LOCUS       sequence-256078-        7499 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Double floxed soma-targeted ChRmine-eYFP under the control of Ef1a 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7499)
  AUTHORS   Marshel JH, Kim YS, Machado TA, Quirin S, Benson B, Kadmon J, Raja 
            C, Chibukhchyan A, Ramakrishnan C, Inoue M, Shane JC, McKnight DJ, 
            Yoshizawa S, Kato HE, Ganguli S, Deisseroth K
  TITLE     Cortical layer-specific critical dynamics triggering perception.
  JOURNAL   Science. 2019 Aug 9;365(6453). pii: science.aaw5202. doi: 
            10.1126/science.aaw5202. Epub 2019 Jul 18.
  PUBMED    31320556
REFERENCE   2  (bases 1 to 7499)
  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: "Science."; 
            date: "2019-08-9"; volume: "365(6453). pii: science.aaw5202. doi"; 
            pages: " 10.1126/science.aaw5202. Epub 2019 Jul 18"
COMMENT     Sequence Label: pAAV-Ef1a-DIO-ChRmine-eYFP-Kv2.1-WPRE
FEATURES             Location/Qualifiers
     source          1..7499
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(41..62)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     253..272
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   434..574
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   445..574
                     /label=AAV2 ITR
     polyA_signal    614..1090
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     872..891
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    1122..1710
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1228..1239
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1637..1657
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(1735..1768)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(1819..1852)
                     /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)."
     regulatory      1857..1866
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2859..2861,2862..3572)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2901..2922)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3162..3181)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3509..3530
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    3803..3836
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3887..3920
                     /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)."
     promoter        complement(3927..5105)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3936..4874
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(3959..3979)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     repeat_region   5206..5335
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(5397..5985)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5477..5496)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6156..6947,6948..7016))
                     /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     6779..6798
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7017..7121)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7189..7207
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7245..7267)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7367..7386
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(join(7403..7499,1..359))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 taaatcaaaa gaatagcccg agatagagtt gagtgttgtt ccagtttgga acaagagtcc
       61 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
      121 cccactacgt gaaccatcac ccaaatcaag ttttttgggg tcgaggtgcc gtaaagcact
      181 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
      241 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct aaggcgctgg caagtgtagc
      301 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgta
      361 ctatggttgc tttgacgtat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
      421 cgcatcaggc gcccctgcag gcagctgcgc gctcgctcgc tcactgaggc cgcccgggca
      481 aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga
      541 gagggagtgg ccaactccat cactaggggt tcctgcggcc gctcggtccg cacgtggtta
      601 cctacaaaat cagaaggaca gggaagggag cagtggttca cgcctgtaat cccagcaatt
      661 tgggaggcca aggtgggtag atcacctgag attaggagtt ggagaccagc ctggccaata
      721 tggtgaaacc ccgtctctac caaaaaaaca aaaattagct gagcctggtc atgcatgcct
      781 ggaatcccaa caactcggga ggctgaggca ggagaatcgc ttgaacccag gaggcggaga
      841 ttgcagtgag ccaagattgt gccactgcac tccagcttgg ttcccaatag accccgcagg
      901 ccctacaggt tgtcttccca acttgcccct tgctccatac cacccccctc caccccataa
      961 tattatagaa ggacacctag tcagacaaaa tgatgcaact taattttatt aggacaaggc
     1021 tggtgggcac tggagtggca acttccaggg ccaggagagg cactggggag gggtcacagg
     1081 gatgccaccc gtagatctct cgagcagcgc tcggtatcga tgcggggagg cggcccaaag
     1141 ggagatccga ctcgtctgag ggcgaaggcg aagacgcgga agaggccgca gagccggcag
     1201 caggccgcgg gaaggaaggt ccgctggatt gagggccgaa gggacgtagc agaaggacgt
     1261 cccgcgcaga atccaggtgg caacataggc gagcagccaa ggaaaggacg atgatttccc
     1321 cgacaacacc acggaattgt cagtgcccaa cagccgagcc cctgtccagc agcgggcaag
     1381 gcaggcggcg atgagttccg ccgtggcaat agggaggggg aaagcgaaag tcccggaaag
     1441 gagctgacag gtggtggcaa tgccccaacc agtgggggtt gcgtcagcaa acacagtgca
     1501 caccacgcca cgttgcctga caacgggcca caactcctca taaagagaca gcaaccagga
     1561 tttatacaag gaggagaaaa tgaaagccat acgggaagca atagcatgat acaaaggcat
     1621 taaagcagcg tatccacata gcgtaaaagg agcaacatag ttaagaatac cagtcaatct
     1681 ttcacaaatt ttgtaatcca gaggttgatt atcgataagc ttgatatcga attcataact
     1741 tcgtatagca tacattatac gaagttattc tttgcaccat tctaaagaat aacagtgata
     1801 atttctgggt taaggcaaat aacttcgtat aggatacttt atacgaagtt atgctagcca
     1861 ccatggcaca cgcaccaggc accgaccaga tgttctacgt gggcacaatg gacggctggt
     1921 atctggatac caagctgaac tccgtggcca tcggcgccca ctggtcttgc tttatcgtgc
     1981 tgacaatcac cacattctac ctgggctatg agtcctggac atctcggggc cctagcaaga
     2041 gaacctcctt ttacgccggc tatcaggagg agcagaacct ggccctgttt gtgaatttct
     2101 ttgccatgct gagctacttc ggcaagatcg tggccgacac actgggccac aacttcggcg
     2161 atgtgggccc cttcatcatc ggcttcggca attacaggta tgccgactac atgctgacct
