LOCUS       sequence-354020-        6894 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent expression of NEUROD1 and mCherry under the 
            constitutive promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6894)
  AUTHORS   Wang LL, Serrano C, Zhong X, Ma S, Zou Y, Zhang CL
  TITLE     Revisiting astrocyte to neuron conversion with lineage tracing in 
            vivo.
  JOURNAL   Cell. 2021 Oct 14;184(21):5465-5481.e16. doi: 
            10.1016/j.cell.2021.09.005. Epub 2021 Sep 27.
  PUBMED    34582787
REFERENCE   2  (bases 1 to 6894)
  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.cell.2021.09.005"; journalName: "Cell"; date: 
            "2021-10-14- 14"; volume: "184"; issue: "21"; pages: "5465-5481.e16"
COMMENT     Sequence Label: AAV-CAG-FLEX-NEUROD1-T2A-mCherry
FEATURES             Location/Qualifiers
     source          1..6894
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        441..719
                     /label=chicken beta-actin promoter
     primer_bind     998..1020
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1082..1101
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1136..1169
                     /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)."
     protein_bind    1232..1265
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1286..1996)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1564..1583)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1833..1851
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      1993..2002
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(2009..2062)
                     /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"
     protein_bind    complement(3159..3192)
                     /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)."
     protein_bind    complement(3263..3296)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3338..3926
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3391..3411)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3809..3820)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3969..4090
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3980..3999)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4034..4053
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4252..4381
                     /label=AAV2 ITR
     rep_origin      complement(4618..5206)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4698..4717)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5377..6168,6169..6237))
                     /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     6000..6019
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6238..6342)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6368..6823)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6505..6526)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6717..6736
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagagta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      241 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      301 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      361 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      421 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc
      481 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat
      541 gggggcgggg gggggggggg gggcgcgcgc caggcggggc ggggcggggc gaggggcggg
      601 gcggggcgag gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc
      661 cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg
      721 gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc
      781 cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc
      841 cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa
      901 gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg ctgtccgcgg
      961 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1021 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1081 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattgga tccccataac
     1141 ttcgtataaa gtatcctata cgaagttata tcaaaatagg aagaccaatg cttcaccatc
     1201 gacccgaatt gccaagcatc accatcgacc cataacttcg tataatgtat gctatacgaa
     1261 gttatactag ggatcccccc tcgagctact tgtacagctc gtccatgccg ccggtggagt
     1321 ggcggccctc ggcgcgttcg tactgttcca cgatggtgta gtcctcgttg tgggaggtga
     1381 tgtccaactt gatgttgacg ttgtaggcgc cgggcagctg cacgggcttc ttggccttgt
     1441 aggtggtctt gacctcagcg tcgtagtggc cgccgtcctt cagcttcagc ctctgcttga
     1501 tctcgccctt cagggcgccg tcctcggggt acatccgctc ggaggaggcc tcccagccca
     1561 tggtcttctt ctgcattacg gggccgtcgg aggggaagtt ggtgccgcgc agcttcacct
     1621 tgtagatgaa ctcgccgtcc tgcagggagg agtcctgggt cacggtcacc acgccgccgt
     1681 cctcgaagtt catcacgcgc tcccacttga agccctcggg gaaggacagc ttcaagtagt
     1741 cggggatgtc ggcggggtgc ttcacgtagg ccttggagcc gtacatgaac tgaggggaca
     1801 ggatgtccca ggcgaagggc agggggccac ccttggtcac cttcagcttg gcggtctggg
     1861 tgccctcgta ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcacgg
     1921 agccctccat gtgcaccttg aagcgcatga actccttgat gatggccatg ttatcctcct
     1981 cgcccttgct caccatggtg gcaccggtgg gtccaggatt ctcttcgacg tctccgcagg
     2041 tcagaagact tcctctcccc tcgctagcat catgaaatat ggcattgagc tgggcactca
     2101 tgactcgctc atgatgtgaa tggctatcga aggacataat attgtctatg gggatctcgc
     2161 agcgaggggc agcggtgcct gagaagattg atccgtggct ttgggcccct gccagtgtcg
     2221 ctgcaggata gtgcatggta aaggcataat ttttctcaaa ctcggcggac ggttcgtgtt
     2281 tgaaagagaa gttgccattg atgctgagcg gcgggctgag gggtccatca aaggaagggc
     2341 tggtgcaatc agtcagaggg ctttcaaaga agggctccag cgctgcgctg taggcgtgcg
     2401 gcggaggctt aacgtggaag acatgggagc tgtccatggt accgtaaggc ggactgggca
     2461 gcccaggcga ctggtaggag taggggtgta cagggaagga agcgctggcc gtcggcaggt
     2521 gggggggcat gtcctggttc tgctcaggca gaaaagtccg aggattgagt tgcaggcagc
     2581 ccgcaaccag gttggtggtg ggttgggata agcccttgca aagcgtctga acgaaggaga
     2641 ccaggtctgg gcttttgcct gagcgcagga tctccgacag agcccagatg tagttcttgg
     2701 ccaagcgcag agtctcgatt ttggacagct tctgcgtctt agaatagcaa ggcaccacct
     2761 tgcgcaggtt gtctagcgcc gcgttcagtc cgtgcatgcg gttccgctcc cgggcgttag
     2821 ccttcatgcg tctcaattta aaacgctcca ggcgagcctt agtcatcttc ttctttttgg
     2881 ggccgcgtct cttgggcttt tgatcgtcat cctcctcttc ctcttcttcc tcctcttcca
     2941 ggtcctcatc ttcgtcctcc tcctctcccc cgttcctcag tgagtcctcc tctgcgttca
     3001 tggcttcgag gtcgtcctcc ttcttgtctg cctcgtgctc ctcgtcctga gaactgagac
     3061 actcgtctgt ccagcttgga ggaccttggg gctgaggctc gcccatcagc ccactctcgc
     3121 tgtacgattt ggtcatggtg gcggatccga attctagtat aacttcgtat aggatacttt
     3181 atacgaagtt atcattggga ttcttcctat tttgatccaa gcatcaccat cgaccctcta
     3241 gtccagatct caccatcgac ccataacttc gtatagcata cattatacga agttatgtcc
     3301 ctcgaagagg ttcgaattcg atatcaagct tatcgataat caacctctgg attacaaaat
     3361 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     3421 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     3481 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     3541 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     3601 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     3661 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     3721 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct
     3781 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     3841 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     3901 gatctccctt tgggccgcct ccccgcatcg ataccgtcga cccgggcggc cgcttcgagc
     3961 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     4021 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     4081 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggagatgtg
     4141 ggaggttttt taaagcaagt aaaacctcta caaatgtggt aaaatcgata aggatcttcc
     4201 tagagcatgg ctacgtagat aagtagcatg gcgggttaat cattaactac aaggaacccc
     4261 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     4321 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     4381 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4441 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4501 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4561 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4621 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4681 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4741 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4801 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4861 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4921 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4981 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5041 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5101 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     5161 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     5221 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5281 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5341 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5401 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5461 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5521 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5581 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5641 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5701 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5761 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5821 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5881 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5941 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6001 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6061 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6121 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     6181 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6241 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6301 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6361 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     6421 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     6481 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     6541 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     6601 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     6661 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     6721 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     6781 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg
     6841 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccag
//
