LOCUS       sequence-237365-        2993 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMAGIC L1-R5 entry plasmid, contains empty human U6-driven 
            xCas9(3.7) (i.e. SpCas9) 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 2993)
  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 2993)
  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: LF701: pMAGIC (L1-R5) hU6::xCas9(3.7) gRNA sca...
FEATURES             Location/Qualifiers
     source          1..2993
                     /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    1056..1179
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1234..1253)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1235..1253)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1258..1274)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1258..1274)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1376..1395)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1387..2196
                     /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(1454..1473)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2343..2931
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2832..2851
                     /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 tctcagtttc agagctatgc
      961 tggaaacagc atagcaagtt gaaataaggc tagtccgtta tcaacttgaa aaagtggcac
     1021 cgagtcggtg cttttttgcg gccgcaggaa gcttacaact ttgtatacaa aagttgaacg
     1081 agaaacgtaa aatgatataa atatcaatat attaaattag attttgcata aaaaacagac
     1141 tacataatac tgtaaaacac aacatatcca gtcactatga atcaactact tagatggtat
     1201 tagtgacctg tactgcaggg cggccgcgat atcccctata gtgagtcgta ttacatggtc
     1261 atagctgttt cctggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa
     1321 gataaaaata tatcatcatg aacaataaaa ctgtctgctt acataaacag taatacaagg
     1381 ggtgttatga gccatattca acgggaaacg tcgaggccgc gattaaattc caacatggat
     1441 gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc
     1501 tatcgcttgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc
     1561 gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct
     1621 cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg
     1681 atccccggaa aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt
     1741 gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct
     1801 tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg
     1861 gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa
     1921 gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca
     1981 cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc
     2041 ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct
     2101 ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa
     2161 ttgcagtttc atttgatgct cgatgagttt ttctaatcag aattggttaa ttggttgtaa
     2221 cactggcaga gcattacgct gacttgacgg gacggcgcaa gctcatgacc aaaatccctt
     2281 aacgtgagtt acgcgtcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2341 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2401 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2461 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     2521 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     2581 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     2641 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     2701 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     2761 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     2821 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     2881 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     2941 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gtt
//
