LOCUS       Exported                7846 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Bicistronic Cre-dependent expression of TRPV4.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-CMV-DIO-TRPV4-p2A-ferritin-sNRPpA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7846)
  AUTHORS   Wheeler MA, Smith CJ, Ottolini M, Barker BS, Purohit AM, Grippo RM, 
            Gaykema RP, Spano AJ, Beenhakker MP, Kucenas S, Patel MK, Deppmann 
            CD, Guler AD
  TITLE     Genetically targeted magnetic control of the nervous system.
  JOURNAL   Nat Neurosci. 2016 Mar 7. doi: 10.1038/nn.4265.
  PUBMED    26950006
REFERENCE   2  (bases 1 to 7846)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7846
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     enhancer        207..510
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        511..714
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     665..685
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    777..810
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    861..894
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             complement(942..965)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(1560..1583)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(2154..2177)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(2211..2267)
                     /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             complement(2283..2306)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      4916..4925
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    complement(4938..4971)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(5022..5055)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     repeat_region   5109..5249
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5109..5238
                     /label=AAV2 ITR
     rep_origin      5324..5779
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5411..5430)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5621..5642
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5796..5815)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5915..5937
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5975..5993)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6061..6165
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6166..7026
                     /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(6384..6403)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7197..7785
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7686..7705
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgcctagt aatcaattac ggggtcatta
      181 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      241 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      301 ccaataggga ctttccattg acgtcaatgg gtggactatt tacggtaaac tgcccacttg
      361 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      421 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      481 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
      541 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
      601 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
      661 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg
      721 ctaactagag aacccactgc ttactggctt atcgaaattc tagagtcgac tccggaataa
      781 cttcgtatag gatactttat acgaagttat gcagaatggt agctggattg tagctgctat
      841 tagcaatatg aaacctctta ataacttcgt atagcataca ttatacgaag ttatggcgcg
      901 ccggatcctt agctagccac tcgggaggag cctccacccc ctttgtcatc gtcgtcttta
      961 tagtcgctcc cgccaccccc gcgtgcgctt tcattatcac tgtctcccag ggtgtgcttg
     1021 tcaaagagat attccgccaa gccagattcg ggcgctccca tcttgcgcaa gttggtcacg
     1081 tggtcaccca attctttgat ggctttcacc tgctcattca ggtaatgtgt ctcaatgaag
     1141 tcacacaaat gggggtcatt tttgtcagtg gccagtttgt gcagttccag tagtgactga
     1201 ttcacatttt tttccaaatg taatgcacac tccattgcat tcagcccgct ctcccagtca
     1261 tcacagtctg gtttcttgat atcctgaagg aagattcggc cacctcgttg gttctgcagc
     1321 ttcatcagtt tctcagcatg ttccctctcc tcatgagatt ggtgaagaaa gtatttggca
     1381 aagttcttca aagccacatc atcgcggtca aagtagtaag acatggacag gtaaacgtag
     1441 gaggcgtaga gctccaggtt gatctggcgg ttgatggcgg cctctgagtc ctggtggtag
     1501 ttctggcgca cctgcgaggt ggacgcggtc gtcatgacgc gggaggagcc gccgccgcct
     1561 ttatcatcat catctttata atcggagccg ccgccgccgc gggcgtcgtg cttgagagtg
     1621 agcctttcga agagatactc gcccagccca gcctccgggc cacccagcct gtggaggttg
     1681 gtcaggtggt cacccatctt cttgataagc ttcacttcct catctaggaa gtgagtctcc
     1741 aggaagtcac agagatgggg gtccgtgcgg gcagaaccca gggcatgaag atccaaaagg
     1801 gcctggttca gctttttctc cagggccatg gcagctttca tggcgtctgg ggttttaccc
     1861 cactcatctt cagctggctt cttgatgtcc tggaagagag cgcggccgcc acgctggttt
     1921 tgcatcttca ggagacgctc gtagccctcg cgcttctcct cggccaattc gcggaagaag
     1981 tggctcacgc cttccagagc cacatcatcg cggtcgaaat agaagcccag agagaggtag
     2041 gtgtaggagg cctgcaggta caaattgacc aggctgttga cggctgcctc cacgtcggtg
     2101 gaataattct gacgaatctg ggagcccata acccgagagg agccgcctcc tcctttgtcg
     2161 tcgtcgtcct tataatcaga accgcccccg ccccgtgctc cggactcgag ggggccgggg
     2221 ttctcctcca cgtcgccggc ctgcttcagc aggctgaagt tggtggcgcc gctgccggat
     2281 cccttatcgt catcgtcctt gtaatccagt ggtgcgtcct ccgccctcca cttgggagca
     2341 taaccttgct ggtggccgtc acagttgggg ttccctaggt tatccagggg gaccaccact
     2401 tcatctgtgc ctgagttctt gttcagctcc accacgcggg gcaccactga ggaccagcga
     2461 tccctgcgga ggcgccccat ggtatgggag aagccatagt actggtagat ctcgctcttg
     2521 ccggggtcct cgttaatgat gcccaggttc tggttccagt gagaccagtt cacctcgtcc
     2581 accctgaagc accacctgcg gtctggagtg ccatccgagc tcttgcccac tgtcaccatc
     2641 tctccggagc ggaaggcctt cctcaggaac acagggaagg agcgctcgat gtccaggatg
     2701 gtggtggccc actgcagctt ccagatgtgc ttgctctcct tggacacctg gcccacggtc
