LOCUS       sequence-475634-        7654 bp    DNA     circular SYN 11-JUN-2025
DEFINITION  L3 pyramidal neuron Enhancer to drive transgene expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7654)
  AUTHORS   He J, Phan BN, Kerkhoff WG, Alikaya A, Hong T, Brull OR, Fredericks 
            JM, Sedorovitz M, Srinivasan C, Leone MJ, Wirfel OM, Brown A, Dauby 
            S, Tittle RK, Lin MK, Hooks BM, Bostan AC, Gharbawie OA, Byrne LC, 
            Pfenning AR, Stauffer WR
  TITLE     Machine learning identification of enhancers in the rhesus macaque 
            genome.
  JOURNAL   Neuron. 2025 May 21;113(10):1548-1561.e8. doi: 
            10.1016/j.neuron.2025.04.030.
  PUBMED    40403706
REFERENCE   2  (bases 1 to 7654)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 11, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.neuron.2025.04"; journalName: "Neuron"; date: 
            "2025-05-21- 21"; volume: "113"; issue: "10"; pages: "1548-1561.e8"
COMMENT     Sequence Label: pAAV.RMacL3-01.ChR2(H134R)-GFP.BCC10
FEATURES             Location/Qualifiers
     source          1..7654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     regulatory      1593..1602
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1599..2525
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      2532..2541
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2538..2540,2541..2543,2544..3257)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2538..2540,2541..2543,2544..3254)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2583..2604)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2844..2863)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3191..3212
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3864..3884)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4282..4293)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4414..4535
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4425..4444)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4479..4498
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4704..4833
                     /label=AAV2 ITR
     primer_bind     complement(4852..4869)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4852..4868)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4861..4883)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5010..5465
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5097..5116)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5307..5328
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5491..5595
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5596..5664,5665..6456)
                     /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(5814..5833)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6627..7215
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7116..7135
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    7503..7524
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7539..7544,7545..7562,7563..7569)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7577..7593
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     7582..7604
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7601..7617
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7601..7617
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcg gaattcgtca ggaaactcgg gctgaaatcc cactactggg ggaaatctgc
      241 taacccctct gaacctcatt tttcacttgc atcaaagggg ataaagtgct tcttgcctct
      301 gggccgtggg catctgtggg attggactgg gtcatgggga gccgtggggt ggcactgggg
      361 agctgcatca gccgccagca tctctgaggc ctttgctttc tctgagggaa ggcccagttt
      421 gatttaggtc ctgagaatcc cagaatggcc atttctggaa aaaataaccc aaagcccaga
      481 tttccctttt agttttatcc cacacgggcc agcaacatgc ataacaaaat attccatttc
      541 tcaaaggaag tcttagaata ttggatgaat gcactgtgtg ccttgacttc attcgcaaag
      601 cacccatgtc ccttggatcc cttccgctct ccctcccgcc cccacgctgc ccttgttctg
      661 ggaaatccag ctctccagga gcaatcagca cagaggccag gaacatccaa actgagcagc
      721 cggggtcccc cccacccccc accccgcccc acgcggcaac tttgagcctg tgctgggaca
      781 gagcctctag ttcctaaatt agtccatgag gtcagaggca gcactgccat tgtaacgcga
      841 ttggagagga tcacgtcacc ggacacgccc ccaggcatct ccctgggtct cctaaacttg
      901 gcggggagaa gttttagccc ttaagtttta gcctttaacc cccatattca gaactgtgcg
      961 agttggcgaa accccacaaa tcacaacaaa ctgtacacaa caccgagcta gaggtgatct
     1021 ttcttgtcca ttccacacag gccttagtaa tgcgtcgcca tagcaacagt gtcactagta
     1081 gcaccagcac ttccccacac cctccccctc aggaatccgt actctccagt gaaccccaga
     1141 aacctctgga gagttctgga caagggcgga acccacaact ccgattactc aagggaggcg
     1201 gggaagctcc accagacgcg aaactgctgg aagattcctg gccccaaggc ctcctccggc
     1261 tcgctgattg gcccagcgga gagtgggcgg ggccggtgaa gactccttaa aggcgcaggg
     1321 cggcgagcag gtcaccagac gctgacagct actcagaacc aaatctggtt ccatccagag
     1381 acaagcgaag acaagagaag cagagcgagc ggcgcgttcc cgatcctcgg ccaggaccag
     1441 ccttccccag agcatccctg ccgcggagcg caaccttccc aggagcatcc ctgccgcgga
     1501 gcgcaacttt ccccggagca tccacgccgc ggagcgcagc cttccagaag cagagcgcgg
     1561 cgccggagcg ggatcagcca ccgcgggcta gcgccaccat ggactatggc ggcgctttgt
     1621 ctgccgtcgg acgcgaactt ttgttcgtta ctaatcctgt ggtggtgaac gggtccgtcc
     1681 tggtccctga ggatcaatgt tactgtgccg gatggattga atctcgcggc acgaacggcg
     1741 ctcagaccgc gtcaaatgtc ctgcagtggc ttgcagcagg attcagcatt ttgctgctga
     1801 tgttctatgc ctaccaaacc tggaaatcta catgcggctg ggaggagatc tatgtgtgcg
     1861 ccattgaaat ggttaaggtg attctcgagt tcttttttga gtttaagaat ccctctatgc
     1921 tctaccttgc cacaggacac cgggtgcagt ggctgcgcta tgcagagtgg ctgctcactt
     1981 gtcctgtcat ccttatccgc ctgagcaacc tcaccggcct gagcaacgac tacagcagga
     2041 gaaccatggg actccttgtc tcagacatcg ggactatcgt gtggggggct accagcgcca
     2101 tggcaaccgg ctatgttaaa gtcatcttct tttgtcttgg attgtgctat ggcgcgaaca
     2161 cattttttca cgccgccaaa gcatatatcg agggttatca tactgtgcca aagggtcggt
     2221 gccgccaggt cgtgaccggc atggcatggc tgtttttcgt gagctggggt atgttcccaa
