LOCUS       sequence-237364-        3052 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pMAGIC L1-R5 entry plasmid, contains empty mouse 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 3052)
  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 3052)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: LG002: pMAGIC (L1-R5) mU6::xCas9(3.7) gRNA sca...
FEATURES             Location/Qualifiers
     source          1..3052
                     /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        671..984
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     903..923
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    1115..1238
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1293..1312)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1294..1312)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1317..1333)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1317..1333)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1435..1454)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1446..2255
                     /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(1513..1532)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2402..2990
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2891..2910
                     /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 agcaggctta gatccgacgc cgccatctct aggcccgcgc cggccccctc gcacagactt
      721 gtgggagaag ctcggctact cccctgcccc ggttaatttg catataatat ttcctagtaa
      781 ctatagaggc ttaatgtgcg ataaaagaca gataatctgt tctttttaat actagctaca
      841 ttttacatga taggcttgga tttctataag agatacaaat actaaattat tattttaaaa
      901 aacagcacaa aaggaaactc accctaactg taaagtaatt gtgtgttttg agactataaa
      961 tatcccttgg agaaaagcct tgttggagac cacggcaggt ctcagtttca gagctatgct
     1021 ggaaacagca tagcaagttg aaataaggct agtccgttat caacttgaaa aagtggcacc
     1081 gagtcggtgc ttttttgcgg ccgcaggaag cttacaactt tgtatacaaa agttgaacga
     1141 gaaacgtaaa atgatataaa tatcaatata ttaaattaga ttttgcataa aaaacagact
     1201 acataatact gtaaaacaca acatatccag tcactatgaa tcaactactt agatggtatt
     1261 agtgacctgt actgcagggc ggccgcgata tcccctatag tgagtcgtat tacatggtca
     1321 tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct ctgatgttac attgcacaag
     1381 ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg
     1441 gtgttatgag ccatattcaa cgggaaacgt cgaggccgcg attaaattcc aacatggatg
     1501 ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct
     1561 atcgcttgta tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg
     1621 ttgccaatga tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc
     1681 ttccgaccat caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga
     1741 tccccggaaa aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg
     1801 ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt
     1861 ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg
     1921 ttgatgcgag tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag
     1981 aaatgcataa acttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac
     2041 ttgataacct tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg
     2101 gaatcgcaga ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc
     2161 cttcattaca gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat
     2221 tgcagtttca tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac
     2281 actggcagag cattacgctg acttgacggg acggcgcaag ctcatgacca aaatccctta
     2341 acgtgagtta cgcgtcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     2401 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     2461 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     2521 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     2581 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     2641 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     2701 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     2761 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     2821 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     2881 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     2941 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     3001 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg tt
//
