LOCUS       sequence-486316-        2209 bp    DNA     circular SYN 16-SEP-2025
DEFINITION  sgRNA-E compatible with TCTP-SynPro-04, TCTP-SynPro-06 and CaMV 
            35S-SynPro-03 as Level-0 part (Golden Gate cloning).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2209)
  AUTHORS   Khan MA, Herring G, Zhu JY, Oliva M, Fourie E, Johnston B, Zhang Z, 
            Potter J, Pineda L, Pflueger J, Swain T, Pflueger C, Lloyd JPB, 
            Secco D, Small I, Kidd BN, Lister R
  TITLE     CRISPRi-based circuits to control gene expression in plants.
  JOURNAL   Nat Biotechnol. 2025 Mar;43(3):416-430. doi: 
            10.1038/s41587-024-02236-w. Epub 2024 May 20.
  PUBMED    38769424
REFERENCE   2  (bases 1 to 2209)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 16, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41587-024-02236-w"; journalName: "Nat Biotechnol"; date: 
            "2025-03"; volume: "43"; issue: "3"; pages: "416-430"
COMMENT     Sequence Label: LLP995
FEATURES             Location/Qualifiers
     source          1..2209
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             285..1076
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1417..2005
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1906..1925
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    2058..2077
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        2098..2173
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    2174..2193
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
ORIGIN
        1 gtgccgaggc acgaacccag tggacataag cctgttcggt tcgtaagctg taatgcaagt
       61 agcgtatgcg ctcacgcaac tggtccagaa ccttgaccga acgcagcggt ggtaacggcg
      121 cagtggcggt tttcatggct tgttatgact gtttttttgg ggtacagtct atgcctcggg
      181 catccaagca gcaagcgcgt tacgccgtgg gtcgatgttt gatgttatgg agcagcaacg
      241 atgttacgca gcagggcagt cgccctaaaa caaagttaaa catcatgggg gaagcggtga
      301 tcgccgaagt atcgactcaa ctatcagagg tagttggcgt catcgagcgc catctcgaac
      361 cgacgttgct ggccgtacat ttgtacggct ccgcagtgga tggcggcctg aagccacaca
      421 gcgatattga tttgctggtt acggtgaccg taaggcttga tgaaacaacg cggcgagctt
      481 tgatcaacga ccttttggaa acttcggctt cccctggaga gagcgagatt ctccgcgctg
      541 tagaagtcac cattgttgtg cacgacgaca tcattccgtg gcgttatcca gctaagcgcg
      601 aactgcaatt tggagaatgg cagcgcaatg acattcttgc aggtatcttc gagccagcca
      661 cgatcgacat tgatctggct atcttgctga caaaagcaag agaacatagc gttgccttgg
      721 taggtccagc ggcggaggaa ctctttgatc cggttcctga acaggatcta tttgaggcgc
      781 taaatgaaac cttaacgcta tggaactcgc cgcccgactg ggctggcgat gagcgaaatg
      841 tagtgcttac gttgtcccgc atttggtaca gcgcagtaac cggcaaaatc gcgccgaagg
      901 atgtcgctgc cgactgggca atggagcgcc tgccggccca gtatcagccc gtcatacttg
      961 aagctagaca ggcttatctt ggacaagaag aagatcgctt ggcctcgcgc gcagatcagt
     1021 tggaagaatt tgtccattac gtgaaaggcg agatcaccaa ggtagtcggc aaataatgtc
     1081 tagctagaaa ttcgttcaag ccgacgccgc ttcgcggcgc ggcttaactc aagcgttaga
     1141 tgcactaagc acataattgc tcacagccaa actatcaggt caagtctgct tttattattt
     1201 ttaagcgtgc ataataagcc ctacacaaat tgggagatat atcatgctgt cagaccaagt
     1261 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     1321 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     1381 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     1441 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     1501 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     1561 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     1621 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     1681 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     1741 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     1801 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     1861 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     1921 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     1981 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctgca
     2041 ctctgtggtc tcatactgtt cactgccgta taggcagcat tgccatacac cttgagggtt
     2101 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
     2161 accgagtcgg tgcgttcact gccgtatagg caggcttcga gaccacgaa
//
