LOCUS       sequence-422161-        4717 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Modular Unit carrying pegRNA and ngRNA scaffolds flanked by attR4 
            and attR3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4717)
  AUTHORS   Gupta A, Liu B, Raza S, Chen QJ, Yang B
  TITLE     Modularly assembled multiplex prime editors for simultaneous editing
            of agronomically important genes in rice.
  JOURNAL   Plant Commun. 2023 Oct 26:100741. doi: 10.1016/j.xplc.2023.100741.
  PUBMED    37897041
REFERENCE   2  (bases 1 to 4717)
  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 
            Commun. 2023 Oct 26:100741. doi: 10.1016/j.xplc.2023.100741."
COMMENT     Sequence Label: pCR8-attR4-pegRNA-attR3
FEATURES             Location/Qualifiers
     source          1..4717
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      882..1470
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1371..1390
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1624..1641
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(1800..1827)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(1919..2005)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2054..2076
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2068..2085
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2069..2085
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2101..2225)
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     ncRNA           3472..3539
                     /product="hepatitis delta virus self-cleaving ribozyme 
                     (Ferre-D'Amare and Scott, 2010)"
                     /label=HDV ribozyme
                     /ncRNA_class="ribozyme"
     misc_RNA        3645..3720
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     ncRNA           3730..3797
                     /product="hepatitis delta virus self-cleaving ribozyme 
                     (Ferre-D'Amare and Scott, 2010)"
                     /label=HDV ribozyme
                     /ncRNA_class="ribozyme"
     terminator      3829..4077
                     /label=HSP terminator
                     /note="efficient transcription terminator from the 
                     Arabidopsis thaliana heat shock protein 18.2 gene (Nagaya 
                     et al., 2010)"
     protein_bind    4085..4208
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4244..4263)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4245..4263)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4268..4284)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4268..4284)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4715..4717,1..789)
                     /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"
ORIGIN
        1 agggaagcgg tgatcgccga agtatcgact caactatcag aggtagttgg cgtcatcgag
       61 cgccatctcg aaccgacgtt gctggccgta catttgtacg gctccgcagt ggatggcggc
      121 ctgaagccac acagtgatat tgatttgctg gttacggtga ccgtaaggct tgatgaaaca
      181 acgcggcgag ctttgatcaa cgaccttttg gaaacttcgg cttcccctgg agagagcgag
      241 attctccgcg ctgtagaagt caccattgtt gtgcacgacg acatcattcc gtggcgttat
      301 ccagctaagc gcgaactgca atttggagaa tggcagcgca atgacattct tgcaggtatc
      361 ttcgagccag ccacgatcga cattgatctg gctatcttgc tgacaaaagc aagagaacat
      421 agcgttgcct tggtaggtcc agcggcggag gaactctttg atccggttcc tgaacaggat
      481 ctatttgagg cgctaaatga aaccttaacg ctatggaact cgccgcccga ctgggctggc
      541 gatgagcgaa atgtagtgct tacgttgtcc cgcatttggt acagcgcagt aaccggcaaa
      601 atcgcgccga aggatgtcgc tgccgactgg gcaatggagc gcctgccggc ccagtatcag
      661 cccgtcatac ttgaagctag acaggcttat cttggacaag aagaagatcg cttggcctcg
      721 cgcgcagatc agttggaaga atttgtccac tacgtgaaag gcgagatcac caaggtagtc
      781 ggcaaataac cctcgagcca cccatgacca aaatccctta acgtgagtta cgcgtcgttc
      841 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
      901 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
