LOCUS       sequence-461588-        6721 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  L5 extratelencephalic projection neuron Enhancer to drive transgene 
            expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6721)
  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 6721)
  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"; 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.RMacL5ET-04.EGFP.BCB2.WPRE
FEATURES             Location/Qualifiers
     source          1..6721
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     regulatory      1599..1608
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1605..1607,1608..1610,1611..2324)
                     /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(1605..1607,1608..1610,1611..2321)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1650..1671)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1911..1930)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2258..2279
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2931..2951)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3349..3360)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3481..3602
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3492..3511)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3546..3565
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3771..3900
                     /label=AAV2 ITR
     primer_bind     complement(3919..3936)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3919..3935)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3928..3950)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4077..4532
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4164..4183)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4374..4395
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4558..4662
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4663..4731,4732..5523)
                     /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(4881..4900)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5694..6282
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6183..6202
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6570..6591
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6606..6611,6612..6629,6630..6636)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6644..6660
                     /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     6649..6671
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6668..6684
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6668..6684
                     /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 gaattcactg ttgtctgccg tacgcaactg aacagactcg cccatatcct
      241 caagaagctc gtggtccact gtggggacag gcaggtaacg tgcaatgcgt gcaggaagga
      301 ggaagcagag gttgctgggg agtgtggtag aggctctgcc tgtgatagct gcggggcctt
      361 gggcagggct gcgggccgcc ctgcatctcc acatcacctg ttgcctgcaa gtttcagaac
      421 ccaaaacggg ccgatcaaac ccccgtcata ctggttctgg ccatcagagg gcgcgcgaag
      481 cagcatttag cgggacggag cagcaccccc aactctgcag acccctgggc tgtccgggag
      541 ctgggtactt atgccgctgg gccaccaaca gagagaggac ataggcgcga tgtccacaaa
      601 actctggctt taccccaaac cgttcccggc tggaggaatc ttccaacgac atggcacata
      661 gtctccttgt cttccacgct gttgggtctg tggggactcc tagtttccat gctctagcct
      721 gcaggcagga acatccaaac tgagcagccg gggtcccccc caccccccac cccgccccac
      781 gcggcaactt tgagcctgtg ctgggacaga gcctctagtt cctaaattag tccatgaggt
      841 cagaggcagc actgccattg taacgcgatt ggagaggatc acgtcaccgg acacgccccc
      901 aggcatctcc ctgggtctcc taaacttggc ggggagaagt tttagccctt aagttttagc
      961 ctttaacccc catattcaga actgtgcgag ttggcgaaac cccacaaatc acaacaaact
     1021 gtacacaaca ccgagctaga ggtgatcttt cttgtccatt ccacacaggc cttagtaatg
     1081 cgtcgccata gcaacagtgt cactagtagc accagcactt ccccacaccc tccccctcag
     1141 gaatccgtac tctccagtga accccagaaa cctctggaga gttctggaca agggcggaac
     1201 ccacaactcc gattactcaa gggaggcggg gaagctccac cagacgcgaa actgctggaa
     1261 gattcctggc cccaaggcct cctccggctc gctgattggc ccagcggaga gtgggcgggg
     1321 ccggtgaaga ctccttaaag gcgcagggcg gcgagcaggt caccagacgc tgacagctac
     1381 tcagaaccaa atctggttcc atccagagac aagcgaagac aagagaagca gagcgagcgg
     1441 cgcgttcccg atcctcggcc aggaccagcc ttccccagag catccctgcc gcggagcgca
     1501 accttcccag gagcatccct gccgcggagc gcaactttcc ccggagcatc cacgccgcgg
     1561 agcgcagcct tccagaagca gagcgcggcg ccgctagcgc caccatggtg agcaagggcg
     1621 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     1681 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     1741 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
     1801 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
     1861 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1921 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     1981 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     2041 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     2101 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     2161 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     2221 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     2281 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaagtttaa acacagggtt
     2341 aatcattaac tacaggtcgc tctcatcatc ggatcttacc gtgcctgaag atagctgcaa
     2401 cgaccaagta ggaactcgtg ttgaagctcc ctccacctta attgtcatgt acgcgcacca
     2461 gtaccggacc acagacgtac aacctcacct ctcgcctacc accatgaacg accgatgatg
     2521 ccgtagactg gtgttcacaa cctgacctgc atggtatgtc actctcacgt ctggaggtaa
     2581 tgatgaactg gtcggaggtc cgaagaagct aggacgtgtg ttcgatctaa tacgaccgat
     2641 atagtcgtcg gtcatcctcc gcgtaatggt aagtgtagaa ccagattgac tgtgaggatc
     2701 tctggactat gttggagagt aacagagcgg gattcgcagt gttaccctta acgaacatgg
     2761 tcgaccatat tgtagcagga ctaggattgg agtctggaat gagtatcagg tatcaatggt
     2821 tcgagcgtta atcttacctt atacgtgaac atcatactgc ttcgagcggc cgcgatcgct
     2881 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     2941 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     3001 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     3061 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     3121 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     3181 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     3241 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     3301 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     3361 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     3421 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcgatc gcagacatga
     3481 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     3541 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     3601 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt
     3661 tttaaagcaa gtaaaacctc tacaaatgtg gtagtcgaaa ttcccgataa ggatcttcct
     3721 agagcatggc tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct
     3781 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     3841 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     3901 ccttaattaa cctaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     3961 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     4021 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     4081 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     4141 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     4201 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     4261 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     4321 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     4381 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     4441 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     4501 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     4561 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4621 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     4681 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4741 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4801 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4861 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4921 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     4981 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     5041 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     5101 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     5161 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     5221 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     5281 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     5341 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     5401 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     5461 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     5521 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     5581 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5641 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5701 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5761 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5821 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5881 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5941 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     6001 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     6061 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     6121 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     6181 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     6241 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6301 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     6361 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     6421 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     6481 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     6541 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     6601 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     6661 tttcacacag gaaacagcta tgaccatgat tacgccagat ttaattaagg ccttaattag
     6721 g
//
