6326
            pOTTC1553 was constructed by Chris Richie at GEVVC/NIDA (CR1852_D).
            
            
            bakcbone was pOTTC407 - pAAV EF1a eGFP was created by Mari 
            Savolainen on 09/28/2012 (MS08).  Addgene 60058
            Described in Savolainen et al. (2014).  The beneficial effect of a 
            prolyl oligopeptidase inhibitor, KYP-2047, on alpha-synuclein 
            clearance and autophagy in A30P transgenic mouse.  Neurobiology of 
            Disease.  DOI: 10.1016/j.nbd.2014.04.003
FEATURES             Location/Qualifiers
     source          1..7197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4; direction: RIGHT"
     promoter        167..480
                     /label=mU6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    483..502
                     /label=CR1996_mTacr1_A55
                     /note="color: #00ff00"
     misc_RNA        503..578
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
                     /note="color: #00ccff; direction: RIGHT"
     misc_RNA        507..578
                     /label=Constant region for chimeric gRNA
                     /note="color: #800080; direction: RIGHT"
     promoter        609..1872
                     /label=EF-1a promoter
                     /note="color: #408080"
     promoter        671..926
                     /label=EFS (EF1a short) promoter
                     /note="color: #8000ff"
     intron          926..1863
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
                     /note="color: #------"
     misc_feature    990..1019
                     /label=+C in EF1a promoter
                     /note="GAATTACTTCCACTGGCTGCAGTACGTGAT -> GAATTACTTCCAC c 
                     TGGCTGCAGTACGTGAT
                     this is an existing -1nuc SNP in the EF1a promoter of many 
                     GEVVC vectors.
                     stemming from ""founder effect"", likely going back to KD's
                     optogenetic vectors.
                     
                     added to GEVVC Common Features on 10-02-23"
                     /note="color: #a6acb3"
     regulatory      1892..1901
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     CDS             1898..2614
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /note="This feature has 3 segments:
                        1: 1898 .. 1900 / #05fd14
                        2: 1901 .. 1903 / #05fd14 / 1a
                        3: 1904 .. 2614 / #05fd14"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2633..2881
                     /label=KASH domain of Nesprin2
                     /note="/dnas_title=KASH domain of Nesprin2"
                     /note="/vntifkey=4"
                     /note="color: #0080c0; direction: RIGHT"
     misc_RNA        2896..3504
                     /label=WPRE
                     /note="color: #800080; direction: RIGHT"
     polyA_signal    3525..4005
                     /label=hGH polyA
                     /note="color: #8080ff"
     misc_RNA        complement(4059..4134)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
                     /note="color: #00ccff; direction: LEFT"
     protein_bind    complement(4135..4154)
                     /label=CR1996_mTacr1_B108
                     /note="color: #ffa500; direction: LEFT"
     promoter        complement(4156..4421)
                     /locus_tag="hU6 promoter"
                     /label=hU6 promoter
                     /note="Human U6 promoter (polIII)"
                     /note="color: #804000; direction: LEFT"
     repeat_region   complement(4460..4600)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4; direction: LEFT"
     rep_origin      4675..5130
                     /label=M13 origin
                     /note="color: #808080"
     promoter        5412..5516
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             5517..6374
                     /label=AmpR
                     /note="color: #0080c0; direction: RIGHT"
     rep_origin      6472..7154
                     /label=ColE1 origin
                     /note="color: #808080"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtcgtct cagatagatc cgacgccgcc
      181 atctctaggc ccgcgccggc cccctcgcac agacttgtgg gagaagctcg gctactcccc
      241 tgccccggtt aatttgcata taatatttcc tagtaactat agaggcttaa tgtgcgataa
      301 aagacagata atctgttctt tttaatacta gctacatttt acatgatagg cttggatttc
      361 tataagagat acaaatacta aattattatt ttaaaaaaca gcacaaaagg aaactcaccc
      421 taactgtaaa gtaattgtgt gttttgagac tataaatatc ccttggagaa aagccttgtt
      481 tgccgtatag gcggctgccc aagttttaga gctagaaata gcaagttaaa ataaggctag
      541 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt tttttgtact gagacgccgc
      601 ggtcgcgtaa gctttgcaaa gatggataaa gttttaaaca gagaggaatc tttgcagcta
      661 atggaccttc taggtcttga aaggagtggg aattggctcc ggtgcccgtc agtgggcaga
      721 gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gaaccggtgc
      781 ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt
      841 tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg
      901 caacgggttt gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg ggcctggcct
      961 ctttacgggt tatggccctt gcgtgccttg aattacttcc actggctgca gtacgtgatt
     1021 cttgatcccg agcttcgggt tggaagtggg tgggagagtt cgaggccttg cgcttaagga
     1081 gccccttcgc ctcgtgcttg agttgaggcc tggcctgggc gctggggccg ccgcgtgcga
     1141 atctggtggc accttcgcgc ctgtctcgct gctttcgata agtctctagc catttaaaat
     1201 ttttgatgac ctgctgcgac gctttttttc tggcaagata gtcttgtaaa tgcgggccaa
     1261 gatctgcaca ctggtatttc ggtttttggg gccgcgggcg gcgacggggc ccgtgcgtcc
     1321 cagcgcacat gttcggcgag gcggggcctg cgagcgcggc caccgagaat cggacggggg
     1381 tagtctcaag ctggccggcc tgctctggtg cctggcctcg cgccgccgtg tatcgccccg
     1441 ccctgggcgg caaggctggc ccggtcggca ccagttgcgt gagcggaaag atggccgctt
     1501 cccggccctg ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga gcgggcgggt
     1561 gagtcaccca cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc atgtgactcc
     1621 acggagtacc gggcgccgtc caggcacctc gattagttct cgagcttttg gagtacgtcg
