LOCUS       sequence-474024-        9340 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  T-DNA vector for A3A/Y130F-CBE_V01 based C-to-T base editing for 
            monocot plants; NG or NA PAM ; wide working window; 
            A3A/Y130F-CBE_V01 was driven by ZmUbi1 and the sgRNA was driven by 
            OsU3 promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9340)
  AUTHORS   Cheng Y, Li G, Qi A, Mandlik R, Pan C, Wang D, Ge S, Qi Y
  TITLE     A comprehensive all-in-one CRISPR toolbox for large-scale screens in
            plants.
  JOURNAL   Plant Cell. 2025 Apr 9:koaf081. doi: 10.1093/plcell/koaf081.
  PUBMED    40202185
REFERENCE   2  (bases 1 to 9340)
  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 Cell.
            2025 Apr 9:koaf081. doi: 10.1093/plcell/koaf081."
COMMENT     Sequence Label: pYPQAT22
FEATURES             Location/Qualifiers
     source          1..9340
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             946..1575
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             2004..3077
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      3143..3337
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(3573..3595)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3681..3821
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3736..3755
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3836..3853)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4007..4595)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4087..4106)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4682..5476)
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     promoter        complement(6736..8732)
                     /label=Ubi promoter
                     /note="maize polyubiquitin gene promoter"
     primer_bind     complement(8748..8765)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8748..8764)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8757..8779)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    8967..8991
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
ORIGIN
        1 atgcccgcgt cagcaccgac gaccaggact tgaccaacca acgggccgaa ctgcacgcgg
       61 ccggctgcac caagctgttt tccgagaaga tcaccggcac caggcgcgac cgcccggagc
      121 tggccaggat gcttgaccac ctacgccctg gcgacgttgt gacagtgacc aggctagacc
      181 gcctggcccg cagcacccgc gacctactgg acattgccga gcgcatccag gaggccggcg
      241 cgggcctgcg tagcctggca gagccgtggg ccgacaccac cacgccggcc ggccgcatgg
      301 tgttgaccgt gttcgccggc attgccgagt tcgagcgttc cctaatcatc gaccgcaccc
      361 ggagcgggcg cgaggccgcc aaggcccgag gcgtgaagtt tggcccccgc cctaccctca
      421 ccccggcaca gatcgcgcac gcccgcgagc tgatcgacca ggaaggccgc accgtgaaag
      481 aggcggctgc actgcttggc gtgcatcgct cgaccctgta ccgcgcactt gagcgcagcg
      541 aggaagtgac gcccaccgag gccaggcggc gcggtgccta ggacgcattg accgaggccg
      601 acgccctggc ggccgccgag aatgaacgcc aagaggaaca agcatgaaac cgcaccagga
      661 cggccaggac gaaccgtttt tcattaccga agagatcgag gcggagatga tcgcggccgg
      721 gtacgtgttc gagccgcccg cgcacgtctc aaccgtgcgg ctgcatgaaa tcctggccgg
      781 tttgtctgat gccaagctgg cggcctggcc ggccagcttg gccgctgaag aaaccgagcg
      841 ccgccgtcta aaaaggtgat gtgtatttga gtaaaacagc ttgcgtcatg cggtcgctgc
      901 gtatatgatg cgatgagtaa ataaacaaat acgcaagggg aacgcatgaa ggttatcgct
      961 gtacttaacc agaaaggcgg gtcaggcaag acgaccatcg caacccatct agcccgcgcc
