LOCUS       sequence-181860-        4978 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Module B, Promoter: PvUbi1, 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 4978)
  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 4978)
  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_B2112
FEATURES             Location/Qualifiers
     source          1..4978
                     /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"
     CDS             complement(1589..1600)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    2867..2886
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        2887..2962
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    2963..2982
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     CDS             3302..3607
                     /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     3486..3505
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    3635..3809
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     promoter        3886..3990
                     /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 agagcacgtc agtgtttggt
      901 ttccactagc acgagtagcg caatcagaaa attttcaatg catgaagtac taaacgaagt
      961 ttatttagaa atttttttaa gaaatgagtg taattttttg cgacgaattt aatgacaata
     1021 attaatcgat gattgcctac agtaatgcta cagtaaccaa cctctaatca tgcgtcgaat
     1081 gcgtcattag attcgtctcg caaaatagca caagaattat gaaattaatt ttacaaacta
     1141 tttttattta atactaataa ttaactgtca aagtttgtgc tactcgcaag agtagcgcga
     1201 accaaacacg gcctggagga gcacggtaac ggcgtcgaca aactaacggc caccacccgc
     1261 caacgcaaag gagacggatg agagttgact tcttgacggt tctccacccc tctgtctctc
     1321 tgtcactggg ccctgggtcc ccctctcgaa agttcctctg gccgaaattg cgcggcggag
     1381 acgaggcggg cggaaccgtc acggcagagg attccttccc caccctgcct ggcccggcca
     1441 tatataaaca gccaccgccc ctccccgttc cccatcgcgt ctcgtctcgt gttgttccca
     1501 gaacacaacc aaaatccaaa tcctcctcct cctcccgagc ctcgtcgatc cctcacccgc
     1561 ttcaaggtac ggcgatcctc ctctcccttc tcccctcgat cgattatgcg tgttccgttt
     1621 ccgtttccga tcgagcgaat cgatggttag gacccatggg ggacccatgg ggtgtcgtgt
     1681 ggtggtctgg tttgatccgc gatatttctc cgttcgtagt gtagatctga tcgaatccct
     1741 ggtgaaatcg ttgatcgtgc tattcgtgtg agggttctta ggtttggagt tgtggaggta
     1801 gttctgatcg gtttgtaggt gagattttcc ccatgatttt gcttggctcg tttgtcttgg
     1861 ttagattaga tctgcccgca ttttgttcga tatttctgat gcagatatga tgaataattt
     1921 cgtccttgta tcccgcgtcc gtatgtgtat taagtttgca ggtcctagtt aggtttttcc
     1981 tactgatttg tcttatccat tctgtttagc ttgcaaggtt tggtaatggt ccggcatgtt
     2041 tgtctctata gattagagta gaataagatt atctcaacaa gctgttggct tatcaatttt
     2101 ggatctgcat gtgtttcgca tctatatctt tgcaattaag atggtagatg gacatatgct
     2161 cctgttgagt tgatgttgta ccttttacct gaggtctgag gaacatgcat cctcctgcta
     2221 ctttgtgctt atacagatca tcaagattat gtagctaata ttcgatcagt ttctagtatc
     2281 tacatggtaa acttgcatgc acttgctact tatttttgat atacttggat gataacatat
     2341 gctgctggtt gattcctacc tacatgatga acattttaca ggccattagt gtctgtctgt
     2401 atgtgttgtt cctgtttgct tcagtctatt tctgtttcat tcctagttta ttggttctct
     2461 gctagatact taccctgctg ggcttagtta tcatcttatc tcgaatgcat tttcatgttt
     2521 atagatgaat atacactcag ataggtgtag atgtatgcta ctgtttctct acgttgctgt
     2581 aggttttacc tgtggcaact gcatactcct gttgcttcgc tagatatgta tgtgcttata
     2641 tagattaaga tatgtgtgat ggttctttag tatatctgat gatcatgtat gctcttttaa
     2701 cttcttgcta cacttggtaa catgctgtga tgctgtttgt tgattctgta gcactaccaa
     2761 tgatgacctt atctctcttt gtatatgatg tttctgtttg tttgaggctt gtgttactgc
     2821 tagttactta ccctgttgcc tggctaatct tctgcagatc ctgcaggttc actgccgtat
     2881 aggcaggttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
     2941 aaaagtggca ccgagtcggt gcgttcactg ccgtataggc agaagccaga taacagtatg
     3001 cgtatttgcg cgctgatttt tgcggtataa gaatatatac tgatatgtat acccgaagta
     3061 tgtcaaaagg aggtatgcta tgaagcagcg tattacagtg acagttgaca gcgacagcta
     3121 tcagttgctc aaggcatata tgatgtcaat atctccggtc tggtaagcac aaccatgcag
     3181 aatgaagccc gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga agggatggct
     3241 gaggtcgccc ggtttattga aatgaacggc tcttttgctg acgagaacag gggctggtga
     3301 aatgcagttt aaggtttaca cctataaaag agagagccgt tatcgtctgt ttgtggatgt
     3361 acagagtgat attattgaca cgcccgggcg acggatggtg atccccctgg ccagtgcacg
     3421 tctgctgtca gataaagtct cccgtgaact ttacccggtg gtgcatatcg gggatgaaag
     3481 ctggcgcatg atgaccaccg atatggccag tgtgccggtt tccgttatcg gggaagaagt
     3541 ggctgatctc agccaccgcg aaaatgacat caaaaacgcc attaacctga tgttctgggg
     3601 aatataagct cttcagtcga tcgacaagct cgagtttctc cataataatg tgtgagtagt
     3661 tcccagataa gggaattagg gttcctatag ggtttcgctc atgtgttgag catataagaa
     3721 acccttagta tgtatttgta tttgtaaaat acttctatca ataaaatttc taattcctaa
     3781 aaccaaaatc cagtactaaa atccagatcc cccgaattag agctctaccg gtgacgcagg
     3841 tgatgaaaag ccttagggtc aggtggcact tttcggggaa atgtgcgatg aacccctatt
     3901 tgtttatttt tctaaataca ttcaaatatg tatccgctaa tgagacaata accctgataa
     3961 atgcttcaat tatattgaaa aaggaagagt atgagtatcc aacatttccg tgtcgccctt
     4021 attccctttt ttgcggcctt ctgccttcct gtttttgctc acccagaaac gctggtgaaa
     4081 gtaaaagatg ctgaagatca gttgggtgcc cgagtgggtt acattgaact ggatctcaac
     4141 agcggcaaaa tcttagagag ttttcgcccc gaagaacgct tcccaatgat gagcactttc
     4201 aaagttctgc tatgtggcgc ggtgttatcc cgtattgatg ctgggcaaga gcaactcggt
     4261 cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac agaaaagcat
     4321 cttacggatg gcatgacagt aagagaatta tgtagtgctg ccataaccat gagtgataac
     4381 actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac cgcttttttg
     4441 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     4501 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac tctccgcaaa
     4561 ctattaactg gcgaactact tactcttgct tcacgccagc aactcattga ctggatggag
     4621 gcggacaaag ttgcaggacc acttctgcgc tcggcacttc cggctggctg gtttattgct
     4681 gataaatctg gagccggtga gcgtggctct cgcggtatca ttgcagcact ggggccagat
     4741 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     4801 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     4861 gaagggaaat aaatagtagc ccgcctgatg tgcgggcttt tttttggtgg tggttaccag
     4921 tgtcgcttag ctaccgtagg tcctcttagg cggtccgatt agcgacatag tcatcaat
//