     2761 tcacccatga gggcgatgag catgttcagc aggagcacga aggtgaggat gatgtaggta
     2821 accagcagga gaatgaagac cacggggtac ttagcgctgc tcagcatctc caggtcgccc
     2881 atgccgatgg tgagcttgaa gaggtccagt aggaaggcgc tgaaggtctc gctgtcccgg
     2941 cacgcggggt atgagggcac cgtgcagttg ctctggtcct cgttacagac cttcatgttg
     3001 gtgcacggat tcaggagggt gaccagagct gaggcatagc caatcataaa aagcaggtag
     3061 accagcagaa agcggaagag atctttgaag aggatcttct gaatcatgat gctgtaggtc
     3121 cctgtcagct tcagcccacg ggtgaagtaa agggcattca tccagcccag gaccagggca
     3181 aagaccatca cagccagata ggcctcgatc cctgccaggt agagcgccgc agacacaacc
     3241 accagcactg agtagatgaa gtagagcaac tggaaggagc catcgacgaa gagagaattc
     3301 actccagggc atttcttcat gaacaagtct ttgatactgg taaagaagaa caggactcct
     3361 gtgaggagcg tgatgacctc accagccagc ctcaggtagt ccaccgtggt acggtaaggg
     3421 tagggtggcg tgccctccag tggctgatag taggctgtga gggtgaagat gaccatggca
     3481 cacagatagg agacaacgtt gatgtagaag gacacggccc cgaacttacg ccacttgtcc
     3541 ctcagcagtt cgttaatggg ctccacagcc agcatctcat ggcggttctc gatcttgctg
     3601 ttgtaaacca ggatctccag cacggacact tcctccccgc acgtatccag ggaggagagg
     3661 tcgtagagag aagaatacac aggcccgtag gcccagtcct tgaacttgcg agacaggtgc
     3721 cgtgtgtcct catctgtcac ctcccgtcgg atgatgtgct gaaagacccc gatcttgcca
     3781 gtcttggcag ccatcatgag gggcgaaaga ccgtcattgt taagcacagt ctccaggttg
     3841 ctgtctggga agaggcggga gcacttgaga agcaacaggt catacatctt ggtgacaaac
     3901 ttggtgttct ctcgggtgtt gtcagcgatg gccaccagcg cgtggagcac cgtgttgcct
     3961 ctggagtcct gtcgcctcat atcggctttc ttgtgagggt tctctgtcag gtagttgacg
     4021 atgtgcggct ggttggtgca ggctgccaag gacaagggca gctccccaaa gtagaagtag
     4081 ccaccctcat ccttgggctg gaagaaccgc cctcgggcct gcgcgtgcac atcggctccc
     4141 tgggccacca ggagctccac gtaatgcttg cagcgccgtt caatggcgat gtgcagtgcc
     4201 gtctgccctc ggtagtagat gtctctgaag ggcgagttga tgaactcccg catgttgccc
     4261 gtgcgttccg caatgtccag caacactggg atggtgtcgt ttcggccatt gcttaagttc
     4321 agaagtgcct tgggcaggca ggtcttccct gtggatggtt cccggaactc ctcatcagtc
     4381 aggcgcttct tgtgggtcag caagtaggag agcagtccgt ccaggtcggc agtggagccc
     4441 cgggacacga tgtcaaagag gatgggccgg ttgaagactt tgaggatggg gggtggctgg
     4501 ggggcgggag ctttggggct ctgtggctgc ttctctacga ccttcctcct ccatctcttg
     4561 ttgtcactgg ggtggtgccg gtaagtgcca tagtcgaaca acgaatccat gggcgctttc
     4621 ttgggcccag gcactactga ggactcatac agggtggact ccagcaggtc aatggggttg
     4681 ggaaccccct tgcggaaagc gccctggaac ttcatacgca ggtttggacg tccatccccg
     4741 gggccagcag ggcggctagc atccactggt gaaagagaag aggagccttc ctctccctca
     4801 aacaggttgg ccagggaaga gagggggaag gcctccccac cagaagtgcc actctcgtct
     4861 ccagggggct cagccacatc cccaggcgct gcacgggggc catcaccagg atctgccatg
     4921 gtggcggtac ctctagcata acttcgtata aagtatccta tacgaagtta tttgccttaa
     4981 cccagaaatt atcactgtta ttctttagaa tggtgcaaag aataacttcg tataatgtat
     5041 gctatacgaa gttatgaatt caaataaaat acgaaatgca cgtaaccacg tgcggaccga
     5101 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     5161 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     5221 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     5281 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     5341 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     5401 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     5461 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     5521 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     5581 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     5641 ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     5701 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     5761 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     5821 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     5881 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     5941 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     6001 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     6061 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     6121 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     6181 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     6241 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     6301 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     6361 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     6421 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     6481 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     6541 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     6601 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     6661 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     6721 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     6781 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     6841 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     6901 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     6961 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     7021 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     7081 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     7141 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     7201 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     7261 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     7321 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     7381 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     7441 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     7501 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     7561 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     7621 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     7681 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     7741 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     7801 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