     1021 ctgcaactcg ccggggccga tgttctgtta gtcgattccg atccccaggg cagtgcccgc
     1081 gattgggcgg ccgtgcggga agatcaaccg ctaaccgttg tcggcatcga ccgcccgacg
     1141 attgaccgcg acgtgaaggc catcggccgg cgcgacttcg tagtgatcga cggagcgccc
     1201 caggcggcgg acttggctgt gtccgcgatc aaggcagccg acttcgtgct gattccggtg
     1261 cagccaagcc cttacgacat atgggccacc gccgacctgg tggagctggt taagcagcgc
     1321 attgaggtca cggatggaag gctacaagcg gcctttgtcg tgtcgcgggc gatcaaaggc
     1381 acgcgcatcg gcggtgaggt tgccgaggcg ctggccgggt acgagctgcc cattcttgag
     1441 tcccgtatca cgcagcgcgt gagctaccca ggcactgccg ccgccggcac aaccgttctt
     1501 gaatcagaac ccgagggcga cgctgcccgc gaggtccagg cgctggccgc tgaaattaaa
     1561 tcaaaactca tttgagttaa tgaggtaaag agaaaatgag caaaagcaca aacacgctaa
     1621 gtgccggccg tccgagcgca cgcagcagca aggctgcaac gttggccagc ctggcagaca
     1681 cgccagccat gaagcgggtc aactttcagt tgccggcgga ggatcacacc aagctgaaga
     1741 tgtacgcggt acgccaaggc aagaccatta ccgagctgct atctgaatac atcgcgcagc
     1801 taccagagta aatgagcaaa tgaataaatg agtagatgaa ttttagcggc taaaggaggc
     1861 ggcatggaaa atcaagaaca accaggcacc gacgccgtgg aatgccccat gtgtggagga
     1921 acgggcggtt ggccaggcgt aagcggctgg gttgcctgcc ggccctgcaa tggcactgga
     1981 acccccaagc ccgaggaatc ggcgtgagcg gtcgcaaacc atccggcccg gtacaaatcg
     2041 gcgcggcgct gggtgatgac ctggtggaga agttgaaggc cgcgcaggcc gcccagcggc
     2101 aacgcatcga ggcagaagca cgccccggtg aatcgtggca agcggccgct gatcgaatcc
     2161 gcaaagaatc ccggcaaccg ccggcagccg gtgcgccgtc gattaggaag ccgcccaagg
     2221 gcgacgagca accagatttt ttcgttccga tgctctatga cgtgggcacc cgcgatagtc
     2281 gcagcatcat ggacgtggcc gttttccgtc tgtcgaagcg tgaccgacga gctggcgagg
     2341 tgatccgcta cgagcttcca gacgggcacg tagaggtttc cgcagggccg gccggcatgg
     2401 ccagtgtgtg ggattacgac ctggtactga tggcggtttc ccatctaacc gaatccatga
     2461 accgataccg ggaagggaag ggagacaagc ccggccgcgt gttccgtcca cacgttgcgg
     2521 acgtactcaa gttctgccgg cgagccgatg gcggaaagca gaaagacgac ctggtagaaa
     2581 cctgcattcg gttaaacacc acgcacgttg ccatgcagcg tacgaagaag gccaagaacg
     2641 gccgcctggt gacggtatcc gagggtgaag ccttgattag ccgctacaag atcgtaaaga
     2701 gcgaaaccgg gcggccggag tacatcgaga tcgagctagc tgattggatg taccgcgaga
     2761 tcacagaagg caagaacccg gacgtgctga cggttcaccc cgattacttt ttgatcgatc
     2821 ccggcatcgg ccgttttctc taccgcctgg cacgccgcgc cgcaggcaag gcagaagcca
     2881 gatggttgtt caagacgatc tacgaacgca gtggcagcgc cggagagttc aagaagttct
     2941 gtttcaccgt gcgcaagctg atcgggtcaa atgacctgcc ggagtacgat ttgaaggagg
     3001 aggcggggca ggctggcccg atcctagtca tgcgctaccg caacctgatc gagggcgaag
     3061 catccgccgg ttcctaatgt acggagcaga tgctagggca aattgcccta gcaggggaaa
     3121 aaggtcgaaa aggtctgttt cctgtggata gcacgtacat tgggaaccca aagccgtaca
     3181 ttgggaaccg gaacccgtac attgggaacc caaagccgta cattgggaac cggtcacaca
     3241 tgtaagtgac tgatataaaa gagaaaaaag gcgatttttc cgcctaaaac tctttaaaac
     3301 ttattaaaac tcttaaaacc cgcctggcct gtgcataact gtctggccag cgcacagccg
     3361 aagagctgca aaaagcgcct acccttcggt cgctgcgctc cctacgcccc gccgcttcgc
     3421 gtcggcctat cgcggccgct ggccgctcaa aaatggctgg cctacggcca ggcaatctac
     3481 cagggcgcgg acaagccgcg ccgtcgccac tcgaccgccg gcgcccacat caaggcaccc
     3541 tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     3601 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     3661 ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat
     3721 actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg
