LOCUS       sequence-490874-        7432 bp    DNA     circular SYN 28-OCT-2025
DEFINITION  Donor I vector in DASH and omega1 vector in GoldenBraid/DASH, with 
            attBTT and attBCC-FRT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7432)
  AUTHORS   Zhao C, Stepanova AN, Alonso JM
  TITLE     DASH: a versatile and high-capacity gene stacking system for plant 
            synthetic biology.
  JOURNAL   Plant Biotechnol J. 2025 Jun 10. doi: 10.1111/pbi.70179.
  PUBMED    40493021
REFERENCE   2  (bases 1 to 7432)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 28, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plant 
            Biotechnol J. 2025 Jun 10. doi: 10.1111/pbi.70179."
COMMENT     Sequence Label: pDASH-DI-omega-1
FEATURES             Location/Qualifiers
     source          1..7432
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..25
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     protein_bind    48..81
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     primer_bind     411..433
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     425..442
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     426..442
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    443..499
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(512..528)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(512..528)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(525..547)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    536..552
                     /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)."
     promoter        complement(join(560..566,567..584,585..590))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    605..626
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     protein_bind    complement(754..787)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    794..818
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             2119..2748
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             3177..4250
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      4316..4510
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(4746..4768)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4854..4994
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5009..5026)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5180..5768)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5260..5279)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5856..6647)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
ORIGIN
        1 tggcaggata tattgtggtg taaacataac ggatccccgc ggtgcgggtg ccagggcgtg
       61 cccttgggct ccccgggcgc gtactccggt ctcaggagtg agacgcacag cttgtctgta
      121 agcggatgcc gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg
      181 gggctggctt aactatgcgg catcagagca gattgtactg agagtgcacc atatgcggtg
      241 tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgccatt cgccattcag
      301 gctgcgcaac tgttgggaag ggcgatcggc gcaggcctct tcgctattac gccagctggc
      361 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg
      421 acgttgtaaa acgacggcca gtgaattcga gctcggtacc cggggatcct ctagagtcga
      481 cctgcaggca tgcaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
      541 atccgctcac aattccacac aacatacgag ccggaagcat atagtgtaaa gcctggggtg
      601 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
      661 gaaacctgtc gtgccagctg cgtctcacgc tgtcatgaga ccccgcggtg cgggtgccag
      721 ggcgtgcccc cgggctcccc gggcgcgtac tccgaagttc ctatactttc tagagaatag
      781 gaacttcgga tcctgacagg atatattggc gggtaaacct aagagaaaag agcgtttatt
      841 agaataatcg gatatttaaa agggcgtgaa aaggtttatc cgttcgtcca tttgtatgtg
      901 catgccaacc acagggttcc cctcgggatc aaagtacttt gatccaaccc ctccgctgct
      961 atagtgcagt cggcttctga cgttcagtgc agccgtcatc tgaaaacgac atgtcgcaca
