LOCUS       sequence-353970-        6881 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 6881)
  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 6881)
  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..6881
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     polyA_signal    292..413
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(329..348)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     383..402
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    456..1044
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             562..573
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     971..991
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    1086..1119
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1190..1223
                     /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)."
     CDS             2320..2373
                     /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      2380..2389
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2386..3096
                     /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(2531..2549)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2799..2818
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(3117..3150)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(3213..3246)
                     /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)."
     primer_bind     complement(3281..3300)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(3362..3384)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     repeat_region   4239..4368
                     /label=AAV2 ITR
     rep_origin      4440..4895
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4527..4546)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4737..4758
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4921..5025
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5026..5094,5095..5886)
                     /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(5244..5263)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6057..6645
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6546..6565
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcc ttatcgattt taccacattt gtagaggttt tacttgcttt aaaaaacctc
      241 ccacatctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
      301 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
      361 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
      421 tgctcgaagc ggccgcccgg gtcgacggta tcgatgcggg gaggcggccc aaagggagat
      481 ccgactcgtc tgagggcgaa ggcgaagacg cggaagaggc cgcagagccg gcagcaggcc
      541 gcgggaagga aggtccgctg gattgagggc cgaagggacg tagcagaagg acgtcccgcg
      601 cagaatccag gtggcaacac aggcgagcag ccaaggaaag gacgatgatt tccccgacaa
      661 caccacggaa ttgtcagtgc ccaacagccg agcccctgtc cagcagcggg caaggcaggc
      721 ggcgatgagt tccgccgtgg caatagggag ggggaaagcg aaagtcccgg aaaggagctg
      781 acaggtggtg gcaatgcccc aaccagtggg ggttgcgtca gcaaacacag tgcacaccac
      841 gccacgttgc ctgacaacgg gccacaactc ctcataaaga gacagcaacc aggatttata
      901 caaggaggag aaaatgaaag ccatacggga agcaatagca tgatacaaag gcattaaagc
      961 agcgtatcca catagcgtaa aaggagcaac atagttaaga ataccagtca atctttcaca
     1021 aattttgtaa tccagaggtt gattatcgat aagcttgata tcgaattcga acctcttcga
     1081 gggacataac ttcgtataat gtatgctata cgaagttatg ggtcgatggt gagatctgga
     1141 ctagagggtc gatggtgatg cttggatcaa aataggaaga atcccaatga taacttcgta
     1201 taaagtatcc tatacgaagt tatactagaa ttcggatccg ccaccatgac caaatcgtac
     1261 agcgagagtg ggctgatggg cgagcctcag ccccaaggtc ctccaagctg gacagacgag
     1321 tgtctcagtt ctcaggacga ggagcacgag gcagacaaga aggaggacga cctcgaagcc
     1381 atgaacgcag aggaggactc actgaggaac gggggagagg aggaggacga agatgaggac
     1441 ctggaagagg aggaagaaga ggaagaggag gatgacgatc aaaagcccaa gagacgcggc
     1501 cccaaaaaga agaagatgac taaggctcgc ctggagcgtt ttaaattgag acgcatgaag
     1561 gctaacgccc gggagcggaa ccgcatgcac ggactgaacg cggcgctaga caacctgcgc
     1621 aaggtggtgc cttgctattc taagacgcag aagctgtcca aaatcgagac tctgcgcttg
     1681 gccaagaact acatctgggc tctgtcggag atcctgcgct caggcaaaag cccagacctg
     1741 gtctccttcg ttcagacgct ttgcaagggc ttatcccaac ccaccaccaa cctggttgcg
     1801 ggctgcctgc aactcaatcc tcggactttt ctgcctgagc agaaccagga catgcccccc
     1861 cacctgccga cggccagcgc ttccttccct gtacacccct actcctacca gtcgcctggg
     1921 ctgcccagtc cgccttacgg taccatggac agctcccatg tcttccacgt taagcctccg
     1981 ccgcacgcct acagcgcagc gctggagccc ttctttgaaa gccctctgac tgattgcacc
     2041 agcccttcct ttgatggacc cctcagcccg ccgctcagca tcaatggcaa cttctctttc
     2101 aaacacgaac cgtccgccga gtttgagaaa aattatgcct ttaccatgca ctatcctgca
     2161 gcgacactgg caggggccca aagccacgga tcaatcttct caggcaccgc tgcccctcgc
     2221 tgcgagatcc ccatagacaa tattatgtcc ttcgatagcc attcacatca tgagcgagtc
     2281 atgagtgccc agctcaatgc catatttcat gatgctagcg aggggagagg aagtcttctg
     2341 acctgcggag acgtcgaaga gaatcctgga cccaccggtg ccaccatggt gagcaagggc
     2401 gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc
     2461 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
     2521 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc
     2581 ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc
     2641 gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag
     2701 gacggcggcg tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac
     2761 aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc
     2821 atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag
     2881 atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc
     2941 tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac
     3001 atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc
     3061 cactccaccg gcggcatgga cgagctgtac aagtagctcg aggggggatc cctagtataa
     3121 cttcgtatag catacattat acgaagttat gggtcgatgg tgatgcttgg caattcgggt
     3181 cgatggtgaa gcattggtct tcctattttg atataacttc gtataggata ctttatacga
     3241 agttatgggg atccaattct ttgccaaaat gatgagacag cacaataacc agcacgttgc
     3301 ccaggagctg taggaaaaag aagaaggcat gaacatggtt agcagaggct ctagagccgc
     3361 cggtcacacg ccagaagccg aaccccgccc tgccccgtcc cccccgaagg cagccgtccc
     3421 cccgcggaca gccccgagcc gctccccccg cacaaagggc cctcccggag cccctcaagg
     3481 ctttcacgca gccacagaaa agaaacaagc cgtcattaaa ccaagcgcta attacagccc
     3541 ggaggagaag ggccgtcccg cccgctcacc tgtgggagta acgcggtcag tcagagccgg
     3601 ggcgggcggc gcgaggcggc ggcggagcgg ggcacggggc gaaggcagcg cgcagcgact
     3661 cccgcccgcc gcgcgcttcg ctttttatag ggccgccgcc gccgccgcct cgccataaaa
     3721 ggaaactttc ggagcgcgcc gctctgattg gctgccgccg cacctctccg cctcgccccg
     3781 ccccgcccct cgccccgccc ccgcgccccc cccccccccc ccgcccccat cgctgcacaa
     3841 aataattaaa aaataaataa atacaaaatt gggggtgggg agggggggga gatggggaga
     3901 gtgaagcaga acgtggggct cacctcgacc atggtaatag cgatgactaa tacgtagatg
     3961 tactgccaag taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat
     4021 ttaccgtcat tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc
     4081 aagtgggcag tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc
     4141 gttactatgg gaacatacgt cattattgac gtcaatggta ctcttcctag agcatggcta
     4201 cgtagataag tagcatggcg ggttaatcat taactacaag gaacccctag tgatggagtt
     4261 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     4321 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct ggcgtaatag
     4381 cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggga
     4441 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     4501 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     4561 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4621 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     4681 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4741 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     4801 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4861 cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg
     4921 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     4981 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     5041 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     5101 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     5161 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     5221 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     5281 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     5341 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     5401 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     5461 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     5521 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     5581 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     5641 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     5701 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     5761 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     5821 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     5881 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     5941 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     6001 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     6061 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     6121 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     6181 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     6241 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     6301 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     6361 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     6421 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     6481 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     6541 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     6601 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     6661 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     6721 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     6781 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     6841 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca g
//