     1681 tctttaggtt ggggggaggg gttttatgcg atggagtttc cccacactga gtgggtggag
     1741 actgaagtta ggccagcttg gcacttgatg taattctcct tggaatttgc cctttttgag
     1801 tttggatctt ggttcattct caagcctcag acagtggttc aaagtttttt tcttccattt
     1861 caggtgtcgt gaggtaccgg atccaccggt cgccaccatg gtgagcaagg gcgaggagct
     1921 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     1981 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat
     2041 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg
     2101 cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     2161 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     2221 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     2281 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag
     2341 ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat
     2401 ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
     2461 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct
     2521 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
     2581 cgggatcact ctcggcatgg acgagctgta caagtccgga ctcagatctc gagaggagga
     2641 ggaggagaca gacagcagga tgccccacct cgacagcccc ggcagctccc agccgagacg
     2701 ctccttcctc tcaagggtga tcagggcagc gctaccgttg cagctgcttc tgctgctgct
     2761 gctgctcctg gcctgcctgc tacctgcctc tgaagatgac tacagctgca cccaggccaa
     2821 caactttgcc cgatccttct accccatgct gcggtacacc aacgggccac ctcccaccta
     2881 ggaattcgat atcaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt
     2941 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     3001 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     3061 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     3121 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     3181 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     3241 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     3301 atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc gcgggacgtc
     3361 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     3421 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     3481 ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctacgggtg gcatccctgt
     3541 gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt
     3601 gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta taatattatg
     3661 gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc
     3721 ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc
     3781 gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca
     3841 tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg
     3901 gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg
     3961 ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt gtaggtaacc
     4021 acaccgcggc gtctcagtac gctcagcgga gaaaaaaagc accgactcgg tgccactttt
     4081 tcaagttgat aacggactag ccttatttta acttgctatt tctagctcta aaacctacgt
     4141 tgcccaccac ggaacggtgt ttcgtccttt ccacaagata tataaagcca agaaatcgaa
     4201 atactttcaa gttacggtaa gcatatgata gtccatttta aaacataatt ttaaaactgc
     4261 aaactaccca agaaattatt actttctacg tcacgtattt tgtactaata tctttgtgtt
     4321 tacagtcaaa ttaattccaa ttatctctct aacagccttg tatcgtatat gcaaatatga
     4381 aggaatcatg ggaaataggc cctcttcctg cccgaccttg gatcgatccg cctgtgagac
     4441 gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     4501 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
     4561 gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt
     4621 ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc
     4681 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     4741 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     4801 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     4861 tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc
     4921 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     4981 tgttccaaac tggaacaaca ctcaacccta tctcgggcta ttcttttgat ttataaggga
     5041 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     5101 attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg
     5161 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     5221 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     5281 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     5341 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     5401 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     5461 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     5521 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     5581 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     5641 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     5701 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     5761 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     5821 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     5881 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     5941 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     6001 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     6061 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     6121 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     6181 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     6241 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     6301 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     6361 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     6421 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     6481 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     6541 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     6601 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     6661 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     6721 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     6781 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     6841 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     6901 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     6961 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     7021 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     7081 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     7141 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgt
//