     2221 gcccaatgct ggtgtacgat ctgctgtatc agctgagggc cccctatcgc gtgtcttgta
     2281 gcgccatcat ctttgccatc ctgatgtctg gcgtgctggc cgagttctac gcagagggcg
     2341 accctaggct gaggaatggc gcctacgcct ggtatggctt tggctgtttc tggtttatct
     2401 tcgcctactc tatcgtgatg agcatcgtgg ccaagcagta tagccggctg gcccagctgg
     2461 cccaggatac aggcgccgag cacagcctgc acgtgctgaa gttcgccgtg tttaccttct
     2521 ccatgctgtg gattctgttt cccctggtgt gggccatctg ccctagaggc ttcggctgga
     2581 tcgacgataa ctggacagag gtggcccact gcgtgtgcga catcgtggcc aagtcctgtt
     2641 acggctttgc cctggcccgg ttcagaaaga cctatgatga ggagctgttt cggctgctgg
     2701 agcagctggg acacgacgag gatgagttcc agaagctgga gctggatatg aggctgagca
     2761 gcaatggcga gcgcctgcgg agactgtctg cggccgccaa gagcaggatc accagcgagg
     2821 gcgagtacat ccccctggac cagatcgaca tcaacgtggt gagcaagggc gaggagctgt
     2881 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     2941 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
     3001 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccttc ggctacggcc
     3061 tgcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     3121 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     3181 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     3241 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     3301 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     3361 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     3421 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagctaccag tccgccctga
     3481 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     3541 ggatcactct cggcatggac gagctgtaca agcagtccca gcccatcctc aacaccaagg
     3601 agatggcccc gcagagcaag cctccagagg agctggagat gagcagcatg cccagccccg
     3661 tggcccctct gcccgcacgc acggagggcg tcatcgacat gcggagcatg tccagcattg
     3721 acagcttcat cagctgtgcc acggacttcc ctgaagccac cagattcttc tgctacgaga
     3781 acgaggtgta atagggcgcg ccataacttc gtataatgta tgctatacga agttattaag
     3841 aggtttcata ttgctaatag cagctacaat ccagctacca ttctgcataa cttcgtataa
     3901 agtatcctat acgaagttat ggatcctcac gacacctgaa atggaagaaa aaaactttga
     3961 accactgtct gaggcttgag aatgaaccaa gatccaaact caaaaagggc aaattccaag
     4021 gagaattaca tcaagtgcca agctggccta acttcagtct ccacccactc agtgtgggga
     4081 aactccatcg cataaaaccc ctccccccaa cctaaagacg acgtactcca aaagctcgag
     4141 aactaatcga ggtgcctgga cggcgcccgg tactccgtgg agtcacatga agcgacggct
     4201 gaggacggaa aggccctttt cctttgtgtg ggtgactcac ccgcccgctc tcccgagcgc
     4261 cgcgtcctcc attttgagct ccctgcagca gggccgggaa gcggccatct ttccgctcac
     4321 gcaactggtg ccgaccgggc cagccttgcc gcccagggcg gggcgataca cggcggcgcg
     4381 aggccaggca ccagagcagg ccggccagct tgagactacc cccgtccgat tctcggtggc
     4441 cgcgctcgca ggccccgcct cgccgaacat gtgcgctggg acgcacgggc cccgtcgccg
     4501 cccgcggccc caaaaaccga aataccagtg tgcagatctt ggcccgcatt tacaagacta
     4561 tcttgccaga aaaaaagcgt cgcagcaggt catcaaaaat tttaaatggc tagagactta
     4621 tcgaaagcag cgagacaggc gcgaaggtgc caccagattc gcacgcggcg gccccagcgc
     4681 ccaggccagg cctcaactca agcacgaggc gaaggggctc cttaagcgca aggcctcgaa
     4741 ctctcccacc cacttccaac ccgaagctcg ggatcaagaa tcacgtactg cagccaggtg
     4801 gaagtaattc aaggcacgca agggccataa cccgtaaaga ggccaggccc gcgggaacca
     4861 cacacggcac ttacctgtgt tctggcggca aacccgttgc gaaaaagaac gttcacggcg
     4921 actactgcac ttatatacgg ttctccccca ccctcgggaa aaaggcggag ccagtacacg
     4981 acatcacttt cccagtttac cccgcgccac cttctctagg caccggttca attgccgacc
     5041 cctcccccca acttctcggg gactgtgggc gatgtgcgct ctgcccactg acgggcaccg
     5101 gagccaattc ccactccttt caagacctag aaggtccatt agctgcaaag attcctctct
     5161 gtttaaaact ttatccatct ttgcaaagct tacgcgtgcg gccgcaggaa cccctagtga
     5221 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg
     5281 tcgcccgacg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggacatg
     5341 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5401 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5461 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5521 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5581 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5641 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5701 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5761 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5821 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5881 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     5941 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     6001 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     6061 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     6121 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     6181 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6241 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6301 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6361 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6421 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6481 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6541 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6601 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6661 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6721 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6781 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6841 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6901 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6961 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     7021 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7081 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     7141 ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc
     7201 acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     7261 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag
     7321 ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag
     7381 attgtactga gagtgcacca taaaattgta aacgttaata ttttgttaaa attcgcgtta
     7441 aatttttgtt aaatcagctc attttttaac caatagaccg aaatcggcaa aatccctta
//
