LOCUS       sequence-463012-        7341 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral expression of mLhx3 with mRuby2 fluorescent reporter with
            upstream open reading frame.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7341)
  AUTHORS   Wang NB, Lende-Dorn BA, Beitz AM, Han P, Adewumi HO, O'Shea TM, 
            Galloway KE
  TITLE     Proliferation history and transcription factor levels drive direct 
            conversion to motor neurons.
  JOURNAL   Cell Syst. 2025 Apr 16;16(4):101205. doi: 
            10.1016/j.cels.2025.101205. Epub 2025 Mar 13.
  PUBMED    40086434
REFERENCE   2  (bases 1 to 7341)
  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."; date: "2025-04-16"; volume: "16(4):101205. doi"; pages: " 
            10.1016/j.cels.2025.101205. Epub 2025 Mar 13"
COMMENT     Sequence Label: pMXs-uORFx2-mLhx3-P2A-mRuby2
FEATURES             Location/Qualifiers
     source          1..7341
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     117..133
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     protein_bind    222..246
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1538..1594
                     /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             1595..2305
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     protein_bind    complement(2322..2346)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2363..2951
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2416..2436)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2834..2845)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             3131..3600
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(3730..3747)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3901..4489)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3981..4000)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4653..5444,5445..5513))
                     /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     5276..5295
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5514..5618)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             5648..6241
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    6304..6661
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             6720..7136
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     7066..7088
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     7104..7120
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    join(7146..7341,1..179)
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
ORIGIN
        1 tcggcgacac agtgtgggtc cgccgacacc agactaagaa cctagaacct cgctggaaag
       61 gaccttacac agtcctgctg accaccccca ccgccctcaa agtagacggc atcgcagctt
      121 ggatacacgc cgcccacgtg aaggctgccg accccggggg tggaccatcc tctagactgc
      181 cggatctagc tagttaatcg acggtatcga taagctatca aacaagtttg tacaaaaaag
      241 caggctccac catgggaacc aattcagtcg agtcgactgg atccaccatg ggttgaacca
      301 tgggttgaga tccggtacta ctgccaccat gctgctagaa gcagaactcg attgccaccg
      361 agagaggccc ggtgcccctg gagcttctgc cctctgtacc ttcagcagga ctccagagat
      421 cccgatgtgt gcaggctgtg accagcacat cttggaccgt ttcatcctta aggctctgga
      481 ccgacattgg cacagcaagt gtctcaagtg cagtgactgc cacgtccctc tggctgagcg
      541 ctgcttcagc cgcggggaga gcgtctactg caaagacgac ttctttaagc gcttcgggac
      601 caagtgcgcc gcatgccagc tgggcatccc gcccacgcag gtggtgcgcc gcgcccagga
      661 cttcgtgtac cacctgcatt gcttcgcctg cgtggtctgc aagcggcagc tggccacggg
      721 cgacgagttc tacctcatgg aagacagccg gctggtgtgc aaggcggact acgaaacagc
      781 caagcagcga gaagccgagg ccacagccaa gcggccgcgc accaccatca ccgccaagca
      841 gctggagacg ctgaagagcg cctacaacac ttcgcccaag ccggcgcgcc acgtgcgcga
      901 gcagctctcc tccgagactg gcctggacat gcgagtggtg caggtgtggt tccagaatcg
      961 ccgggctaag gaaaagagac tgaagaaaga cgctggccgg cagcgctggg gacagtattt
     1021 ccgcaatatg aagcgctccc gcggcagttc caagtccgac aaggacagca tccaggaggg
     1081 acaagacagc gacgccgaag tctccttcac tgatgagccg tccatggctg acatggggcc
     1141 tgctaatggc ctgtacagca gcctgggaga gcctgcccct gcgttgggcc ggcccgtagg
     1201 aggcctgggc agctttaccc tggatcacgg aggcttgacg ggtccagagc agtaccgaga
     1261 gctacgccca ggcagcccct atggcatccc cccatctcct gcagcccccc agagccttcc
     1321 tggcccccag cctctcctct ccagcctggt atacccagac accaacttga gccttgtccc
     1381 ttcagggccc ccaggtggac ccccacccat gagggtgctg gctggaaatg ggcccagctc
     1441 cgacctgtcc acagagagca gttctggcta cccagacttt cctgctagcc ctgcttcctg
     1501 gctggatgaa gtagaccatg ctcagttcgg aagcggagct actaacttca gcctgctgaa
     1561 gcaggctgga gacgtggagg agaaccctgg acctatggtg tctaagggcg aagagctgat
     1621 caaggaaaat atgcgtatga aggtggtcat ggaaggttcg gtcaacggcc accaattcaa
     1681 atgcacaggt gaaggagaag gcaatccgta catgggaact caaaccatga ggatcaaagt
     1741 catcgaggga ggacccctgc catttgcctt tgacattctt gccacgtcgt tcatgtatgg
     1801 cagccgtact tttatcaagt acccgaaagg cattcctgat ttctttaaac agtcctttcc
     1861 tgagggtttt acttgggaaa gagttacgag atacgaagat ggtggagtcg tcaccgtcat
     1921 gcaggacacc agccttgagg atggctgtct cgtttaccac gtccaagtca gaggggtaaa
     1981 ctttccctcc aatggtcccg tgatgcagaa gaagaccaag ggttgggagc ctaatacaga
     2041 gatgatgtat ccagcagatg gtggtctgag gggatacact catatggcac tgaaagttga
     2101 tggtggtggc catctgtctt gctctttcgt aacaacttac aggtcaaaaa agaccgtcgg
     2161 gaacatcaag atgcccggta tccatgccgt tgatcaccgc ctggaaaggt tagaggaaag
     2221 tgacaatgaa atgttcgtag tacaacgcga acacgcagtt gccaagttcg ccgggcttgg
     2281 tggtgggatg gacgagctgt acaagtaacg ctgatatcta gacccagctt tcttgtacaa
     2341 agtggttcga tagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac
     2401 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     2461 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     2521 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     2581 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     2641 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     2701 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc
     2761 atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     2821 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     2881 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     2941 cgcctccccg catcgatacc gtcgacgata aaataaaaga ttttatttag tctccagaaa
     3001 aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt
     3061 tgcaaggcat ggaaaaatac ataactgaga atagagaagt tcagatcaag gtcaggaaca
     3121 gatggaacag ctgaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc
     3181 tcagggccaa gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc
     3241 agttcctgcc ccggctcagg gccaagaaca gatggtcccc agatgcggtc cagccctcag
     3301 cagtttctag agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt
     3361 gccttatttg aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc
     3421 cgagctcaat aaaagagccc acaacccctc actcggggcg ccagtcctcc gattgactga
     3481 gtcgcccggg tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct
     3541 cgctgttcct tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca
     3601 catgcagcat gtatcaaaat taatttggtt ttttttctta agtatttaca ttaaatggcc
     3661 atagttgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     3721 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     3781 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     3841 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     3901 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     3961 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4021 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4081 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4141 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4201 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4261 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4321 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     4381 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4441 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     4501 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4561 catgagatta tcaaaaagga tcttcaccta gatccttttg cggccggccg caaatcaatc
     4621 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4681 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4741 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4801 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4861 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4921 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4981 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5041 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5101 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5161 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5221 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5281 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5341 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5401 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5461 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5521 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5581 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5641 cctgcataat gaaagacccc acctgtaggt ttggcaagct agcttaagta acgccatttt
     5701 gcaaggcatg gaaaaataca taactgagaa tagaaaagtt cagatcaagg tcaggaacag
     5761 atggaacagc tgaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct
     5821 cagggccaag aacagatgga acagctgaat atgggccaaa caggatatct gtggtaagca
     5881 gttcctgccc cggctcaggg ccaagaacag atggtcccca gatgcggtcc agccctcagc
     5941 agtttctaga gaaccatcag atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg
     6001 ccttatttga actaaccaat cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc
     6061 gagctcaata aaagagccca caacccctca ctcggcgcgc cagtcctccg attgactgag
     6121 tcgcccgggt acccgtgtat ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc
     6181 gctgttcctt gggagggtct cctctgagtg attgactacc cgtcagcggg ggtctttcat
     6241 ttgggggctc gtccgggatc gggagacccc tgcccaggga ccaccgaccc accaccggga
     6301 ggtaagctgg ccagcaactt atctgtgtct gtccgattgt ctagtgtcta tgactgattt
     6361 tatgcgcctg cgtcggtact agttagctaa ctagctctgt atctggcgga cccgtggtgg
     6421 aactgacgag ttcggaacac ccggccgcaa ccctgggaga cgtcccaggg acttcggggg
     6481 ccgtttttgt ggcccgacct gagtccaaaa atcccgatcg ttttggactc tttggtgcac
     6541 ccccctaata ggagggatat gtggttctgg taggagacga gaacctaaaa cagttcccgc
     6601 ctccgtctga atttttgctt tcggtttggg accgaagccg cgccgcgcgt cttgtctgct
     6661 gcagcatcgt tctgtgttgt ctctgtctga ctgtgtttct gtatttgtct gaaaattagg
     6721 gccagactgt taccactccc ttaagtttga ccttaggtca ctggaaagat gtcgagcgga
     6781 tcgctcacaa ccagtcggta gatgtcaaga agagacgttg ggttaccttc tgctctgcag
     6841 aatggccaac ctttaacgtc ggatggccgc gagacggcac ctttaaccga gacctcatca
     6901 cccaggttaa gatcaaggtc ttttcacctg gcccgcatgg acacccagac caggtcccct
     6961 acatcgtgac ctgggaagcc ttggcttttg acccccctcc ctgggtcaag ccctttgtac
     7021 accctaagcc tccgcctcct cttcctccat ccgccccgtc tctccccctt gaacctcctc
     7081 gttcgacccc gcctcgatcc tccctttatc cagccctcac tccttctcta ggcgccccca
     7141 tatggccata tgagatctta tatggggcac ccccgcccct tgtaaacttc cctgaccctg
     7201 acatgacaag agttactaac agcccctctc tccaagctca cttacaggct ctctacttag
     7261 tccagcacga agtctggaga cctctggcgg cagcctacca agaacaactg gaccgaccgg
     7321 tggtacctca cccttaccga g
//