      961 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     1021 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     1081 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     1141 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     1201 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     1261 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     1321 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     1381 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     1441 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     1501 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     1561 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     1621 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     1681 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     1741 agtgagcgca acgcaattaa tacgcgtacc gctagccagg aagagtttgt agaaacgcaa
     1801 aaaggccatc cgtcaggatg gccttctgct tagtttgatg cctggcagtt tatggcgggc
     1861 gtcctgcccg ccaccctccg ggccgttgct tcacaacgtt caaatccgct cccggcggat
     1921 ttgtcctact caggagagcg ttcaccgaca aacaacagat aaaacgaaag gcccagtctt
     1981 ccgactgagc ctttcgtttt atttgatgcc tggcagttcc ctactctcgc gttaacgcta
     2041 gcatggatgt tttcccagtc acgacgttgt aaaacgacgg ccagtcttaa gctcgggccc
     2101 catagtgact ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc
     2161 taatttaata tattgatatt tatatcattt tacgtttctc gttcaacttt tctatacaaa
     2221 gttggccgaa ttatggagtc aaagattcaa atagaggacc taacagaact cgccgtaaag
     2281 actggcgaac agttcataca gagtctctta cgactcaatg acaagaagaa aatcttcgtc
     2341 aacatggtgg agcacgacac acttgtctac tccaaaaata tcaaagatac agtctcagaa
     2401 gaccaaaggg caattgagac ttttcaacaa agggtaatat ccggaaacct cctcggattc
     2461 cattgcccag ctatctgtca ctttattgtg aagatagtgg aaaaggaagg tggctcctac
     2521 aaatgccatc attgcgataa aggaaaggcc atcgttgaag atgcctctgc cgacagtggt
     2581 cccaaagatg gacccccacc cacgaggagc atcgtggaaa aagaagacgt tccaaccacg
     2641 tcttcaaagc aagtggattg atgtgattgg cagacatact gtcccacaaa tgaagatgga
     2701 atctgtaaaa gaaaacgcgt gaaataatgc gtctgacaaa ggttaggtcg gctgccttta
     2761 atcaatacca aagtggtccc taccacgatg gaaaaactgt gcagtcggtt tggctttttc
     2821 tgacgaacaa ataagattcg tggccgacag gtgggggtcc accatgtgaa ggcatcttca
     2881 gactccaata atggagcaat gacgtaaggg cttacgaaat aagtaagggt agtttgggaa
     2941 atgtccactc acccgtcagt ctataaatac ttagcccctc cctcattgtt aagggagcaa
     3001 aatctcagag agatagtcct agagagagaa agagagcaag tagcctagaa gtagtcaagg
     3061 cggcgaagta ttcaggcacg tggccaggaa gaagaaaagc caagacgacg aaaacaggta
     3121 agagctaagc atctagaaag ttgaaaacaa tcttcaaaag tcccacatcg cttagataag
     3181 aaaacgaagc tgagtttata tacagctaga gtcgaagtag tgattgaaca aagcaccagt
     3241 ggtctagtgg tagaatagta ccctgccacg gtacagaccc gggttcgatt cccggctggt
     3301 gcaagagacg agtcacgtct cagtttaaga gctatgctgg aaacagcata gcaagtttaa
     3361 ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgcag agaccacagg
     3421 atccttggtc tcacgcggtt ctatctagtt acgcgttaaa ccaactagaa aggccggcat
     3481 ggtcccagcc tcctcgctgg cgccggctgg gcaacatgct tcggcatggc gaatgggaca
     3541 acaacaaatc agagtggcgc agcggaagcg tggtgggccc ataacccaca ggtcccagga
     3601 tcgaaacctg gctctgataa gagaccacag gatccttggt ctcagtttta gagctagaaa
     3661 tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc gagtcggtgc
     3721 tttttttttg gccggcatgg tcccagcctc ctcgctggcg ccggctgggc aacatgcttc
     3781 ggcatggcga atgggacttt ttttttgata tctccggggc taattgaata tgaagatgaa
     3841 gatgaaatat ttggtgtgtc aaataaaaag ctggtgtgct taagtttgtg tttttttctt
     3901 ggcttgttgt gttatgaatt tgtggctttt tctaatatta aatgaatgta agatctcatt
     3961 ataatgaata aacaaatgtt tctataatcc attgtgaatg ttttgttgga tctcttctgc
     4021 agcatataac tactgtatgt gctatggtat ggactatgga atatgattaa agataaggaa
     4081 ttcgcaactt tgtataataa agttgaacga gaaacgtaaa atgatataaa tatcaatata
     4141 ttaaattaga ttttgcataa aaaacagact acataatact gtaaaacaca acatatccag
     4201 tcactatgaa aataaaatca ttatttgcca tccagctgat atcccctata gtgagtcgta
     4261 ttacatggtc atagctgttt cctggcagct ctggcccgtg tctcaaaatc tctgatgtta
     4321 cattgcacaa gataaaaata tatcatcatg cctcctctag accagccagg acagaaatgc
     4381 ctcgacttcg ctgctaccca aggttgccgg gtgacgcaca ccgtggaaac ggatgaaggc
     4441 acgaacccag tggacataag cctgttcggt tcgtaagctg taatgcaagt agcgtatgcg
     4501 ctcacgcaac tggtccagaa ccttgaccga acgcagcggt ggtaacggcg cagtggcggt
     4561 tttcatggct tgttatgact gtttttttgg ggtacagtct atgcctcggg catccaagca
     4621 gcaagcgcgt tacgccgtgg gtcgatgttt gatgttatgg agcagcaacg atgttacgca
     4681 gcagggcagt cgccctaaaa caaagttaaa cattatg
//
