LOCUS       sequence-256089-        7478 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Double floxed soma-targeted ChRmine-mScarlet under the control of 
            Ef1a promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7478)
  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 7478)
  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-mScarlet-Kv2.1-WPRE
FEATURES             Location/Qualifiers
     source          1..7478
                     /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             2859..3551
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="VSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGGA
                     VTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKM
                     ALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS
                     TGGMDELYK"
     primer_bind     complement(2989..3007)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3257..3276
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    3782..3815
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3866..3899
                     /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(3906..5084)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3915..4853
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(3938..3958)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     repeat_region   5185..5314
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(5376..5964)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5456..5475)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6135..6926,6927..6995))
                     /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     6758..6777
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6996..7100)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7168..7186
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7224..7246)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7346..7365
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(join(7382..7478,1..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 gaggcagtga
     2881 tcaaggagtt catgcggttc aaggtgcaca tggagggctc catgaacggc cacgagttcg
     2941 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
     3001 tgaccaaggg tggccccctg cccttctcct gggacatcct gtcccctcag ttcatgtacg
     3061 gctccagggc cttcaccaag caccccgccg acatccccga ctactataag cagtccttcc
     3121 ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgcc gtgaccgtga
     3181 cccaggacac ctccctggag gacggcaccc tgatctacaa ggtgaagctc cgcggcacca
     3241 acttccctcc tgacggcccc gtaatgcaga agaagacaat gggctgggaa gcgtccaccg
     3301 agcggttgta ccccgaggac ggcgtgctga agggcgacat taagatggcc ctgcgcctga
     3361 aggacggcgg gcgctacctg gcggacttca agaccaccta caaggccaag aagcccgtgc
     3421 agatgcccgg cgcctacaac gtggaccgca agttggacat cacctcccac aacgaggact
     3481 acaccgtggt ggaacagtac gaacgctccg agggccgcca ctccaccggc ggcatggacg
     3541 agctgtacaa gcagtcccag cccatcctca acaccaagga gatggccccg cagagcaagc
     3601 ctccagagga gctggagatg agcagcatgc ccagccccgt ggcccctctg cccgcacgca
     3661 cggagggcgt catcgacatg cggagcatgt ccagcattga cagcttcatc agctgtgcca
     3721 cggacttccc tgaagccacc agattcttct gctacgagaa cgaggtgtaa tagggcgcgc
     3781 cataacttcg tataatgtat gctatacgaa gttattaaga ggtttcatat tgctaatagc
     3841 agctacaatc cagctaccat tctgcataac ttcgtataaa gtatcctata cgaagttatg
     3901 gatcctcacg acacctgaaa tggaagaaaa aaactttgaa ccactgtctg aggcttgaga
     3961 atgaaccaag atccaaactc aaaaagggca aattccaagg agaattacat caagtgccaa
     4021 gctggcctaa cttcagtctc cacccactca gtgtggggaa actccatcgc ataaaacccc
     4081 tccccccaac ctaaagacga cgtactccaa aagctcgaga actaatcgag gtgcctggac
     4141 ggcgcccggt actccgtgga gtcacatgaa gcgacggctg aggacggaaa ggcccttttc
     4201 ctttgtgtgg gtgactcacc cgcccgctct cccgagcgcc gcgtcctcca ttttgagctc
     4261 cctgcagcag ggccgggaag cggccatctt tccgctcacg caactggtgc cgaccgggcc
     4321 agccttgccg cccagggcgg ggcgatacac ggcggcgcga ggccaggcac cagagcaggc
     4381 cggccagctt gagactaccc ccgtccgatt ctcggtggcc gcgctcgcag gccccgcctc
     4441 gccgaacatg tgcgctggga cgcacgggcc ccgtcgccgc ccgcggcccc aaaaaccgaa
     4501 ataccagtgt gcagatcttg gcccgcattt acaagactat cttgccagaa aaaaagcgtc
     4561 gcagcaggtc atcaaaaatt ttaaatggct agagacttat cgaaagcagc gagacaggcg
     4621 cgaaggtgcc accagattcg cacgcggcgg ccccagcgcc caggccaggc ctcaactcaa
     4681 gcacgaggcg aaggggctcc ttaagcgcaa ggcctcgaac tctcccaccc acttccaacc
     4741 cgaagctcgg gatcaagaat cacgtactgc agccaggtgg aagtaattca aggcacgcaa
     4801 gggccataac ccgtaaagag gccaggcccg cgggaaccac acacggcact tacctgtgtt
     4861 ctggcggcaa acccgttgcg aaaaagaacg ttcacggcga ctactgcact tatatacggt
     4921 tctcccccac cctcgggaaa aaggcggagc cagtacacga catcactttc ccagtttacc
     4981 ccgcgccacc ttctctaggc accggttcaa ttgccgaccc ctccccccaa cttctcgggg
     5041 actgtgggcg atgtgcgctc tgcccactga cgggcaccgg agccaattcc cactcctttc
     5101 aagacctaga aggtccatta gctgcaaaga ttcctctctg tttaaaactt tatccatctt
     5161 tgcaaagctt acgcgtgcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc
     5221 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggcggcc
     5281 tcagtgagcg agcgagcgcg cagctgcctg caggacatgt gagcaaaagg ccagcaaaag
     5341 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5401 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5461 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5521 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5581 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5641 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     5701 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5761 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     5821 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5881 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5941 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6001 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6061 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6121 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6181 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6241 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     6301 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     6361 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6421 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     6481 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6541 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     6601 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     6661 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     6721 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     6781 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     6841 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6901 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6961 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     7021 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     7081 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     7141 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     7201 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     7261 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     7321 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     7381 aaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     7441 ttttttaacc aatagaccga aatcggcaaa atccctta
//
