LOCUS       sequence-152040-        1024 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  tRNA and scaffold for the assembly of GBoligomers for the first 
            position (positon [M1_2]) of a polycistronic tRNA-gRNA regulated by 
            the (monocot) U3 promoter (3-part multiplexing).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1024)
  AUTHORS   Vazquez-Vilar M, Bernabe-Orts JM, Fernandez-Del-Carmen A, Ziarsolo 
            P, Blanca J, Granell A, Orzaez D
  TITLE     A modular toolbox for gRNA-Cas9 genome engineering in plants based 
            on the GoldenBraid standard.
  JOURNAL   Plant Methods. 2016 Feb 1;12:10. doi: 10.1186/s13007-016-0101-2. 
            eCollection 2016.
  PUBMED    26839579
REFERENCE   2  (bases 1 to 1024)
  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: "Plant 
            Methods."; date: "2016-02-1"; volume: "12:10. doi"; pages: " 
            10.1186/s13007-016-0101-2. eCollection 2016"
COMMENT     Sequence Label: tRNA-gRNA position [M1_2] (GB1209)
FEATURES             Location/Qualifiers
     source          1..1024
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     misc_RNA        175..250
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      279..288
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      283..292
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(322..341)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(323..341)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(348..365)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(348..364)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(357..379)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      505..960
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(592..611)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     802..823
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 acgcgttggg agctctccca tatggtcgac ctgcaggcgg ccgcgaattc actagtgatc
       61 ccgtctcgct cgggcaaaca aagcaccagt ggtctagtgg tagaatagta ccctgccacg
      121 gtacagaccc gggttcgatt cccggctggt gcagagacgg cgcgcgccgt ctcagtttta
      181 gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc
      241 gagtcggtgc tgagcgagac gggaatcgaa ttcccgcggc cgccatggcg gccgggagca
      301 tgcgacgtcg ggcccaattc gccctatagt gagtcgtatt acaattcact ggccgtcgtt
      361 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
      421 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
      481 ttgcgcagcc tgaatggcga atggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
      541 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
      601 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
      661 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
      721 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
      781 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
      841 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
      901 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
      961 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tcaggtggca
     1021 cttt
//
