LOCUS       sequence-485613-        5430 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A suicide vector for Agrobacterium with kanamycin resistance..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5430)
  AUTHORS   Liu MS, Huang TK, Wang YC, Wang SC, Wu CH, Kuo CH, Lai EM
  TITLE     Floral stage optimization and immune evasion enhance 
            Agrobacterium-mediated genome editing in Arabidopsis.
  JOURNAL   New Phytol. 2025 Nov 30. doi: 10.1111/nph.70795.
  PUBMED    41321035
REFERENCE   2  (bases 1 to 5430)
  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: "New Phytol.
            2025 Nov 30. doi: 10.1111/nph.70795."
COMMENT     Sequence Label: EML8543_pEML41003
FEATURES             Location/Qualifiers
     source          1..5430
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    79..100
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(115..120,121..138,139..145)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    153..169
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     158..180
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     177..193
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     177..193
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             189..512
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    206..262
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(263..280)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(263..279)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(272..294)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(488..507)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     terminator      617..711
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(810..829)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             821..1636
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1816..1833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1950..2495)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(2030..2049)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     oriT            2869..2978
                     /note="incP origin of transfer"
     promoter        3358..3803
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(3804..3890,3891..5225)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     terminator      complement(5335..5421)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gaattcggat ccggagtgag accgcagctg gcacgacagg tttgccgact ggaaagcggg
       61 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
      121 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
      181 aaacagctat gaccatgatt acgccaagct tgcatgcctg caggtcgact ctagaggatc
      241 cccgggtacc gagctcgaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
      301 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
      361 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc
      421 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca
      481 ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac
      541 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
      601 cggtctcacg ctcttggact cctgttgata gatccagtaa tgacctcaga actccatctg
      661 gatttgttca gaacgctcgg ttgccgccgg gcgtttttta ttggtgagaa tccaggcaat
      721 tgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat catgaacaat
      781 aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata ttcagcgtga
      841 aacgagctgt agccgtccgc gtctgaacag caacatggat gcggatctgt atggctataa
      901 atgggcgcgt gataacgtgg gtcagagcgg cgcgaccatt tatcgtctgt atggcaaacc
      961 ggatgcgccg gaactgtttc tgaaacatgg caaaggcagc gtggcgaacg atgtgaccga
     1021 tgaaatggtg cgtctgaact ggctgaccga atttatgccg ctgccgacca ttaaacattt
     1081 tattcgcacc ccggatgatg cgtggctgct gaccaccgcg attccgggca aaaccgcgtt
     1141 tcaggtgctg gaagaatatc cggatagcgg cgaaaacatt gtggatgcgc tggccgtgtt
     1201 tctgcgtcgt ctgcatagca ttccggtgtg caactgcccg tttaacagcg atcgtgtgtt
     1261 tcgtctggcc caggcgcaga gccgtatgaa caacggcctg gtggatgcga gcgattttga
     1321 tgatgaacgt aacggctggc cggtggaaca ggtgtggaaa gaaatgcata aactgctgcc
     1381 gtttagcccg gatagcgtgg tgacccacgg cgattttagc ctggataacc tgattttcga
     1441 tgaaggcaaa ctgattggct gcattgatgt gggccgtgtg ggcattgcgg atcgttatca
     1501 ggatctggcc attctgtgga actgcctggg cgaatttagc ccgagcctgc aaaaacgtct
     1561 gtttcagaaa tatggcattg ataatccgga tatgaacaaa ctgcaatttc atctgatgct
     1621 ggatgaattt ttctaataat taattgaact atgcgtaacg gcaaaagcac cgccggacat
     1681 cagcgctagc ggagtgtata ctggcttact atgttggcac tgatgagggt gtcagtgaag
     1741 tgcttcatgt ggcaggagaa aaaaggctgc accggtgcgt cagcagaata tgtgatacag
