LOCUS       sequence-242014-        7308 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent soma-localized ChrimsonR for optogenetic stimulation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7308)
  AUTHORS   Chettih SN, Harvey CD
  TITLE     Single-neuron perturbations reveal feature-specific competition in 
            V1.
  JOURNAL   Nature. 2019 Mar 6. pii: 10.1038/s41586-019-0997-6. doi: 
            10.1038/s41586-019-0997-6.
  PUBMED    30842660
REFERENCE   2  (bases 1 to 7308)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2019 Mar 6. pii: 10.1038/s41586-019-0997-6. doi: 
            10.1038/s41586-019-0997-6."
COMMENT     Sequence Label: pAAV-EF1a-DIO-ChrimsonR-mRuby2-KV2.1-WPRE-SV40
FEATURES             Location/Qualifiers
     source          1..7308
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1408
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1399
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1356..1376
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1439..1472)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1523..1556)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1763..2470)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="VSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTMR
                     IKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGGV
                     VTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTHM
                     ALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAVA
                     KFAGLGGGMDELYK"
     CDS             complement(2480..3529)
                     /codon_start=1
                     /product="red light-drivable channelrhodopsin from 
                     Chlamydomonas noctigama (Klapoetke et al., 2014)"
                     /label=ChrimsonR
                     /note="K176R mutant with accelerated closing kinetics"
                     /translation="MAELISSATRSLFAAGGINPWPNPYHHEDMGCGGMTPTGECFSTE
                     WWCDPSYGLSDAGYGYCFVEATGGYLVVGVEKKQAWLHSRGTPGEKIGAQVCQWIAFSI
                     AIALLTFYGFSAWKATCGWEEVYVCCVEVLFVTLEIFKEFSSPATVYLSTGNHAYCLRY
                     FEWLLSCPVILIRLSNLSGLKNDYSKRTMGLIVSCVGMIVFGMAAGLATDWLKWLLYIV
                     SCIYGGYMYFQAAKCYVEANHSVPKGHCRMVVKLMAYAYFASWGSYPILWAVGPEGLLK
                     LSPYANSIGHSICDIIAKEFWTFLAHHLRIKIHEHILIHGDIRKTTKMEIGGEEVEVEE
                     FVEEEDEDTV"
     regulatory      3526..3535
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3540..3573
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3624..3657
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3682..4270
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3735..3755)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4153..4164)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4315..4436
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4326..4345)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4380..4399
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4571..4711
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4571..4700
                     /label=AAV2 ITR
     rep_origin      4786..5241
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4873..4892)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5083..5104
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5258..5277)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5377..5399
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5437..5455)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5523..5627
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5628..5696,5697..6488)
                     /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(5846..5865)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6659..7247
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7148..7167
                     /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 gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccactg
      541 gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag
      601 gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg
      661 gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc
      721 tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct
      781 tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga
      841 cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc
      901 gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc
      961 gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc
     1021 ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc
     1081 gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt
     1141 cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag
     1201 cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca
     1261 cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga
     1321 atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag
     1381 tttttttctt ccatttcagg tgtcgtgagg taccggatcc tctagagtcg actccggaat
     1441 aacttcgtat aggatacttt atacgaagtt atgcagaatg gtagctggat tgtagctgct
     1501 attagcaata tgaaacctct taataacttc gtatagcata cattatacga agttatggcg
     1561 cgccttagaa tctggtggct tcagggaagt ccgtggcaca gctgatgaag ctgtcaatgc
     1621 tggacatgct ccgcatgtcg atgacgccct ccgtgcgtgc gggcagaggg gccacggggc
     1681 tgggcatgct gctcatctcc agctcctctg gaggcttgct ctgcggggcc atctccttgg
     1741 tgttgaggat gggctgggac tgcttgtaca gctcgtccat cccaccacca agcccggcga
     1801 acttggcaac tgcgtgttcg cgttgtacta cgaacatttc attgtcactt tcctctaacc
     1861 tttccaggcg gtgatcaacg gcatggatac cgggcatctt gatgttcccg acggtctttt
     1921 ttgacctgta agttgttacg aaagagcaag acagatggcc accaccatca actttcagtg
     1981 ccatatgagt gtatcccctc agaccaccat ctgctggata catcatctct gtattaggct
     2041 cccaaccctt ggtcttcttc tgcatcacgg gaccattgga gggaaagttt acccctctga
     2101 cttggacgtg gtaaacgaga cagccatcct caaggctggt gtcctgcatg acggtgacga
