LOCUS       sequence-307437-        3501 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intermediate sgRNA array cloning vector position 4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3501)
  AUTHORS   Stuttmann J, Barthel K, Martin P, Ordon J, Erickson JL, Herr R, 
            Ferik F, Kretschmer C, Berner T, Keilwagen J, Marillonnet S, Bonas U
  TITLE     Highly efficient multiplex editing: One-shot generation of 8x 
            Nicotiana benthamiana and 12x Arabidopsis mutants.
  JOURNAL   Plant J. 2021 Feb 12. doi: 10.1111/tpj.15197.
  PUBMED    33577114
REFERENCE   2  (bases 1 to 3501)
  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 J. 
            2021 Feb 12. doi: 10.1111/tpj.15197."
COMMENT     Sequence Label: pDGE506
FEATURES             Location/Qualifiers
     source          1..3501
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     199..216
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        310..412
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             413..1096
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     VDPAY"
     primer_bind     complement(479..498)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1416..1721
                     /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     1600..1619
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     complement(1802..1820)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1888..1992
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1993..2787
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSLREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      join(2958..3501,1..45)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3447..3466
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
       61 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
      121 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
      181 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
      241 aaaccgcctc tccccgcgcg ttggccgatt cattaatcac tctgtgaaga catgactttg
      301 ctgagacctt gatcggcacg taagaggttc caactttcac cataatgaaa taagatcact
      361 accgggcgta ttttttgagt tatcgagatt ttcaggagct aaggaagcta aaatggagaa
      421 aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga
      481 ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc
      541 ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct
      601 tgcccgcctg atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt
      661 gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc
      721 atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga
      781 tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt
      841 tttcgtttca gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat
      901 ggacaacttc ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt
      961 gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag
     1021 aatgcttaat gaattacaac agtactgcga tgagtggcag ggggggcgta aacgccgcgt
     1081 ggatccggct tactaaaagc cagataacag tatgcgtatt tgcgcgctga tttttgcggt
     1141 ataagaatat atactgatat gtatacccga agtatgtcaa aaagaggtat gctatgaagc
     1201 agcgtattac agtgacagtt gacagcgaca gctatcagtt gctcaaggca tatatgatgt
     1261 caatatctcc ggtctggtaa gcacaaccat gcagaatgaa gcccgtcgtc tgcgtgccga
     1321 acgctggaaa gcggaaaatc aggaagggat ggctgaggtc gcccggttta ttgaaatgaa
     1381 cggctctttt gctgacgaga acaggggctg gtgaaatgca gtttaaggtt tacacctata
     1441 aaagagagag ccgttatcgt ctgtttgtgg atgtacagag tgatattatt gacacgcctg
     1501 ggcgacggat ggtgatcccc ctggccagtg cacgtctgct gtcagataaa gtctcccgtg
     1561 aactttaccc ggtggtgcat atcggggatg aaagctggcg catgatgacc accgatatgg
     1621 ccagtgtgcc ggtttccgtt atcggggaag aagtggctga tctcagccac cgcgaaaatg
     1681 acatcaaaaa cgccattaac ctgatgttct ggggaatata aatgtcaggc tcccttatac
     1741 acagccagtc ggtctcagct tccagttgtc ttcacgaagt ggctcttcag tggacgaaag
     1801 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     1861 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     1921 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     1981 aaaaggaaga gtatggctaa aatgagaata tcaccggaat tgaaaaaact gatcgaaaaa
     2041 taccgctgcg taaaagatac ggaaggaatg tctcctgcta aggtatataa gctggtggga
     2101 gaaaatgaaa acctatattt aaaaatgacg gacagccggt ataaagggac cacctatgat
     2161 gtggaacggg aaaaggacat gatgctatgg ctggaaggaa agctgcctgt tccaaaggtc
     2221 ctgcactttg aacggcatga tggctggagc aatctgctca tgagtgaggc cgatggcgtc
     2281 ctttgctcgg aagagtatga agatgaacaa agccctgaaa agattatcga gctgtatgcg
     2341 gagtgcatca ggctctttca ctccatcgac atatcggatt gtccctatac gaatagctta
     2401 gacagccgct tagccgaatt ggattactta ctgaataacg atctggccga tgtggattgc
     2461 gaaaactggg aagaggacac tccatttaaa gatccgcgcg agctgtatga ttttttaaag
     2521 acggaaaagc ccgaagagga acttgtcttt tcccacggcg acctgggaga cagcaacatc
     2581 tttgtgaaag atggcaaagt aagtggcttt attgatcttg ggagaagcgg cagggcggac
     2641 aagtggtatg acattgcctt ctgcgtccgg tcgctcaggg aggatatcgg ggaagaacag
     2701 tatgtcgagc tattttttga cttactgggg atcaagcctg attgggagaa aataaaatat
     2761 tatattttac tggatgaatt gttttagctg tcagaccaag tttactcata tatactttag
     2821 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     2881 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     2941 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     3001 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     3061 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     3121 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     3181 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     3241 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     3301 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     3361 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     3421 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     3481 tttcgccacc tctgacttga g
//
