LOCUS       sequence-181873-        3656 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Module B, Promoter: 35S, Gene: SapI ccdb cassette for cloning 
            multiple gRNA spacers with tRNA spacers , Terminator: 35S.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3656)
  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 3656)
  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_B2301
FEATURES             Location/Qualifiers
     source          1..3656
                     /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"
     promoter        1068..1413
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1356..1376
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     misc_RNA        1508..1583
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             1980..2285
                     /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     2164..2183
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    2313..2487
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     promoter        2564..2668
                     /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 agagcatgga gtcaaagatt
      901 caaatagagg acctaacaga actcgccgta aagactggcg aacagttcat acagagtctc
      961 ttacgactca atgacaagaa gaaaatcttc gtcaacatgg tggagcacga cacacttgtc
     1021 tactccaaaa atatcaaaga tacagtctca gaagaccaaa gggcaattga gacttttcaa
     1081 caaagggtaa tatccggaaa cctcctcgga ttccattgcc cagctatctg tcactttatt
     1141 gtgaagatag tggaaaagga aggtggctcc tacaaatgcc atcattgcga taaaggaaag
     1201 gccatcgttg aagatgcctc tgccgacagt ggtcccaaag atggaccccc acccacgagg
     1261 agcatcgtgg aaaaagaaga cgttccaacc acgtcttcaa agcaagtgga ttgatgtgat
     1321 atctccactg acgtaaggga tgacgcacaa tcccactatc cttcgcaaga cccttcctct
     1381 atataaggaa gttcatttca tttggagaga acacggggga ctcctgcagg aacaaagcac
     1441 cagtggtcta gtggtagaat agtaccctgc cacggtacag acccgggttc gattcccggc
     1501 tggtgcagtt ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg
     1561 aaaaagtggc accgagtcgg tgcaacaaag caccagtggt ctagtggtag aatagtaccc
     1621 tgccacggta cagacccggg ttcgattccc ggctggtgca aagccagata acagtatgcg
     1681 tatttgcgcg ctgatttttg cggtataaga atatatactg atatgtatac ccgaagtatg
     1741 tcaaaaggag gtatgctatg aagcagcgta ttacagtgac agttgacagc gacagctatc
     1801 agttgctcaa ggcatatatg atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa
     1861 tgaagcccgt cgtctgcgtg ccgaacgctg gaaagcggaa aatcaggaag ggatggctga
     1921 ggtcgcccgg tttattgaaa tgaacggctc ttttgctgac gagaacaggg gctggtgaaa
     1981 tgcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     2041 agagtgatat tattgacacg cccgggcgac ggatggtgat ccccctggcc agtgcacgtc
     2101 tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     2161 ggcgcatgat gaccaccgat atggccagtg tgccggtttc cgttatcggg gaagaagtgg
     2221 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     2281 tataagctct tcagtcgatc gacaagctcg agtttctcca taataatgtg tgagtagttc
     2341 ccagataagg gaattagggt tcctataggg tttcgctcat gtgttgagca tataagaaac
     2401 ccttagtatg tatttgtatt tgtaaaatac ttctatcaat aaaatttcta attcctaaaa
     2461 ccaaaatcca gtactaaaat ccagatcccc cgaattagag ctctaccggt gacgcaggtg
     2521 atgaaaagcc ttagggtcag gtggcacttt tcggggaaat gtgcgatgaa cccctatttg
     2581 tttatttttc taaatacatt caaatatgta tccgctaatg agacaataac cctgataaat
     2641 gcttcaatta tattgaaaaa ggaagagtat gagtatccaa catttccgtg tcgcccttat
     2701 tccctttttt gcggccttct gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2761 aaaagatgct gaagatcagt tgggtgcccg agtgggttac attgaactgg atctcaacag
     2821 cggcaaaatc ttagagagtt ttcgccccga agaacgcttc ccaatgatga gcactttcaa
     2881 agttctgcta tgtggcgcgg tgttatcccg tattgatgct gggcaagagc aactcggtcg
     2941 ccgcatacac tattctcaga atgacttggt tgagtattca ccagtcacag aaaagcatct
     3001 tacggatggc atgacagtaa gagaattatg tagtgctgcc ataaccatga gtgataacac
     3061 tgcggccaac ttacttctga caacgattgg aggaccgaag gagctaaccg cttttttgca
     3121 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     3181 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaactc tccgcaaact
     3241 attaactggc gaactactta ctcttgcttc acgccagcaa ctcattgact ggatggaggc
     3301 ggacaaagtt gcaggaccac ttctgcgctc ggcacttccg gctggctggt ttattgctga
     3361 taaatctgga gccggtgagc gtggctctcg cggtatcatt gcagcactgg ggccagatgg
     3421 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     3481 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacga
     3541 agggaaataa atagtagccc gcctgatgtg cgggcttttt tttggtggtg gttaccagtg
     3601 tcgcttagct accgtaggtc ctcttaggcg gtccgattag cgacatagtc atcaat
//
