LOCUS       sequence-465054-        7026 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV expression of Lhx3, Ngn2x3HA, and Isl1 for motor neuron 
            reprogramming.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7026)
  AUTHORS   Wang NB, Adewumi HO, Lende-Dorn BA, Beitz AM, O'Shea TM, Galloway KE
  TITLE     Compact transcription factor cassettes generate functional, 
            engraftable motor neurons by direct conversion.
  JOURNAL   Cell Syst. 2025 Mar 10:101206. doi: 10.1016/j.cels.2025.101206.
  PUBMED    40086435
REFERENCE   2  (bases 1 to 7026)
  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: "Cell Syst. 
            2025 Mar 10:101206. doi: 10.1016/j.cels.2025.101206."
COMMENT     Sequence Label: pAAV-CMV-LNI-bGH
FEATURES             Location/Qualifiers
     source          1..7026
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   64..193
                     /label=AAV2 ITR
     protein_bind    206..230
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        273..576
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        577..780
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     731..751
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     782..806
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2050..2106
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2935..2961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2971..2997
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3007..3033
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3043..3096
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      3887..3896
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4158..4382
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(4396..4420)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     repeat_region   4447..4576
                     /label=AAV2 ITR
     rep_origin      complement(4813..5401)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4893..4912)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5572..6363,6364..6432))
                     /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     6195..6214
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6433..6537)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6563..7018)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6700..6721)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6912..6931
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
       61 cagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc
      121 tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc
      181 actaggggtt ccttgtagtt aatgaacaag tttgtacaaa aaagcaggct ccaccatggg
      241 aaccaattca gtcgactgga tccggtacta ctcgttacat aacttacggt aaatggcccg
      301 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      361 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      421 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      481 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      541 cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc
      601 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
      661 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
      721 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct
      781 gagctcgttt agtgaaccgt cagatcgcct ggagacgcca tccacgctgt aatggccacc
      841 atgctgctag aagcagaact cgattgccac cgagagaggc ccggtgcccc tggagcttct
      901 gccctctgta ccttcagcag gactccagag atcccgatgt gtgcaggctg tgaccagcac
      961 atcttggacc gtttcatcct taaggctctg gaccgacatt ggcacagcaa gtgtctcaag
     1021 tgcagtgact gccacgtccc tctggctgag cgctgcttca gccgcgggga gagcgtctac
     1081 tgcaaagacg acttctttaa gcgcttcggg accaagtgcg ccgcatgcca gctgggcatc
     1141 ccgcccacgc aggtggtgcg ccgcgcccag gacttcgtgt accacctgca ttgcttcgcc
     1201 tgcgtggtct gcaagcggca gctggccacg ggcgacgagt tctacctcat ggaagacagc
     1261 cggctggtgt gcaaggcgga ctacgaaaca gccaagcagc gagaagccga ggccacagcc
     1321 aagcggccgc gcaccaccat caccgccaag cagctggaga cgctgaagag cgcctacaac
     1381 acttcgccca agccggcgcg ccacgtgcgc gagcagctct cctccgagac tggcctggac
     1441 atgcgagtgg tgcaggtgtg gttccagaat cgccgggcta aggaaaagag actgaagaaa
     1501 gacgctggcc ggcagcgctg gggacagtat ttccgcaata tgaagcgctc ccgcggcagt
     1561 tccaagtccg acaaggacag catccaggag ggacaagaca gcgacgccga agtctccttc
     1621 actgatgagc cgtccatggc tgacatgggg cctgctaatg gcctgtacag cagcctggga
     1681 gagcctgccc ctgcgttggg ccggcccgta ggaggcctgg gcagctttac cctggatcac
     1741 ggaggcttga cgggtccaga gcagtaccga gagctacgcc caggcagccc ctatggcatc
     1801 cccccatctc ctgcagcccc ccagagcctt cctggccccc agcctctcct ctccagcctg
     1861 gtatacccag acaccaactt gagccttgtc ccttcagggc ccccaggtgg acccccaccc
     1921 atgagggtgc tggctggaaa tgggcccagc tccgacctgt ccacagagag cagttctggc
     1981 tacccagact ttcctgctag ccctgcttcc tggctggatg aagtagacca tgctcagttc
     2041 ggaagcggag ctactaactt cagcctgctg aagcaggctg gagacgtgga ggagaaccct
     2101 ggacctatgt tcgtcaaatc tgagactctg gagttgaagg aggaagagga ggtactgatg
     2161 ctgctgggct cggcttcccc ggcctcggcg accctgaccc cgatgtcctc cagcgcggac
     2221 gaggaggagg acgaggagct gcgccggccg ggctccgcgc gtgggcagcg tggagcggaa
     2281 gccgggcagg gggtgcaggg cagtccggcg tcgggtgccg ggggttgccg gccagggcgg
