LOCUS       sequence-237358-        2983 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMAGIC L1-R5 entry plasmid, contains empty human U6-driven SaCas9 
            gRNA scaffold for 4-component MultiSite Gateway Pro assembly. 
            Protospacer motif can be inserted after BsaI digestion..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2983)
  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 2983)
  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: JJ802: pMAGIC (L1-R5) hU6::SaCas9 gRNA scaffold
FEATURES             Location/Qualifiers
     source          1..2983
                     /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    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        677..925
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     677..697
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     848..867
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     protein_bind    1046..1169
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1224..1243)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1225..1243)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1248..1264)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1248..1264)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1366..1385)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1377..2186
                     /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(1444..1463)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2333..2921
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2822..2841
                     /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 tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctag gtacctgagg gcctatttcc catgattcct tcatatttgc atatacgata
      721 caaggctgtt agagagataa ttggaattaa tttgactgta aacacaaaga tattagtaca
      781 aaatacgtga cgtagaaagt aataatttct tgggtagttt gcagttttaa aattatgttt
      841 taaaatggac tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct tggctttata
      901 tatcttgtgg aaaggacgaa acaccggaga ccacggcagg tctcagttta agtactctgt
      961 gctggaaaca gcacagaatc tactaaacaa ggcaaaatgc cgtgtttatc tcgtcaactt
     1021 gttggcgaga tttttgcggc cgctgcaact ttgtatacaa aagttgaacg agaaacgtaa
     1081 aatgatataa atatcaatat attaaattag attttgcata aaaaacagac tacataatac
     1141 tgtaaaacac aacatatcca gtcactatga atcaactact tagatggtat tagtgacctg
     1201 tactgcaggg cggccgcgat atcccctata gtgagtcgta ttacatggtc atagctgttt
     1261 cctggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa gataaaaata
     1321 tatcatcatg aacaataaaa ctgtctgctt acataaacag taatacaagg ggtgttatga
     1381 gccatattca acgggaaacg tcgaggccgc gattaaattc caacatggat gctgatttat
     1441 atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgcttgt
     1501 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg
     1561 atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca
     1621 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atccccggaa
     1681 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc
     1741 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg
     1801 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga
     1861 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata
     1921 aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc
     1981 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag
     2041 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac
     2101 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc
     2161 atttgatgct cgatgagttt ttctaatcag aattggttaa ttggttgtaa cactggcaga
     2221 gcattacgct gacttgacgg gacggcgcaa gctcatgacc aaaatccctt aacgtgagtt
     2281 acgcgtcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     2341 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     2401 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     2461 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     2521 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     2581 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     2641 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     2701 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     2761 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     2821 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     2881 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     2941 ttttacggtt cctggccttt tgctggcctt ttgctcacat gtt
//