     3781 aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc gcttcctcgc
     3841 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     3901 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     3961 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4021 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4081 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4141 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4201 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4261 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4321 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4381 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4441 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4501 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4561 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4621 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg cattctaggt
     4681 actaaaacaa ttcatccagt aaaatataat attttatttt ctcccaatca ggcttgatcc
     4741 ccagtaagtc aaaaaatagc tcgacatact gttcttcccc gatatcctcc ctgatcgacc
     4801 ggacgcagaa ggcaatgtca taccacttgt ccgccctgcc gcttctccca agatcaataa
     4861 agccacttac tttgccatct ttcacaaaga tgttgctgtc tcccaggtcg ccgtgggaaa
     4921 agacaagttc ctcttcgggc ttttccgtct ttaaaaaatc atacagctcg cgcggatctt
     4981 taaatggagt gtcttcttcc cagttttcgc aatccacatc ggccagatcg ttattcagta
     5041 agtaatccaa ttcggctaag cggctgtcta agctattcgt atagggacaa tccgatatgt
     5101 cgatggagtg aaagagcctg atgcactccg catacagctc gataatcttt tcagggcttt
     5161 gttcatcttc atactcttcc gagcaaagga cgccatcggc ctcactcatg agcagattgc
     5221 tccagccatc atgccgttca aagtgcagga cctttggaac aggcagcttt ccttccagcc
     5281 atagcatcat gtccttttcc cgttccacat cataggtggt ccctttatac cggctgtccg
     5341 tcatttttaa atataggttt tcattttctc ccaccagctt atatacctta gcaggagaca
     5401 ttccttccgt atcttttacg cagcggtatt tttcgatcag ttttttcaat tccggtgata
     5461 ttctcatttt agccatttat tatttccttc ctcttttcta cagtatttaa agatacccca
     5521 agaagctaag gaaggtgcga ataagcgggg aaattcttct cggctgactc agtcatttca
     5581 tttcttcatg tttgagccga ttttttctcc cgtaaatgcc ttgaatcagc ctatttagac
     5641 cgtttcttcg ccatttaagg cgttatcccc agtttttagt gagatctctc ccactgacgt
     5701 atcatttggt ccgcccgaaa caggttggcc agcgtgaata acatcgccag ttggttatcg
     5761 tttttcagca accccttgta tctggctttc acgaagccga actgtcgctt gatgatgcga
     5821 aatgggtgct ccaccctggc ccggatgctg gctttcatgt attcgatgtt gatggccgtt
     5881 ttgttcttgc gtggatgctg tttcaaggtt cttaccttgc cggggcgctc ggcgatcagc
     5941 cagtccacat ccacctcggc cagctcctcg cgctgtggcg ccccttggta gccggcatcg
     6001 gctgagacaa attgctcctc tccatgcagc agattaccca gctgattgag gtcatgctcg
     6061 ttggccgcgg tggtgaccag gctgtgggtc aggccactct tggcatcgac accaatgtgg
     6121 gccttcatgc caaagtgcca ctgattgcct ttcttggtct gatgcatctc cggatcgcgt
     6181 tgctgctctt tgttcttggt cgagctgggt gcctcaatga tggtggcatc gaccaaggtg
     6241 ccttgagtca tcatgacgcc tgcttcggcc agccagcgat tgatggtctt gaacaattgg
     6301 cgggccagtt gatgctgctc cagcaggtgg cggaaattca tgatggtggt gcggtccggc
     6361 aaggcgctat ccagggataa ccgggcaaac agacgcatgg aggcgatttc gtacagagca
     6421 tcttccatcg cgccatcgct caggttgtac caatgctgca tgcagtgaat gcgtagcatg
     6481 gtttccagcg gataaggtcg ccggccatta ccagccttgg ggtaaaacgg ctcgatgact
     6541 tccaccatgt tttgccatgg cagaatctgc tccatgcggg acaagaaaat ctcttttctg
     6601 gtctgacggc gcttactgct gaattcactg tcggcgaagg taagttgatg actcatgatg
