LOCUS       sequence-237394-        4443 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMAGIC R4-R3 entry plasmid, contains 2x NLS/Dnmt3a-3L scaffold 
            enabling addition of dCas9 mutants into pMAGIC. dCas9 can be 
            inserted via unique SrfI/PmeI restriction sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4443)
  AUTHORS   Haldeman JM, Conway AE, Arlotto ME, Slentz DH, Muoio DM, Becker TC, 
            Newgard CB
  TITLE     Creation of versatile cloning platforms for transgene expression and
            dCas9-based epigenome editing.
  JOURNAL   Nucleic Acids Res. 2018 Dec 27. pii: 5264291. doi: 
            10.1093/nar/gky1286.
  PUBMED    30590691
REFERENCE   2  (bases 1 to 4443)
  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: "Nucleic 
            Acids Res. 2018 Dec 27. pii: 5264291. doi: 10.1093/nar/gky1286."
COMMENT     Sequence Label: KL401: pMAGIC (R4-R3) NLS-(SrfI/PmeI)-NLS-Dnmt...
FEATURES             Location/Qualifiers
     source          1..4443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(603..726)
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      726..735
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             738..758
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             813..860
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     protein_bind    2506..2629
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2684..2703)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2685..2703)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2708..2724)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2708..2724)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2826..2845)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2837..3646
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2904..2923)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3793..4381
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4282..4301
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccct acaggtcact aataccatct aagtagttga
      601 ttcatagtga ctggatatgt tgtgttttac agtattatgt agtctgtttt ttatgcaaaa
      661 tctaatttaa tatattgata tttatatcat tttacgtttc tcgttcaact tttctataca
      721 aagttgccac catggcccca aagaagaagc ggaaggtcgg tatccacgga gtcccagcag
      781 cccgggcgag accatgctcg cggtctcagt ttaaacggcc ggcagccacg aagaaggccg
      841 gccaggcaaa aaagaaaaag ggagggggag gcagcaacca tgaccaggaa tttgaccctc
      901 caaaggttta cccacctgtg ccagctgaga agaggaagcc catccgcgtg ctgtctctct
      961 ttgatgggat tgctacaggg ctcctggtgc tgaaggacct gggcatccaa gtggaccgct
     1021 acattgcctc cgaggtgtgt gaggactcca tcacagtggg catggtgcgg caccagggaa
     1081 agatcatgta cgtcggggac gtccgcagcg tcacacagaa gcatatccag gagtggggcc
     1141 cattcgacct ggtgattgga ggcagtccct gcaatgacct ctccattgtc aaccctgccc
     1201 gcaagggact ttatgagggt actggccgcc tcttctttga gttctaccgc ctcctgcatg
     1261 atgctcggcc caaggaggga gatgatcgcc ccttcttctg gctctttgag aatgtggtgg
     1321 ccatgggcgt tagtgacaag agggacatct cccgatttct tgagtctaac cccgtgatga
     1381 ttgacgccaa agaagtgtct gctgcacaca gggccagata cttctggggt aaccttcctg
     1441 gcatgaacag gcctttggca tccactgtga atgataagct ggagctgcaa gagtgtctgg
     1501 agcacggcag aatagccaag ttcagcaaag tgaggaccat taccaccagg tcaaactcta
     1561 taaagcaggg caaagaccag catttccccg tcttcatgaa cgagaaggag gacatcctgt
     1621 ggtgcactga aatggaaagg gtgtttggct tccccgtcca ctacacagac gtctccaaca
     1681 tgagccgctt ggccaggcag agactgctgg gccgatcctg gagcgtgccg gtcatccgcc
     1741 acctcttcgc tccgctgaag gaatattttg cttgtgtgag cagcggcaac agcaacgcca
     1801 acagccgcgg ccccagcttc agcagcggcc tggtgcccct gagcctgcgc ggcagccaca
     1861 tgggccctat ggagatatac aagacagtgt ctgcatggaa gagacagcca gtgcgggtgc
     1921 tgagcctgtt cagaaacatc gataaagtgc tgaagagttt gggcttcttg gaaagcggtt
     1981 ctggttctgg gggaggaacc ctgaagtacg tggaagatgt cacaaatgtc gtgaggagag
     2041 acgtggagaa atggggcccc tttgacctgg tgtacggctc cactcagccc ctgggcagct
     2101 cttgtgatcg ctgtcccggc tggtacatgt tccagttcca ccgaatcctg cagtatgccc
     2161 tgcctcgcca ggagagtcag cggcccttct tctggatatt catggacaat ctgctgctga
     2221 ctgaggatga ccaagagaca actacccgct tccttcagac agaggctgtg accctccagg
     2281 atgtcagagg cagagactac cagaatgcta tgcgggtgtg gagcaacatt ccagggctga
     2341 agagcaagca tgctcccctg accccaaagg aagaagagta tctgcaagcc caagtcagaa
     2401 gcaggagcaa gctggacgcc ccgaaagttg acctcctggt gaagaactgc cttctcccgc
     2461 tgagagagta cttcaagtat ttttctcaaa actcacttcc tctcgcaact ttgtataata
     2521 aagttgaacg agaaacgtaa aatgatataa atatcaatat attaaattag attttgcata
     2581 aaaaacagac tacataatac tgtaaaacac aacatatcca gtcactatga atcaactact
     2641 tagatggtat tagtgacctg tactgcaggg cggccgcgat atcccctata gtgagtcgta
     2701 ttacatggtc atagctgttt cctggcagct ctggcccgtg tctcaaaatc tctgatgtta
     2761 cattgcacaa gataaaaata tatcatcatg aacaataaaa ctgtctgctt acataaacag
     2821 taatacaagg ggtgttatga gccatattca acgggaaacg tcgaggccgc gattaaattc
     2881 caacatggat gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg
     2941 tgcgacaatc tatcgcttgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg
     3001 caaaggtagc gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga
     3061 atttatgcct cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact
     3121 caccactgcg atccccggaa aaacagcatt ccaggtatta gaagaatatc ctgattcagg
     3181 tgaaaatatt gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg
     3241 taattgtcct tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa
     3301 taacggtttg gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca
     3361 agtctggaaa gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg
     3421 tgatttctca cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt
     3481 tggacgagtc ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg
     3541 tgagttttct ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga
     3601 tatgaataaa ttgcagtttc atttgatgct cgatgagttt ttctaatcag aattggttaa
     3661 ttggttgtaa cactggcaga gcattacgct gacttgacgg gacggcgcaa gctcatgacc
     3721 aaaatccctt aacgtgagtt acgcgtcgtt ccactgagcg tcagaccccg tagaaaagat
     3781 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     3841 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     3901 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     3961 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     4021 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     4081 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     4141 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     4201 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     4261 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     4321 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     4381 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4441 gtt
//
