LOCUS       sequence-203128-        3490 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express single gRNA with gRNA2.0 scaffold (with four MS2 binding 
            sites) under OsU3 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3490)
  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 3490)
  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: pYPQ141D2.0
FEATURES             Location/Qualifiers
     source          1..3490
                     /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"
     promoter        181..558
                     /label=OsU3 promoter
                     /note="Oryza sativa (rice) snRNA U3 promoter"
     misc_RNA        608..626
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        678..696
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     protein_bind    complement(815..914)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(932..950)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(967..985)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(990..1006)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(990..1006)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1437..2228
                     /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      2321..2909
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2810..2829
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3063..3080
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3239..3266)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3358..3444)
                     /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 caggatccta
      181 aagggatctt taaacatacg aacagatcac ttaaagttct tctgaagcaa cttaaagtta
      241 tcaggcatgc atggatcttg gaggaatcag atgtgcagtc agggaccata gcacaggaca
      301 ggcgtcttct actggtgcta ccagcaaatg ctggaagccg ggaacactgg gtacgttgga
      361 aaccacgtga tgtggagtaa gataaactgt aggagaaaag catttcgtag tgggccatga
      421 agcctttcag gacatgtatt gcagtatggg ccggcccatt acgcaattgg acgacaacaa
      481 agactagtat tagtaccacc tcggctatcc acatagatca aagctggttt aaaagagttg
      541 tgcagatgat ccgtggcaga gacgagatct agtctgagtc gaccgtctct gttttagagc
      601 taggccaaca tgaggatcac ccatgtctgc agggcctagc aagttaaaat aaggctagtc
      661 cgttatcaac ttggccaaca tgaggatcac ccatgtctgc agggccaagt ggcaccgagt
      721 cggtgctttt tttgtccctt cgaagggcaa ttctgcagat atccatcaca ctggcggccg
      781 ctcgaggtcg agggtatcga taagcttgaa ttcgacccag ctttcttgta caaagttggc
      841 attataaaaa ataattgctc atcaatttgt tgcaacgaac aggtcactat cagtcaaaat
      901 aaaatcatta tttgccatcc agctgatatc ccctatagtg agtcgtatta catggtcata
      961 gcatccccta tagtgagtcg tattacatgg tcatagctgt ttcctggcag ctctggcccg
     1021 tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca tgcctcctct
     1081 ggaccagcca ggacagaaat gcctcgactt cgctgctacc caaggttgcc gggtgacgca
     1141 caccgtggaa acggatgaag gcacgaaccc agtggacata agcctgttcg gttcgtaagc
     1201 tgtaatgcaa gtagcgtatg cgctcacgca actggtccag aaccttgacc gaacgcagcg
     1261 gtggtaacgg cgcagtggcg gttttcatgg cttgttatga ctgttttttt ggggtacagt
     1321 ctatgcctcg ggcatccaag cagcaagcgc gttacgccgt gggtcgatgt ttgatgttat
     1381 ggagcagcaa cgatgttacg cagcagggca gtcgccctaa aacaaagtta aacattatga
     1441 gggaagcggt gatcgccgaa gtatcgactc aactatcaga ggtagttggc gtcatcgagc
     1501 gccatctcga accgacgttg ctggccgtac atttgtacgg ctccgcagtg gatggcggcc
     1561 tgaagccaca cagtgatatt gatttgctgg ttacggtgac cgtaaggctt gatgaaacaa
     1621 cgcggcgagc tttgatcaac gaccttttgg aaacttcggc ttcccctgga gagagcgaga
     1681 ttctccgcgc tgtagaagtc accattgttg tgcacgacga catcattccg tggcgttatc
     1741 cagctaagcg cgaactgcaa tttggagaat ggcagcgcaa tgacattctt gcaggtatct
     1801 tcgagccagc cacgatcgac attgatctgg ctatcttgct gacaaaagca agagaacata
     1861 gcgttgcctt ggtaggtcca gcggcggagg aactctttga tccggttcct gaacaggatc
     1921 tatttgaggc gctaaatgaa accttaacgc tatggaactc gccgcccgac tgggctggcg
     1981 atgagcgaaa tgtagtgctt acgttgtccc gcatttggta cagcgcagta accggcaaaa
     2041 tcgcgccgaa ggatgtcgct gccgactggg caatggagcg cctgccggcc cagtatcagc
     2101 ccgtcatact tgaagctaga caggcttatc ttggacaaga agaagatcgc ttggcctcgc
     2161 gcgcagatca gttggaagaa tttgtccact acgtgaaagg cgagatcacc aaggtagtcg
     2221 gcaaataacc ctcgagccac ccatgaccaa aatcccttaa cgtgagttac gcgtcgttcc
     2281 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     2341 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     2401 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     2461 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     2521 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     2581 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     2641 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     2701 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     2761 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     2821 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     2881 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     2941 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     3001 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     3061 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     3121 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     3181 gtgagcgcaa cgcaattaat acgcgtaccg cgagccagga agagtttgta gaaacgcaaa
     3241 aaggccatcc gtcaggatgg ccttctgctt agtttgatgc ctggcagttt atggcgggcg
     3301 tcctgcccgc caccctccgg gccgttgctt cacaacgttc aaatccgctc ccggcggatt
     3361 tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttc
     3421 cgactgagcc tttcgtttta tttgatgcct ggcagttccc tactctcgcg ttaacgcttg
     3481 catggatgtt
//
