LOCUS       sequence-151794-         943 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  tRNA and scaffold for the assembly of GBoligomers for the first 
            position (positon [D1_2]) of a polycistronic tRNA-gRNA regulated by 
            the dicot U6-26 or U6-1 promoter (3-part multiplexing).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 943)
  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 943)
  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 [D1_2] (GB1205)
FEATURES             Location/Qualifiers
     source          1..943
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     misc_RNA        170..245
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      274..283
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      278..287
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(317..336)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(318..336)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(343..360)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(343..359)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(352..374)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      500..928
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(587..606)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     797..818
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ttgggagctc tcccatatgg tcgacctgca ggcggccgcg aattcactag tgatcccgtc
       61 tcgctcgatt gaacaaagca ccagtggtct agtggtagaa tagtaccctg ccacggtaca
      121 gacccgggtt cgattcccgg ctggtgcaga gacggcgcgc gccgtctcag ttttagagct
      181 agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
      241 ggtgctgagc gagacgggaa tcgaattccc gcggccgcca tggcggccgg gagcatgcga
      301 cgtcgggccc aattcgccct atagtgagtc gtattacaat tcactggccg tcgttttaca
      361 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
      421 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
      481 cagcctgaat ggcgaatgga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
      541 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
      601 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
      661 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
      721 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
      781 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
      841 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
      901 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taa
//