     2161 ctccaccatc ttcgtatctc gtaactcttt cccaagtaaa accctcagga aaggactgtt
     2221 taaagaaatc aggaatgcct ttcgggtact tgataaaagt acggctgcca tacatgaacg
     2281 acgtggcaag aatgtcaaag gcaaatggca ggggtcctcc ctcgatgact ttgatcctca
     2341 tggtttgagt tcccatgtac ggattgcctt ctccttcacc tgtgcatttg aattggtggc
     2401 cgttgaccga accttccatg accaccttca tacgcatatt ttccttgatc agctcttcgc
     2461 ccttagacac ggcggccgcc actgtgtcct cgtcctcctc ctccacgaac tcttccactt
     2521 ccacctcctc gcctccgatc tccatcttgg tggtcttccg gatgtcgccg tggatcagga
     2581 tgtgctcgtg gatcttgatc ctcaggtggt gggccaggaa ggtccaaaac tccttggcga
     2641 tgatgtcgca gatgctgtgg ccgatgctgt tggcgtaagg gctcagcttc agcagtcctt
     2701 ctggtcccac tgcccagagg attgggtagc tgccccaaga ggcgaagtaa gcgtaggcca
     2761 tcagcttcac gaccatgcgg caatggcctt taggcacgct gtggttggct tccacgtagc
     2821 acttggcggc ctggaagtac atgtagccgc cgtagatgca agacacgata tacagcagcc
     2881 acttgagcca atcggtagcc agtcctgcgg ccatgccgaa cacgatcatt cccacgcaag
     2941 acacgatcag gcccatggtc cgcttgctgt agtcgttctt caggccgctc aggttgctca
     3001 gtctgatcag gatcacgggg caagacagca gccactcgaa gtagcgcagg caataggcgt
     3061 ggttgccggt agacaggtac actgtggcgg ggctgctgaa ctccttgaag atctccaggg
     3121 tcacgaacag cacctcgacg caacagacgt agacctcctc ccaaccgcaa gtggccttcc
     3181 aggcgctgaa gccgtagaat gtcagcaggg cgatggcgat gctgaaagca atccactggc
     3241 agacctgggc gccgatcttt tctcctggtg tgcctctgct gtgcagccaa gcctgcttct
     3301 tctccactcc cacgaccagg tagccgcctg tggcctccac gaagcaatat ccgtatccgg
     3361 cgtcgctcag tccgtaagaa gggtcacacc accactcggt gctgaagcac tcgcctgtag
     3421 gtgtcattcc tccacagccc atgtcctcgt ggtggtaggg gttaggccaa gggttgatgc
     3481 ctccggcggc aaacagagat ctggtggcgc tgctgatcag ctcagccatg gtggctagca
     3541 taacttcgta taaagtatcc tatacgaagt tatttgcctt aacccagaaa ttatcactgt
     3601 tattctttag aatggtgcaa agaataactt cgtataatgt atgctatacg aagttatgaa
     3661 ttcgatatca agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact
     3721 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     3781 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg
     3841 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg
     3901 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccgga
     3961 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc
     4021 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca
     4081 tcgtcctttc cttggctgct cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc
     4141 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct
     4201 ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc
     4261 gcctccccgc atcgataccg agcgctgctc gagagatctt tcgagcagac atgataagat
     4321 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     4381 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     4441 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa
     4501 gcaagtaaaa cctctacaaa tgtggtaaaa tctgattttg taggtaacca cgtgcggacc
     4561 gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg
     4621 ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca
     4681 gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg
     4741 catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg
     4801 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     4861 ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     4921 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     4981 tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga
     5041 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     5101 ctggaacaac actcaactct atctcgggct attcttttga tttataaggg attttgccga
     5161 tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     5221 aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat
     5281 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     5341 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     5401 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat
     5461 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     5521 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5581 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5641 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5701 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     5761 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     5821 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     5881 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     5941 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     6001 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     6061 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     6121 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     6181 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     6241 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     6301 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     6361 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     6421 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     6481 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     6541 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     6601 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     6661 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     6721 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     6781 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     6841 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     6901 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     6961 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     7021 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     7081 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     7141 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     7201 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     7261 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgt
//
