LOCUS       LZF1827_pAAV-hSy        6538 bp ds-DNA     circular SYN 26-OCT-2019
DEFINITION  .
ACCESSION   urn.local...c-a9zsljg (modified)
VERSION     .
KEYWORDS    addgene-plasmid-105677-sequence-204352.
SOURCE      null
  ORGANISM  .
            .
REFERENCE   1  (bases 1 to 6446)
  AUTHORS   Mahn, et. and al.
  TITLE     High-efficiency optogenetic silencing with soma-targeted
            anion-conducting channelrhodopsins
  JOURNAL   BiorXiv
REFERENCE   2  (bases 1 to 6446)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 25, 2018 from SnapGene Viewer 3.1.2
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     repeat_region   1..130
                     /note="Functional equivalent of wild-type AAV2 ITR"
                     /label="AAV2 ITR (alternate)"
     source          1..6538
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        175..622
                     /note="hSyn promoter"
                     /note="human synapsin I promoter; confers neuron-specific
                     expression (Kugler et al., 2003)"
     misc_feature    610..>634
                     /created_by="User"
                     /label="bb_overlap"
     primer_bind     610..699
                     /created_by="kathrynevans"
                     /modified_by="kathrynevans"
                     /Sequence="CCTGAGAGCGCAGTCGAGAAGGTACCGAAGTTCCTATTCTCTAGAAA
                     GTATAGGAACTTCGTACCTACCGTTCGTATAGCATACATTATA"
                     /modified_by="User"
                     /label="KEE_004_F"
     misc_feature    630..635
                     /Recognition_pattern="G^GTACC"
                     /annotation_group="GGTACC"
                     /label="Acc65I, Asp718I"
                     /note="Geneious type: restriction site"
     misc_feature    630..635
                     /Recognition_pattern="GGTAC^C"
                     /annotation_group="GGTACC"
                     /label="KpnI"
                     /note="Geneious type: restriction site"
     misc_feature    632..670
                     /Original_Bases
                     /label
                     /note="Geneious type: Editing History Insertion"
     source          <636..>669
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    636..669
                     /ApEinfo_fwdcolor="#7eff74"
                     /ApEinfo_revcolor="#7eff74"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /label="FRT"
     protein_bind    <675..708
                     /bound_moiety="Cre recombinase"
                     /note="Cre recombinase lox71"
                     /note="loxP mutant. Cre-mediated recombination occurs in 
                     the 8-bp core sequence (GCATACAT)."
                     /label="lox71"
     misc_feature    685..729
                     /created_by="User"
                     /modified_by="User"
                     /label="LZF1825_F1_59"
     misc_feature    709..714
                     /Recognition_pattern="G^AATTC"
                     /Methylation="CpG:variable, EcoBI:variable"
                     /annotation_group="GAATTC"
                     /label="EcoRI"
                     /note="Geneious type: restriction site"
     misc_feature    complement(718..1626)
                     /created_by="User"
                     /modified_by="kathrynevans"
                     /label="CFP_Kv2.1"
     misc_feature    complement(718..912)
                     /created_by="User"
                     /label="Kv2.1"
     misc_feature    1602..1642
                     /created_by="User"
                     /label="LZF1825_F2"
     misc_feature    complement(1611..1626)
                     /created_by="User"
                     /modified_by="User"
                     /label="LZF1825_R1_63"
     misc_feature    complement(1696..2568)
                     /created_by="User"
                     /modified_by="kathrynevans"
                     /label="GtACR2"
     misc_feature    1857..1862
                     /Recognition_pattern="CA^TATG"
                     /Methylation="EcoBI:cut"
                     /annotation_group="CATATG"
                     /label="NdeI"
                     /note="Geneious type: restriction site"
     misc_feature    complement(1879..1899)
                     /created_by="User"
                     /modified_by="User"
                     /label="LZF1825_R2_63"
     regulatory      2565..2574
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong
                     initiationof translation (Kozak, 1987)"
     misc_feature    2565..2570
                     /Recognition_pattern="C^CATGG"
                     /annotation_group="CCATGG"
                     /label="NcoI"
                     /note="Geneious type: restriction site"
     misc_feature    2565..2570
                     /Recognition_pattern="C^CATGG"
                     /annotation_group="CCATGG"
                     /label="NcoI, Bsp19I"
                     /note="Geneious type: restriction site"
     misc_feature    2575..2580
                     /Recognition_pattern="G^GATCC"
                     /Methylation="Dam:cut, Dcm:cut, CpG:cut"
                     /annotation_group="GGATCC"
                     /label="BamHI"
                     /note="Geneious type: restriction site"
     misc_feature    2575..2580
                     /Recognition_pattern="G^GATCC"
                     /annotation_group="GGATCC"
                     /label="BamHI"
                     /note="Geneious type: restriction site"
     protein_bind    complement(2581..2614)
                     /bound_moiety="Cre recombinase"
                     /note="Cre recombinase lox66"
                     /note="loxP mutant. Cre-mediated recombination occurs in 
                     the 8-bp core sequence (GCATACAT)."
