LOCUS       sequence-181862-        3922 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Module B, Promoter: GmUbi, Gene: SapI ccdb cassette for cloning 
            multiple gRNA spacers with Csy4 spacers , Terminator: 35S.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3922)
  AUTHORS   Cermak T, Curtin SJ, Gil-Humanes J, Cegan R, Kono TJY, Konecna E, 
            Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF
  TITLE     A multi-purpose toolkit to enable advanced genome engineering in 
            plants.
  JOURNAL   Plant Cell. 2017 May 18. pii: tpc.00922.2016. doi: 
            10.1105/tpc.16.00922.
  PUBMED    28522548
REFERENCE   2  (bases 1 to 3922)
  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.
            2017 May 18. pii: tpc.00922.2016. doi: 10.1105/tpc.16.00922."
COMMENT     Sequence Label: pMOD_B2121
FEATURES             Location/Qualifiers
     source          1..3922
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      80..668
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     569..588
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     822..839
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1811..1830
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        1831..1906
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    1907..1926
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     CDS             2246..2551
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2430..2449
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    2579..2753
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     promoter        2830..2934
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gccttgacgg caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
       61 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
      121 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ctaactcttt
      181 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
      241 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
      301 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
      361 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgctcacagc
      421 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
      481 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
      541 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
      601 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
      661 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
      721 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
      781 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
      841 aagcgggaga gcgcccacct gccaggggac ggcgcgccga agagcggggc ccaatataac
      901 aacgacgtcg taacagataa agcgaagctt gaaggtgcat gtgactccgt caagattacg
      961 aaaccgccaa ctaccacgca aattgcaatt ctcaatttcc tagaaggact ctccgaaaat
     1021 gcatccaata ccaaatatta cccgtgtcat aggcaccaag tgacaccata catgaacacg
     1081 cgtcacaata tgactggaga agggttccac accttatgct ataaaacgcc ccacacccct
     1141 cctccttcct tcgcagttca attccaatat attccattct ctctgtgtat ttccctacct
     1201 ctcccttcaa ggttagtcga tttcttctgt ttttcttctt cgttctttcc atgaattgtg
     1261 tatgttcttt gatcaatacg atgttgattt gattgtgttt tgtttggttt catcgatctt
     1321 caattttcat aatcagattc agcttttatt atctttacaa caacgtcctt aatttgatga
     1381 ttctttaatc gtagatttgc tctaattaga gctttttcat gtcagatccc tttacaacaa
     1441 gccttaattg ttgattcatt aatcgtagat tagggctttt ttcattgatt acttcagatc
     1501 cgttaaacgt aaccatagat cagggctttt tcatgaatta cttcagatcc gttaaacaac
     1561 agccttattt tttatacttc tgtggttttt caagaaattg ttcagatccg ttgacaaaaa
     1621 gccttattcg ttgattctat atcgtttttc gagagatatt gctcagatct gttagcaact
     1681 gccttgtttg ttgattctat tgccgtggat tagggttttt tttcacgaga ttgcttcaga
     1741 tccgtactta agattacgta atggattttg attctgattt atctgtgatt gttgactcga
     1801 caacctgcag gttcactgcc gtataggcag gttttagagc tagaaatagc aagttaaaat
     1861 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgcgttc actgccgtat
     1921 aggcagaagc cagataacag tatgcgtatt tgcgcgctga tttttgcggt ataagaatat
     1981 atactgatat gtatacccga agtatgtcaa aaggaggtat gctatgaagc agcgtattac
     2041 agtgacagtt gacagcgaca gctatcagtt gctcaaggca tatatgatgt caatatctcc
     2101 ggtctggtaa gcacaaccat gcagaatgaa gcccgtcgtc tgcgtgccga acgctggaaa
     2161 gcggaaaatc aggaagggat ggctgaggtc gcccggttta ttgaaatgaa cggctctttt
     2221 gctgacgaga acaggggctg gtgaaatgca gtttaaggtt tacacctata aaagagagag
     2281 ccgttatcgt ctgtttgtgg atgtacagag tgatattatt gacacgcccg ggcgacggat
     2341 ggtgatcccc ctggccagtg cacgtctgct gtcagataaa gtctcccgtg aactttaccc
     2401 ggtggtgcat atcggggatg aaagctggcg catgatgacc accgatatgg ccagtgtgcc
     2461 ggtttccgtt atcggggaag aagtggctga tctcagccac cgcgaaaatg acatcaaaaa
     2521 cgccattaac ctgatgttct ggggaatata agctcttcag tcgatcgaca agctcgagtt
     2581 tctccataat aatgtgtgag tagttcccag ataagggaat tagggttcct atagggtttc
     2641 gctcatgtgt tgagcatata agaaaccctt agtatgtatt tgtatttgta aaatacttct
     2701 atcaataaaa tttctaattc ctaaaaccaa aatccagtac taaaatccag atcccccgaa
     2761 ttagagctct accggtgacg caggtgatga aaagccttag ggtcaggtgg cacttttcgg
     2821 ggaaatgtgc gatgaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     2881 ctaatgagac aataaccctg ataaatgctt caattatatt gaaaaaggaa gagtatgagt
     2941 atccaacatt tccgtgtcgc ccttattccc ttttttgcgg ccttctgcct tcctgttttt
     3001 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcccgagtg
     3061 ggttacattg aactggatct caacagcggc aaaatcttag agagttttcg ccccgaagaa
     3121 cgcttcccaa tgatgagcac tttcaaagtt ctgctatgtg gcgcggtgtt atcccgtatt
     3181 gatgctgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     3241 tattcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgtagt
     3301 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gattggagga
     3361 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     3421 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     3481 gcaatggcaa caactctccg caaactatta actggcgaac tacttactct tgcttcacgc
     3541 cagcaactca ttgactggat ggaggcggac aaagttgcag gaccacttct gcgctcggca
     3601 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg ctctcgcggt
     3661 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     3721 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     3781 attaagcatt ggtaactgtc agacgaaggg aaataaatag tagcccgcct gatgtgcggg
     3841 cttttttttg gtggtggtta ccagtgtcgc ttagctaccg taggtcctct taggcggtcc
     3901 gattagcgac atagtcatca at
//