     6661 aaccctgttc tatggctcca gatgacaaac atgatctcat atcagggact tgttcgcacc
     6721 ttccctagag tcgacctgca gaagtaacac caaacaacag ggtgagcatc gacaaaagaa
     6781 acagtaccaa gcaaataaat agcgtatgaa ggcagggcta aaaaaatcca catatagctg
     6841 ctgcatatgc catcatccaa gtatatcaag atcaaaataa ttataaaaca tacttgttta
     6901 ttataataga taggtactca aggttagagc atatgaatag atgctgcata tgccatcatg
     6961 tatatgcatc agtaaaaccc acatcaacat gtatacctat cctagatcga tatttccatc
     7021 catcttaaac tcgtaactat gaagatgtat gacacacaca tacagttcca aaattaataa
     7081 atacaccagg tagtttgaaa cagtattcta ctccgatcta gaacgaatga acgaccgccc
     7141 aaccacacca catcatcaca accaagcgaa caaaaagcat ctctgtatat gcatcagtaa
     7201 aacccgcatc aacatgtata cctatcctag atcgatattt ccatccatca tcttcaattc
     7261 gtaactatga atatgtatgg cacacacata cagatccaaa attaataaat ccaccaggta
     7321 gtttgaaaca gaattaattc tactccgatc tagaacgacc gcccaaccag accacatcat
     7381 cacaaccaag acaaaaaaaa gcatgaaaag atgacccgac aaacaagtgc acggcatata
     7441 ttgaaataaa ggaaaagggc aaaccaaacc ctatgcaacg aaacaaaaaa aatcatgaaa
     7501 tcgatcccgt ctgcggaacg gctagagcca tcccaggatt ccccaaagag aaacactggc
     7561 aagttagcaa tcagaacgtg tctgacgtac aggtcgcatc cgtgtacgaa cgctagcagc
     7621 acggatctaa cacaaacacg gatctaacac aaacatgaac agaagtagaa ctaccgggcc
     7681 ctaaccatgg accggaacgc cgatctagag aaggtagaga gggggggggg gggaggacga
     7741 gcggcgtacc ttgaagcgga ggtgccgacg ggtggatttg ggggagatct ggttgtgtgt
     7801 gtgtgcgctc cgaacaacac gaggttgggg aaagagggtg tggagggggt gtctatttat
     7861 tacggcgggc gaggaaggga aagcgaagga gcggtgggaa aggaatcccc cgtagctgcc
     7921 ggtgccgtga gaggaggagg aggccgcctg ccgtgccggc tcacgtctgc cgctccgcca
     7981 cgcaatttct ggatgccgac agcggagcaa gtccaacggt ggagcggaac tctcgagagg
     8041 ggtccagagg cagcgacaga gatgccgtgc cgtctgcttc gcttggcccg acgcgacgct
     8101 gctggttcgc tggttggtgt ccgttagact cgtcgacggc gtttaacagg ctggcattat
     8161 ctactcgaaa caagaaaaat gtttccttag tttttttaat ttcttaaagg gtatttgttt
     8221 aatttttagt cactttattt tattctattt tatatctaaa ttattaaata aaaaaactaa
     8281 aatagagttt tagttttctt aatttagagg ctaaaataga ataaaataga tgtactaaaa
     8341 aaattagtct ataaaaacca ttaaccctaa accctaaatg gatgtactaa taaaatggat
     8401 gaagtattat ataggtgaag ctatttgcaa aaaaaaagga gaacacatgc acactaaaaa
     8461 gataaaactg tagagtcctg ttgtcaaaat actcaattgt cctttagacc atgtctaact
     8521 gttcatttat atgattctct aaaacactga tattattgta gtactataga ttatattatt
     8581 cgtagagtaa agtttaaata tatgtataaa gatagataaa ctgcacttca aacaagtgtg
     8641 acaaaaaaaa tatgtggtaa ttttttataa cttagacatg caatgctcat tatctctaga
     8701 gaggggcacg accgggtcac gctgcactgc aggcatgcaa gcttggcact ggccgtcgtt
     8761 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     8821 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     8881 ttgcgcagcc tgaatggcga atgctagagc agcttgagct tggatcagat tgtcgtttcc
     8941 cgccttcagt ttaaactatc agtgtttgac aggatatatt ggcgggtaaa cctaagagaa
     9001 aagagcgttt attagaataa tcggatattt aaaagggcgt gaaaaggttt atccgttcgt
     9061 ccatttgtat gtgcatgcca accacagggt tcccctcggg atcaaagtac tttgatccaa
     9121 cccctccgct gctatagtgc agtcggcttc tgacgttcag tgcagccgtc ttctgaaaac
     9181 gacatgtcgc acaagtccta agttacgcga caggctgccg ccctgccctt ttcctggcgt
     9241 tttcttgtcg cgtgttttag tcgcataaag tagaatactt gcgactagaa ccggagacat
     9301 tacgccatga acaagagcgc cgccgctggc ctgctgggct
//