     1801 gatatattcc gcttcctcgc tcactgactc gctacgctcg gtcgttcgac tgcggcgagc
     1861 ggaaatggct tacgaacggg gcggagattt cctggaagat gccaggaaga tacttaacag
     1921 ggaagtgaga gggccgcggc aaagccgttt ttccataggc tccgcccccc tgacaagcat
     1981 cacgaaatct gacgctcaaa tcagtggtgg cgaaacccga caggactata aagataccag
     2041 gcgtttcccc ctggcggctc cctcgtgcgc tctcctgttc ctgcctttcg gtttaccggt
     2101 gtcattccgc tgttatggcc gcgtttgtct cattccacgc ctgacactca gttccgggta
     2161 ggcagttcgc tccaagctgg actgtatgca cgaacccccc gttcagtccg accgctgcgc
     2221 cttatccggt aactatcgtc ttgagtccaa cccggaaaga catgcaaaag caccactggc
     2281 agcagccact ggtaattgat ttagaggagt tagtcttgaa gtcatgcgcc ggttcaggct
     2341 aaactgaaag gacaagtttt ggtgactgcg ctcctccaag ccagttacct cggttcaaag
     2401 agttggtagc tcagagaacc ttcgaaaaac cgccctgcaa ggcggttttt tcgttttcag
     2461 agcaagagat tacgcgcaga ccaaaacgat ctcaagaaga tcatcttatt aatcagataa
     2521 aatatttcta gctagagtcg atcttcgcca gcagggcgag gatcgtggca tcaccgaacc
     2581 gcgccgtgcg cgggtcgtcg gtgagccaga gtttcagcag gccgcccagg tcgccattga
     2641 tgcgggccag ctcgcggacg tgctcatagt ccacgacgcc cgtgattttg tagccctggc
     2701 cgacggccag caggtaggcc gacaggctca tgccggccgc cgccgccttt tcctcaatcg
     2761 ctcttcgttc gtctggaagg cagtacacct tgataggtgg gctgcccttc ctggttggct
     2821 tggtttcatc agccatccgc ttgccctcat ctgttacgcc ggcggtagcc ggccagcctc
     2881 gcagagcagg attcccgttg agcaccgcca ggtgcgaata agggacagtg aagaaggaac
     2941 acccgctcgc gggtgggcct acttcaccta tcctgcccgg ctgacgccgt tggatacacc
     3001 aaggaaagtc tacacgaacc ctttggcaaa atcctgtata tcgtgcgaaa aaggatggat
     3061 ataccgaaaa aatcgctata atgaccccga agcagggtta tgcagcggaa aagcgctgct
     3121 tccctgctgt tttgtggaat atctaccgac tggaaacagg caaatgcagg aaattactga
     3181 actgagggga caggcgagac acgatgccaa agagctacac cgacgagctg gccgagtggg
     3241 ttgaatcccg cgcggccaag aagcgccggc gtgatgaggc tgcggttgcg ttcctggcgg
     3301 tgagggcgga tgtcgatcga ctctagctag aggatcgatc ctttttaacc catttaccac
     3361 atatacctgc cgttcactat tatttagtga aatgagatat tatgatattt tctgaattgt
     3421 gattaaaaag gcaactttat gcccatgcaa cagaaactat aaaaaataca gagaatgaaa
     3481 agaaacagat agatttttta gttctttagg cccgtagtct gcaaatcctt ttatgatttt
     3541 ctatcaaaca aaagaggaaa atagaccagt tgcaatccaa acgagagtct aatagaatga
     3601 ggtcgaaaag taaatcgcgc gggtttgtta ctgataaagc aggcaagacc taaaatgtgt
     3661 aaagggcaaa gtgtatactt tggcgtcacc ccttacatat tttaggtctt tttttattgt
     3721 gcgtaactaa cttgccatct tcaaacagga gggctggaag aagcagaccg ctaacacagt
     3781 acataaaaaa ggagacatga acgatgaaca tcaaaaagtt tgcaaaacaa gcaacagtat
     3841 taacctttac taccgcactg ctggcaggag gcgcaactca agcgtttgcg aaagaaacga
     3901 accaaaagcc atataaggaa acatacggca tttcccatat tacacgccat gatatgctgc
     3961 aaatccctga acagcaaaaa aatgaaaaat atcaagttcc tgaattcgat tcgtccacaa
     4021 ttaaaaatat ctcttctgca aaaggcctgg acgtttggga cagctggcca ttacaaaacg
     4081 ctgacggcac tgtcgcaaac tatcacggct accacatcgt ctttgcatta gccggagatc
     4141 ctaaaaatgc ggatgacaca tcgatttaca tgttctatca aaaagtcggc gaaacttcta
     4201 ttgacagctg gaaaaacgct ggccgcgtct ttaaagacag cgacaaattc gatgcaaatg
     4261 attctatcct aaaagaccaa acacaagaat ggtcaggttc agccacattt acatctgacg
     4321 gaaaaatccg tttattctac actgatttct ccggtaaaca ttacggcaaa caaacactga
     4381 caactgcaca agttaacgta tcagcatcag acagctcttt gaacatcaac ggtgtagagg
     4441 attataaatc aatctttgac ggtgacggaa aaacgtatca aaatgtacag cagttcatcg
     4501 atgaaggcaa ctacagctca ggcgacaacc atacgctgag agatcctcac tacgtagaag
     4561 ataaaggcca caaatactta gtatttgaag caaacactgg aactgaagat ggctaccaag
     4621 gcgaagaatc tttatttaac aaagcatact atggcaaaag cacatcattc ttccgtcaag
     4681 aaagtcaaaa acttctgcaa agcgataaaa aacgcacggc tgagttagca aacggcgctc
     4741 tcggtatgat tgagctaaac gatgattaca cactgaaaaa agtgatgaaa ccgctgattg
     4801 catctaacac agtaacagat gaaattgaac gcgcgaacgt ctttaaaatg aacggcaaat
     4861 ggtacctgtt cactgactcc cgcggatcaa aaatgacgat tgacggcatt acgtctaacg
     4921 atatttacat gcttggttat gtttctaatt ctttaactgg cccatacaag ccgctgaaca
     4981 aaactggcct tgtgttaaaa atggatcttg atcctaacga tgtaaccttt acttactcac
     5041 acttcgctgt acctcaagcg aaaggaaaca atgtcgtgat tacaagctat atgacaaaca
     5101 gaggattcta cgcagacaaa caatcaacgt ttgcgccaag cttcctgctg aacatcaaag
     5161 gcaagaaaac atctgttgtc aaagacagca tccttgaaca aggacaatta acagttaaca
     5221 aataaaaacg caaaagaaaa tgccgatggc cgcggcgttg tgacaattta ccgaacaact
     5281 ccgcggccgg gaagccgatc tcggcttgaa cgaattgcag atctagggcg gcggatttgt
     5341 cctactcagg agagcgttca ccgacaaaca acagataaaa cgaaaggccc agtctttcga
     5401 ctgagccttt cgttttattt gatgcctgcc
//