     2281 ttctcttcat tttggggccc gaaggttttg gcgtcctgag cgtctatggc tccaccgtag
     2341 gtcacacgat tattgatctg atgagtaaaa attgttgggg gttgttggga cactacctgc
     2401 gcgtcctgat ccacgagcac atattgattc acggagatat ccgcaaaacc accaaactga
     2461 acatcggcgg aacggagatc gaggtcgaga ctctcgtcga agacgaagcc gaggccggag
     2521 ccgtgccagc ggccgccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     2581 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     2641 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     2701 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     2761 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     2821 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     2881 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     2941 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     3001 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     3061 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     3121 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     3181 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     3241 acgagctgta caagtaagtt taaacacagg gttaatcatt aactacaggt gtggagacgt
     3301 ggagaacctc aatactctct cattgtagtt ggagcggaac tggatattga gagatgtggt
     3361 gccaacgtat gatggtctct cttgcacacc ggaatagtga cgacattgtc gtagacctga
     3421 catacagcta catggtggtg taaccactcg tatcttgata ggataacgcg ctcgccatca
     3481 atacaagtct gcacatggtc agactaagag gataggttag aatccgtgtg ttatcacgct
     3541 cttgacctgt acttccaaag accgcttgtt cgaaggagga tcaggaccgc agcctattgg
     3601 acacacaacg tacacatagg agatcagagg acccagatga taccaaccta cgtgagtcct
     3661 gcactacaag ttcgtccgca ctatcctggt ctcgccgata ggttctgacc tccggtacta
     3721 gctatcgagg tgttgtccaa cagtgctatg aacgagcatc ttaggacctg cctcagacta
     3781 ctccgtggaa taccgcatgc ggccgcgatc gctcaacctc tggattacaa aatttgtgaa
     3841 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     3901 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     3961 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     4021 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     4081 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     4141 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     4201 gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg
     4261 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     4321 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     4381 ctttgggccg cctccccgcg atcgcagaca tgataagata cattgatgag tttggacaaa
     4441 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     4501 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     4561 tgtttcaggt tcagggggag atgtgggagg ttttttaaag caagtaaaac ctctacaaat
     4621 gtggtagtcg aaattcccga taaggatctt cctagagcat ggctacgtag ataagtagca
     4681 tggcgggtta atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct
     4741 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     4801 ccgggcggcc tcagtgagcg agcgagcgcg cagccttaat taacctaatt cactggccgt
     4861 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     4921 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     4981 acagttgcgc agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc
     5041 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     5101 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     5161 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     5221 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     5281 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     5341 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     5401 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac
     5461 aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     5521 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     5581 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     5641 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5701 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5761 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5821 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     5881 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5941 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     6001 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     6061 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     6121 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     6181 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     6241 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     6301 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     6361 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     6421 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     6481 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     6541 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     6601 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     6661 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     6721 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     6781 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     6841 ctgctaatcc tgttaccagt gcctgctgcc agtggcgata agtcgtgtct taccgggttg
     6901 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     6961 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     7021 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     7081 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     7141 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     7201 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     7261 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     7321 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     7381 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     7441 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     7501 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     7561 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
     7621 gattacgcca gatttaatta aggccttaat tagg
//