     2341 ctgctgggcc tgatgcacga gtgcaagcgt cgcccgtcgc gctcacgggc cgtctcccga
     2401 ggtgccaaga cggcggagac ggtgcagcgc atcaagaaga cccgcaggct caaggccaac
     2461 aaccgcgagc gcaaccgcat gcacaaccta aacgccgcgc tggacgcgct gcgcgaggtg
     2521 ctgcccacct tccccgagga tgccaagctc acgaagatcg agacgctgcg cttcgcccac
     2581 aattacatct gggcgctcac cgagactctg cgcctggcgg accactgcgc cggcgccggt
     2641 ggcctccagg gggcgctctt cacggaggcg gtgctcctga gcccgggagc tgcgctcggc
     2701 gccagcgggg acagcccttc tccaccttcc tcctggagct gcaccaacag cccggcgtca
     2761 tcctccaact ccacgtcccc atacagctgc actttatcgc ccgctagccc cgggtcagac
     2821 gtggactact ggcagccccc acctccggag aagcatcgtt atgcgcctca cctgcccctc
     2881 gccagggact gtatcgggtc tggctccgga tcaagtggtg gcagcggatc aggatatcca
     2941 tacgatgtac cagactatgc gggatcgggt tacccctacg acgtccccga ctacgccggc
     3001 tccggctacc cctacgacgt ccctgattat gctggcagtg gtgagggcag aggaagtctg
     3061 ctaacatgcg gtgacgtcga ggagaatcct ggcccaatgg gagacatggg cgatccacca
     3121 aaaaaaaaac gtctgatttc cctgtgtgtt ggttgcggca atcaaattca cgaccagtat
     3181 attctgaggg tttctccgga tttggagtgg catgcagcat gtttgaaatg tgcggagtgt
     3241 aatcagtatt tggacgaaag ctgtacgtgc tttgttaggg atgggaaaac ctactgtaaa
     3301 agagattata tcaggttgta cgggatcaaa tgcgccaagt gcagcatagg cttcagcaag
     3361 aacgacttcg tgatgcgcgc ccgctctaag gtgtaccaca tcgagtgttt ccgctgtgta
     3421 gcctgcagcc gacagctcat cccgggagac gaattcgccc tgcgggagga tgggcttttc
     3481 tgccgtgcag accacgatgt ggtggagaga gccagcctgg gagctggaga tcctctcagt
     3541 cccttgcatc cagcgcggcc tctgcaaatg gcagccgaac ccatctcggc taggcagcca
     3601 gctctgcggc cgcacgtcca caagcagccg gagaagacca cccgagtgcg gactgtgctc
     3661 aacgagaagc agctgcacac cttgcggacc tgctatgccg ccaaccctcg gccagatgcg
     3721 ctcatgaagg agcaactagt ggagatgacg ggcctcagtc ccagagtcat ccgagtgtgg
     3781 tttcaaaaca agcggtgcaa ggacaagaaa cgcagcatca tgatgaagca gctccagcag
     3841 cagcaaccca acgacaaaac taatatccag gggatgacag gaactcccat ggtggctgct
     3901 agtccggaga gacatgatgg tggtttacag gctaacccag tagaggtgca aagttaccag
     3961 ccgccctgga aagtactgag tgacttcgcc ttgcaaagcg acatagatca gcctgctttt
     4021 cagcaactgg tcaatttttc agaaggagga ccaggctcta attctactgg cagtgaagta
     4081 gcatcgatgt cctcgcagct cccagataca cccaacagca tggtagccag tcctattgag
     4141 gcatgacaac gttgcacctg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     4201 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     4261 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     4321 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     4381 ggcgctgata tctagaccca gctttcttgt acaaagtggt gcatggcggg ttaatcatta
     4441 actacaagga acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca
     4501 ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga
     4561 gcgagcgagc gcgcagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4621 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4681 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4741 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4801 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4861 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4921 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4981 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5041 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5101 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5161 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5221 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5281 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5341 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5401 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5461 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5521 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5581 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5641 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5701 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5761 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     5821 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     5881 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     5941 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6001 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6061 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6121 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6181 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6241 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6301 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6361 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6421 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6481 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6541 atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta
     6601 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     6661 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     6721 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     6781 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     6841 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     6901 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     6961 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc
     7021 cattcg
//
