LOCUS       sequence-203127-        3392 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express single gRNA with gRNA2.0 scaffold (with four MS2 binding 
            sites) under OsU6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3392)
  AUTHORS   Lowder LG, Zhou J, Zhang Y, Malzahn A, Zhong Z, Hsieh TF, Voytas DF,
            Zhang Y, Qi Y
  TITLE     Robust transcriptional activation in plants using multiplexed 
            CRISPR-Act2.0 and mTALE-Act systems.
  JOURNAL   Mol Plant. 2017 Nov 29. pii: S1674-2052(17)30344-1. doi: 
            10.1016/j.molp.2017.11.010.
  PUBMED    29197638
REFERENCE   2  (bases 1 to 3392)
  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: "Mol Plant. 
            2017 Nov 29. pii: S1674-2052(17)30344-1. doi: 
            10.1016/j.molp.2017.11.010."
COMMENT     Sequence Label: pYPQ141C2.0
FEATURES             Location/Qualifiers
     source          1..3392
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     3..25
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     17..34
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     18..34
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    49..144
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_RNA        510..528
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        580..598
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     protein_bind    complement(717..816)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(834..852)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(869..887)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(892..908)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(892..908)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1339..2130
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      2223..2811
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2712..2731
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2965..2982
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3141..3168)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3260..3346)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttcccagtca cgacgttgta aaacgacggc cagtcttaag ctcgggccca ataatgattt
       61 tattttgact gatagtgacc tgttcgttgc aacaaattga tgagcaatgc ttttttataa
      121 tgccaacttt gtatacaaaa gttgccccat ggcgttccct ctagataacg caggatccac
      181 tagtaacggc cgccagtgtg ctggaattgc ccttggatca tgaaccaacg gcctggctgt
      241 atttggtggt tgtgtaggga gatggggaga agaaaagccc gattctcttc gctgtgatgg
      301 gctggatgca tgcgggggag cgggaggccc aagtacgtgc acggtgagcg gcccacaggg
      361 cgagtgtgag cgcgagaggc gggaggaaca gtttagtacc acattgccca gctaactcga
      421 acgcgaccaa cttataaacc cgcgcgctgt cgcttgtgtg gagacgagat ctagtctgag
      481 tcgaccgtct ctgttttaga gctaggccaa catgaggatc acccatgtct gcagggccta
      541 gcaagttaaa ataaggctag tccgttatca acttggccaa catgaggatc acccatgtct
      601 gcagggccaa gtggcaccga gtcggtgctt tttttgtccc ttcgaagggc aattctgcag
      661 atatccatca cactggcggc cgctcgaggt cgagggtatc gataagcttg aattcgaccc
      721 agctttcttg tacaaagttg gcattataaa aaataattgc tcatcaattt gttgcaacga
      781 acaggtcact atcagtcaaa ataaaatcat tatttgccat ccagctgata tcccctatag
      841 tgagtcgtat tacatggtca tagcatcccc tatagtgagt cgtattacat ggtcatagct
      901 gtttcctggc agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagataaa
      961 aatatatcat catgcctcct ctggaccagc caggacagaa atgcctcgac ttcgctgcta
     1021 cccaaggttg ccgggtgacg cacaccgtgg aaacggatga aggcacgaac ccagtggaca
     1081 taagcctgtt cggttcgtaa gctgtaatgc aagtagcgta tgcgctcacg caactggtcc
     1141 agaaccttga ccgaacgcag cggtggtaac ggcgcagtgg cggttttcat ggcttgttat
     1201 gactgttttt ttggggtaca gtctatgcct cgggcatcca agcagcaagc gcgttacgcc
     1261 gtgggtcgat gtttgatgtt atggagcagc aacgatgtta cgcagcaggg cagtcgccct
     1321 aaaacaaagt taaacattat gagggaagcg gtgatcgccg aagtatcgac tcaactatca
     1381 gaggtagttg gcgtcatcga gcgccatctc gaaccgacgt tgctggccgt acatttgtac
     1441 ggctccgcag tggatggcgg cctgaagcca cacagtgata ttgatttgct ggttacggtg
     1501 accgtaaggc ttgatgaaac aacgcggcga gctttgatca acgacctttt ggaaacttcg
     1561 gcttcccctg gagagagcga gattctccgc gctgtagaag tcaccattgt tgtgcacgac
     1621 gacatcattc cgtggcgtta tccagctaag cgcgaactgc aatttggaga atggcagcgc
     1681 aatgacattc ttgcaggtat cttcgagcca gccacgatcg acattgatct ggctatcttg
     1741 ctgacaaaag caagagaaca tagcgttgcc ttggtaggtc cagcggcgga ggaactcttt
     1801 gatccggttc ctgaacagga tctatttgag gcgctaaatg aaaccttaac gctatggaac
     1861 tcgccgcccg actgggctgg cgatgagcga aatgtagtgc ttacgttgtc ccgcatttgg
     1921 tacagcgcag taaccggcaa aatcgcgccg aaggatgtcg ctgccgactg ggcaatggag
     1981 cgcctgccgg cccagtatca gcccgtcata cttgaagcta gacaggctta tcttggacaa
     2041 gaagaagatc gcttggcctc gcgcgcagat cagttggaag aatttgtcca ctacgtgaaa
     2101 ggcgagatca ccaaggtagt cggcaaataa ccctcgagcc acccatgacc aaaatccctt
     2161 aacgtgagtt acgcgtcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2221 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2281 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2341 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     2401 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     2461 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     2521 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     2581 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     2641 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     2701 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     2761 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     2821 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     2881 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     2941 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
     3001 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     3061 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atacgcgtac cgcgagccag
     3121 gaagagtttg tagaaacgca aaaaggccat ccgtcaggat ggccttctgc ttagtttgat
     3181 gcctggcagt ttatggcggg cgtcctgccc gccaccctcc gggccgttgc ttcacaacgt
     3241 tcaaatccgc tcccggcgga tttgtcctac tcaggagagc gttcaccgac aaacaacaga
     3301 taaaacgaaa ggcccagtct tccgactgag cctttcgttt tatttgatgc ctggcagttc
     3361 cctactctcg cgttaacgct tgcatggatg tt
//