                     /label="lox66"
     primer_bind     complement(2598..2682)
                     /created_by="kathrynevans"
                     /Sequence="GTAATCCAGAGGTTGATTATCGATAGAAGTTCCTATACTTTCTAGAG
                     AATAGGAACTTCGATAAGCTTATAACTTCGTATAGCAT"
                     /modified_by="User"
                     /label="KEE_004_R"
     primer_bind     complement(2598..2625)
                     /Hairpin_Tm="51.8"
                     /Self_Dimer_Tm="24.1"
                     /created_by="primer3"
                     /Sequence="TCGATAAGCTTATAACTTCGTATAGCAT"
                     /Hairpin_Tm="None"
                     /Tm="58.6"
                     /Self_Dimer_Tm="19.3"
                     /%GC="32.1"
                     /created_by="primer3"
                     /modified_by="User"
                     /label="insert sequecing primer"
     misc_feature    2615..2620
                     /Original_Bases
                     /label
                     /note="Geneious type: Editing History Insertion"
     source          <2615..>2620
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          <2615..>2619
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    2615..2620
                     /Recognition_pattern="A^AGCTT"
                     /annotation_group="AAGCTT"
                     /label="HindIII"
                     /note="Geneious type: restriction site"
     misc_feature    2622..2656
                     /Original_Bases
                     /label
                     /note="Geneious type: Editing History Insertion"
     source          <2624..>2657
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          join(<2624..>2657,<2624..>2657)
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    2624..2657
                     /ApEinfo_fwdcolor="#7eff74"
                     /ApEinfo_revcolor="#7eff74"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}
                     width 5 offset 0"
                     /label="FRT"
     primer_bind     2658..2682
                     /Hairpin_Tm="38.4"
                     /Self_Dimer_Tm="1.7"
                     /created_by="primer3"
                     /Sequence="TATCGATAATCAACCTCTGGATTAC"
                     /Hairpin_Tm="None"
                     /Tm="56.1"
                     /Self_Dimer_Tm="None"
                     /%GC="36.0"
                     /created_by="primer3"
                     /modified_by="User"
                     /label="bb_overlap"
     misc_feature    2665..3253
                     /note="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2718..2738)
                     /note="WPRE, reverse primer"
                     /label="WPRE-R"
     CDS             complement(3136..3147)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /note="Factor Xa site"
                     /translation="IEGR"
                     /label="Factor Xa site"
     misc_feature    3277..3282
                     /Recognition_pattern="A^GATCT"
                     /Methylation="Dam:cut, EcoBI:some blocked"
                     /annotation_group="AGATCT"
                     /label="BglII"
                     /note="Geneious type: restriction site"
     polyA_signal    3285..3761
                     /note="human growth hormone polyadenylation signal"
                     /label="hGH poly(A) signal"
     primer_bind     complement(3484..3503)
                     /note="Human growth hormone terminator, reverse primer"
                     /label="hGH-PA-R"
     misc_feature    3773..3779
                     /Recognition_pattern="G^GTNACC"
                     /Methylation="Dcm:cut, CpG:cut, EcoBI:impaired,
                     EcoKI:impaired"
                     /annotation_group="GGTNACC"
                     /label="BstEII"
                     /note="Geneious type: restriction site"
     repeat_region   3801..3941
                     /note="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus
                     serotype 2"
                     /label="AAV2 ITR"
     repeat_region   3801..3930
                     /note="AAV2 ITR"
                     /label="AAV2 ITR"
     rep_origin      4016..4471
                     /direction="RIGHT"
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /label="f1 ori"
     primer_bind     complement(4103..4122)
                     /note="F1 origin, reverse primer"
                     /label="F1ori-R"
     primer_bind     4313..4334
                     /note="F1 origin, forward primer"
                     /label="F1ori-F"
     primer_bind     complement(4488..4507)
                     /note="pRS-marker"
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
                     /label="pRS-marker"
     primer_bind     4607..4629
                     /note="pGEX vectors, reverse primer"
                     /label="pGEX 3'"
     primer_bind     complement(4667..4685)
                     /note="pBRforEco"
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
                     /label="pBRforEco"
     promoter        4753..4857
                     /gene="bla"
                     /note="bla promoter"
                     /label="AmpR promoter"
     CDS             4858..5718
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="bla"
                     /note="confers resistance to ampicillin, carbenicillin,
                     and related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW"
                     /label="AmpR"
     primer_bind     complement(5076..5095)
                     /note="Ampicillin resistance gene, reverse primer"
                     /label="Amp-R"
     misc_feature    5273..5278
                     /Recognition_pattern="CGAT^CG"
                     /Methylation="Dam:cut, CpG:blocked"
                     /annotation_group="CGATCG"
                     /label="PvuI"
                     /note="Geneious type: restriction site"
     rep_origin      5889..6477
                     /direction="RIGHT"
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /label="ori"
     primer_bind     6378..6397
                     /note="pBR322 origin, forward primer"
                     /label="pBR322ori-F"
ORIGIN      
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtgtgtct agactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc agtcgagaag gtaccgaagt tcctattctc tagaaagtat
      661 aggaacttcg tacctaccgt tcgtatagca tacattatac gaagttatga attcttagaa
      721 tctggtggct tcagggaagt ccgtggcaca gctgatgaag ctgtcaatgc tggacatgct
      781 ccgcatgtcg atgacgccct ccgtgcgtgc gggcagaggg gccacggggc tgggcatgct
