LOCUS       sequence-181865-        3471 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Module B, Promoter: CmYLCV, Gene: SapI ccdb cassette (doesn't 
            include the promoter) 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 3471)
  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 3471)
  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_B2103b
FEATURES             Location/Qualifiers
     source          1..3471
                     /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    1352..1371
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        1380..1455
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    1456..1475
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     CDS             1795..2100
                     /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     1979..1998
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    2128..2302
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     promoter        2379..2483
                     /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 ggcgcgcctg gcagacatac tgtcccacaa
      901 atgaagatgg aatctgtaaa agaaaacgcg tgaaataatg cgtctgacaa aggttaggtc
      961 ggctgccttt aatcaatacc aaagtggtcc ctaccacgat ggaaaaactg tgcagtcggt
     1021 ttggcttttt ctgacgaaca aataagattc gtggccgaca ggtgggggtc caccatgtga
     1081 aggcatcttc agactccaat aatggagcaa tgacgtaagg gcttacgaaa taagtaaggg
     1141 tagtttggga aatgtccact cacccgtcag tctataaata cttagcccct ccctcattgt
     1201 taagggagca aaatctcaga gagatagtcc tagagagaga aagagagcaa gtagcctaga
     1261 agtagtcaag gcggcgaagt attcaggcac gtggccagga agaagaaaag ccaagacgac
     1321 gaaaacaggt aagagctaag cttcctgcag ggttcactgc cgtataggca gcgaagagcg
     1381 ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg
     1441 gcaccgagtc ggtgcgttca ctgccgtata ggcagaagcc agataacagt atgcgtattt
     1501 gcgcgctgat ttttgcggta taagaatata tactgatatg tatacccgaa gtatgtcaaa
     1561 aggaggtatg ctatgaagca gcgtattaca gtgacagttg acagcgacag ctatcagttg
     1621 ctcaaggcat atatgatgtc aatatctccg gtctggtaag cacaaccatg cagaatgaag
     1681 cccgtcgtct gcgtgccgaa cgctggaaag cggaaaatca ggaagggatg gctgaggtcg
     1741 cccggtttat tgaaatgaac ggctcttttg ctgacgagaa caggggctgg tgaaatgcag
     1801 tttaaggttt acacctataa aagagagagc cgttatcgtc tgtttgtgga tgtacagagt
     1861 gatattattg acacgcccgg gcgacggatg gtgatccccc tggccagtgc acgtctgctg
     1921 tcagataaag tctcccgtga actttacccg gtggtgcata tcggggatga aagctggcgc
     1981 atgatgacca ccgatatggc cagtgtgccg gtttccgtta tcggggaaga agtggctgat
     2041 ctcagccacc gcgaaaatga catcaaaaac gccattaacc tgatgttctg gggaatataa
     2101 gctcttcagt cgatcgacaa gctcgagttt ctccataata atgtgtgagt agttcccaga
     2161 taagggaatt agggttccta tagggtttcg ctcatgtgtt gagcatataa gaaaccctta
     2221 gtatgtattt gtatttgtaa aatacttcta tcaataaaat ttctaattcc taaaaccaaa
     2281 atccagtact aaaatccaga tcccccgaat tagagctcta ccggtgacgc aggtgatgaa
     2341 aagccttagg gtcaggtggc acttttcggg gaaatgtgcg atgaacccct atttgtttat
     2401 ttttctaaat acattcaaat atgtatccgc taatgagaca ataaccctga taaatgcttc
     2461 aattatattg aaaaaggaag agtatgagta tccaacattt ccgtgtcgcc cttattccct
     2521 tttttgcggc cttctgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     2581 atgctgaaga tcagttgggt gcccgagtgg gttacattga actggatctc aacagcggca
     2641 aaatcttaga gagttttcgc cccgaagaac gcttcccaat gatgagcact ttcaaagttc
     2701 tgctatgtgg cgcggtgtta tcccgtattg atgctgggca agagcaactc ggtcgccgca
     2761 tacactattc tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg
     2821 atggcatgac agtaagagaa ttatgtagtg ctgccataac catgagtgat aacactgcgg
     2881 ccaacttact tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca
     2941 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     3001 acgacgagcg tgacaccacg atgcctgtag caatggcaac aactctccgc aaactattaa
     3061 ctggcgaact acttactctt gcttcacgcc agcaactcat tgactggatg gaggcggaca
     3121 aagttgcagg accacttctg cgctcggcac ttccggctgg ctggtttatt gctgataaat
     3181 ctggagccgg tgagcgtggc tctcgcggta tcattgcagc actggggcca gatggtaagc
     3241 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     3301 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gacgaaggga
     3361 aataaatagt agcccgcctg atgtgcgggc ttttttttgg tggtggttac cagtgtcgct
     3421 tagctaccgt aggtcctctt aggcggtccg attagcgaca tagtcatcaa t
//
