LOCUS       sequence-354349-        6400 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The pre-miR-9/9* and pre-miR-124 sequences have been inserted into 
            the EcoRV of pCAG-nDsRedFL-intron..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6400)
  AUTHORS   Suzuki F, Okuno M, Tanaka T, Sanuki R
  TITLE     Overexpression of neural miRNAs miR-9/9* and miR-124 suppresses 
            differentiation to Muller glia and promotes differentiation to 
            neurons in mouse retina in vivo.
  JOURNAL   Genes Cells. 2020 Nov;25(11):741-752. doi: 10.1111/gtc.12809. Epub 
            2020 Oct 21.
  PUBMED    32979863
REFERENCE   2  (bases 1 to 6400)
  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: "10.1111/gtc.12809";
            journalName: "Genes Cells"; date: "2020-11"; volume: "25"; issue: 
            "11"; pages: "741-752"
COMMENT     Sequence Label: pCAG-nDsRedFL-miR9/9*/124
FEATURES             Location/Qualifiers
     source          1..6400
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          647..1663
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1587..1609
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1671..1690
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1727..1736
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1739..1759
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(1943..1963)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     3261..3284
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             3350..3415
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3431..3450)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3496..3551
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3550..3569)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3912..3928)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3912..3928)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3925..3947)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3936..3952
                     /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)."
     promoter        complement(join(3960..3966,3967..3984,3985..3990))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4005..4026
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4085..4280
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4131..4266
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4286..4420
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4323..4342)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4377..4396
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4488..4505)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4659..5247)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4739..4758)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5418..6209,6210..6278))
                     /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     6041..6060
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6279..6383)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgccacctgg gtcgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc
      421 actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta
      481 ttttgtgcag cgatgggggc gggggggggg ggggggcgsg gggcggggcg gggcgaggsg
      541 grgrggkgsg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
      601 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg
      661 ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact
      721 gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta
      781 gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct
      841 ccgggagggc cctttgtgcg gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt
      901 ggggagcgcc gcgtgcggct ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc
      961 ggggctttgt gcgctccgca gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg
     1021 gtgcgggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg
     1081 agcagggggt gtgggcgcgt cggtcgggct gcaacccccc ctgcaccccc ctccccgagt
     1141 tgctgagcac ggcccggctt cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc
     1201 cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc
     1261 cggggagggc tcgggggagg ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc
     1321 ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg gacttccttt
     1381 gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg
     1441 cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc
     1501 gccgcgccgc cgtccccttc tccctctcca gcctcggggc tgtccgcggg gggacggctg
     1561 ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg gcggctctag
     1621 agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg gcaacgtgct
     1681 ggttattgtg ctgtctcatc attttggcaa agaattcacc cgggaagcca ccatggctcc
     1741 aaagaagaag cgtaaggtac ggtacgcctc ctccgagaac gtcatcaccg agttcatgcg
     1801 cttcaaggtg cgcatggagg gcaccgtgaa cggccacgag ttcgagatcg agggcgaggg
     1861 cgagggccgc ccctacgagg gccacaacac cgtgaagctg aaggtgacca agggcggccc
     1921 cctgcccttc gcctgggaca tcctgtcccc ccagttccag tacggctcca aggtgtacgt
     1981 gaagcacccc gccgacatcc ccgactacaa gaagctgtcc ttccccgagg gcttcaagtg
     2041 ggagcgcgtg atgaacttcg aggacggcgg cgtggcgacc gtgacccagg actcctccct
     2101 gcaggacggc tgcttcatct acaaggtgaa gttcatcggc gtgaacttcc cctccgacgg
     2161 ccccgtgatg cagaagaaaa ctatgggctg ggaggcctcc accgagcgcc tgtacccccg
     2221 cgacggcgtg ctgaagggcg agacccacaa ggccctgaag ctgaaggacg gcggccacta
     2281 cctggtggag ttcaagtcca tctacatggc caagaagccc gtgcaggtga gtactccctc
     2341 tcaaaagcgg gcatgacttc tgcgctaaga ttgtcagttt ccaaaaacga ggaggatttg
     2401 atattcacct ggcccgcggt gatgcctttg agggatacgc gtcggcggcc tgtcggtccc
     2461 ctctggctct ccgcgtgcgc ccggggctgc gggagcggag tccgctcagg cccccgcgcc
     2521 gggtcggcgg cggcacgtgg ggcccacagc gcggcagcgg tgccggcgaa tgggaggccc
     2581 gtttctctct ttggttatct agctgtatga gtgccacaga gccgtcataa agctagataa
     2641 ccgaaagtag aaatgactct cacaacttct gcgtgcgatg gcccggccgg ccgccctggc
     2701 cacaggagcc agaggagccc agagaagccc acgccaccca gggattccag ctcgacggag
     2761 cggccacgcc gggaccaggt atccgacctc ggctctcctt ctagtaggtg ggagtactgc
     2821 tcagagctac aactctagga gtagggactc caagcctaga gctccaagag agggtgaagg
     2881 gcagggagaa aattatagta atagttgcaa tgagtcactt gcttctagat caagatcaga
     2941 gactctgctc tccgtgttca cagcggacct tgatttaatg tcatacaatt aaggcacgcg
     3001 gtgaatgcca agagcggagc ctacagctgc acttgaagga catccgagag aagttaggaa
     3061 gggtggggag aaacaattct agaatgaacc catcctgtgc gacacgatcg tggccgcgtc
     3121 catctggtca gaaaagacaa tctttttgtt gtcaacgttg aggtgtgaca ggcttgagag
     3181 ctggccatac acttgagtga caatgacatc cactttgcct ttctctccac agctgcccgg
     3241 ctactactac gtggacgcca agctggacat cacctcccac aacgaggact acaccatcgt
     3301 ggagcagtac gagcgcaccg agggccgcca ccacctgttc ctgggtaccg actataagga
     3361 ccacgacgga gactacaagg atcatgatat tgattacaaa gacgatgacg ataagtaact
     3421 cgagtgtaga tctttttccc tctgccaaaa attatgggga catcatgaag ccccttgagc
     3481 atctgacttc tggctaataa aggaaattta ttttcattgc aatagtgtgt tggaattttt
     3541 tgtgtctctc actcggaagg acatatggga gggcaaatca tttaaaacat cagaatgagt
     3601 atttggttta gagtttggca acatatgccc atatgctggc tgccatgaac aaaggttggc
     3661 tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta ttccatagaa
     3721 aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt ttttctttaa
     3781 catccctaaa attttcctta catgttttac tagccagatt tttcctcctc tcctgactac
     3841 tcccagtcat agctgtccct cttctcttat ggagatccct cgacctgcag cccaagcttg
     3901 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3961 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     4021 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagcgg
     4081 atccgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     4141 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     4201 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     4261 gaggcctagg cttttgcaaa aagctaactt gtttattgca gcttataatg gttacaaata
     4321 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4381 tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cgctgcatta atgaatcggc
     4441 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4501 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4561 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4621 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4681 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4741 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4801 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4861 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4921 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4981 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5041 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     5101 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5161 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5221 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5281 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5341 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5401 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5461 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5521 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5581 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5641 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     5701 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     5761 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5821 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5881 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5941 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     6001 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     6061 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     6121 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     6181 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     6241 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6301 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6361 aataaacaaa taggggttcc gcgcacattt ccccgaaaag
//