      841 gctcatctcc agctcctctg gaggcttgct ctgcggggcc atctccttgg tgttgaggat
      901 gggctgggac tgcttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa
      961 ctccagcagg accatgtgat cgcgcttctc gttggggtct ttgctcaggg cggactgggt
     1021 gctcaggtag tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg
     1081 gtagtggtcg gcgagctgca cgctgccgtc ctcgatgttg tggcggatct tgaagtgggc
     1141 cttgatgccg ttcttctgct tgtcggcggt gatatagacg ttgtggctga tgtagttgta
     1201 ctccagcttg tgccccagga tgttgccgtc ctccttgaag tcgatgccct tcagctcgat
     1261 gcggttcacc agggtgtcgc cctcgaactt cacctcggcg cgggtcttgt agttgccgtc
     1321 gtccttgaag aagatggtac gctcctggac gtagccttcg ggcatggcgg acttgaagaa
     1381 gtcgtgctgc ttcatgtggt cggggtagcg gctgaagcac tgcacgcccc aggtcagggt
     1441 ggtcacgagg gtgggccagg gcacgggcag cttgccggtg gtgcagatga acttcagggt
     1501 cagcttgccg taggtggcat cgccctcgcc ctcgccggac acgctgaacc tgtggccgtt
     1561 tacgtcgccg tccagctcga ccaggatggg caccaccccg gtgaacagct cctcgccctt
     1621 gctcaccacc acgttgatgt cgatctggtc cagggggatg tactcgccct cgctggtgat
     1681 cctgctcttg gcggcccttc caaacattct gatattgact tcgtcgttag gaggggcatt
     1741 cttctcgtga ttcaggccaa aaggatcgtc ggcttcgggt ttctcctgcc ctggcagaca
     1801 tgcggggtcc cacttccctt ccagcactcc ccaggcagtg ctccacagga tcactccata
     1861 tgtatttttg cacaggccgt cagccagcag gtacagaatg gcgctcacgt tctcgtggat
     1921 cacgcagact ccggtgggtg aaatcagcca cacgattggg aacatagtcc aagcaataaa
     1981 gaagatcagg cgcagcatct tgatcctcac caccaccttc tgtgccagag gtgtcccgac
     2041 agattcaaag ccatacagcc ccacttgaaa gattgtgaag accacactgt actcgaacac
     2101 caggcaggtg gccccaaaga aaaagaacag ccacttcata tactctgcgg gagaaattgt
     2161 agcggtgatt cccatgacca ctctaatgat ggaagcggcc tgtgcaatgg gattcagtgg
     2221 gatacttttg tacttcacca gagccatgtt agaaatctgt cccagcatga ttggacaggt
     2281 gcacagccag gaggccattc ggctccaagg aaagaatctg ccgtccctca tctgcagggt
     2341 tccgttgcca gtactggcca gtgcataagt gatagattcg acaaagggca ggtacacgga
     2401 ctcccaaggg accttgggag ccctgctaat catgaagatt gtctggcagc ctgagaccac
     2461 ggcccacacc agctgcagca gtgagacgaa ggtggagtcg attctggaca tattgtagca
     2521 ctcggtgtga gttgatgccc attccccata gacgacctgt gatgccatgg tggcggatcc
     2581 taccgttcgt ataatgtatg ctatacgaag ttataagctt atcgaagttc ctattctcta
     2641 gaaagtatag gaacttctat cgataatcaa cctctggatt acaaaatttg tgaaagattg
     2701 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     2761 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     2821 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     2881 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     2941 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     3001 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     3061 tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg cgggacgtcc
     3121 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     3181 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     3241 gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg catccctgtg
     3301 acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc accagccttg
     3361 tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat aatattatgg
     3421 ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt agggcctgcg
     3481 gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact gcaatctccg
     3541 cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga ttccaggcat
     3601 gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc accatattgg
     3661 ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct cccaaattgc
     3721 tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg taggtaacca
     3781 cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg
     3841 cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg
     3901 ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt
     3961 tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg
     4021 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     4081 cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     4141 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     4201 ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg
     4261 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     4321 ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga tttataaggg
     4381 attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     4441 aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct
     4501 gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     4561 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     4621 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc
     4681 ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt
     4741 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     4801 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     4861 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     4921 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     4981 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     5041 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     5101 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     5161 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     5221 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     5281 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     5341 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     5401 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     5461 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     5521 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     5581 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     5641 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     5701 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     5761 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     5821 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     5881 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     5941 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     6001 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     6061 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     6121 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     6181 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     6241 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     6301 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     6361 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     6421 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     6481 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgt
//