     1021 agtcctaagt tacgcgacag gctgccgccc tgcccttttc ctggcgtttt cttgtcgcgt
     1081 gttttagtcg cataaagtag aatacttgcg actagaaccg gagacattac gccatgaaca
     1141 agagcgccgc cgctggcctg ctgggctatg cccgcgtcag caccgacgac caggacttga
     1201 ccaaccaacg ggccgaactg cacgcggccg gctgcaccaa gctgttttcc gagaagatca
     1261 ccggcaccag gcgcgaccgc ccggagctgg ccaggatgct tgaccaccta cgccctggcg
     1321 acgttgtgac agtgaccagg ctagaccgcc tggcccgcag cacccgcgac ctactggaca
     1381 ttgccgagcg catccaggag gccggcgcgg gcctgcgtag cctggcagag ccgtgggccg
     1441 acaccaccac gccggccggc cgcatggtgt tgaccgtgtt cgccggcatt gccgagttcg
     1501 agcgttccct aatcatcgac cgcacccgga gcgggcgcga ggccgccaag gcccgaggcg
     1561 tgaagtttgg cccccgccct accctcaccc cggcacagat cgcgcacgcc cgcgagctga
     1621 tcgaccagga aggccgcacc gtgaaagagg cggctgcact gcttggcgtg catcgctcga
     1681 ccctgtaccg cgcacttgag cgcagcgagg aagtgacgcc caccgaggcc aggcggcgcg
     1741 gtgccttccg tgaggacgca ttgaccgagg ccgacgccct ggcggccgcc gagaatgaac
     1801 gccaagagga acaagcatga aaccgcacca ggacggccag gacgaaccgt ttttcattac
     1861 cgaagagatc gaggcggaga tgatcgcggc cgggtacgtg ttcgagccgc ccgcgcacct
     1921 ctcaaccgtg cggctgcatg aaatcctggc cggtttgtct gatgccaagc tggcggcctg
     1981 gccggccagc ttggccgctg aagaaaccga gcgccgccgt ctaaaaaggt gatgtgtatt
     2041 tgagtaaaac agcttgcgtc atgcggtcgc tgcgtatatg atgcgatgag taaataaaca
     2101 aatacgcaag gggaacgcat gaaggttatc gctgtactta accagaaagg cgggtcaggc
     2161 aagacgacca tcgcaaccca tctagcccgc gccctgcaac tcgccggggc cgatgttctg
     2221 ttagtcgatt ccgatcccca gggcagtgcc cgcgattggg cggccgtgcg ggaagatcaa
     2281 ccgctaaccg ttgtcggcat cgaccgcccg acgattgacc gcgacgtgaa ggccatcggc
     2341 cggcgcgact tcgtagtgat cgacggagcg ccccaggcgg cggacttggc tgtgtccgcg
     2401 atcaaggcag ccgacttcgt gctgattccg gtgcagccaa gcccttacga catatgggcc
     2461 accgccgacc tggtggagct ggttaagcag cgcattgagg tcacggatgg aaggctacaa
     2521 gcggcctttg tcgtgtcgcg ggcgatcaaa ggcacgcgca tcggcggtga ggttgccgag
     2581 gcgctggccg ggtacgagct gcccattctt gagtcccgta tcacgcagcg cgtgagctac
     2641 ccaggcactg ccgccgccgg cacaaccgtt cttgaatcag aacccgaggg cgacgctgcc
     2701 cgcgaggtcc aggcgctggc cgctgaaatt aaatcaaaac tcatttgagt taatgaggta
     2761 aagagaaaat gagcaaaagc acaaacacgc taagtgccgg ccgtccgagc gcacgcagca
     2821 gcaaggctgc aacgttggcc agcctggcag acacgccagc catgaagcgg gtcaactttc
     2881 agttgccggc ggaggatcac accaagctga agatgtacgc ggtacgccaa ggcaagacca
     2941 ttaccgagct gctatctgaa tacatcgcgc agctaccaga gtaaatgagc aaatgaataa
     3001 atgagtagat gaattttagc ggctaaagga ggcggcatgg aaaatcaaga acaaccaggc
     3061 accgacgccg tggaatgccc catgtgtgga ggaacgggcg gttggccagg cgtaagcggc
     3121 tgggttgcct gccggccctg caatggcact ggaaccccca agcccgagga atcggcgtga
     3181 gcggtcgcaa accatccggc ccggtacaaa tcggcgcggc gctgggtgat gacctggtgg
     3241 agaagttgaa ggccgcgcag gccgcccagc ggcaacgcat cgaggcagaa gcacgccccg
     3301 gtgaatcgtg gcaagcggcc gctgatcgaa tccgcaaaga atcccggcaa ccgccggcag
     3361 ccggtgcgcc gtcgattagg aagccgccca agggcgacga gcaaccagat tttttcgttc
     3421 cgatgctcta tgacgtgggc acccgcgata gtcgcagcat catggacgtg gccgttttcc
     3481 gtctgtcgaa gcgtgaccga cgagctggcg aggtgatccg ctacgagctt ccagacgggc
     3541 acgtagaggt ttccgcaggg ccggccggca tggccagtgt gtgggattac gacctggtac
     3601 tgatggcggt ttcccatcta accgaatcca tgaaccgata ccgggaaggg aagggagaca
     3661 agcccggccg cgtgttccgt ccacacgttg cggacgtact caagttctgc cggcgagccg
     3721 atggcggaaa gcagaaagac gacctggtag aaacctgcat tcggttaaac accacgcacg
     3781 ttgccatgca gcgtacgaag aaggccaaga acggccgcct ggtgacggta tccgagggtg
     3841 aagccttgat tagccgctac aagatcgtaa agagcgaaac cgggcggccg gagtacatcg
     3901 agatcgagct agctgattgg atgtaccgcg agatcacaga aggcaagaac ccggacgtgc
     3961 tgacggttca ccccgattac tttttgatcg atcccggcat cggccgtttt ctctaccgcc
     4021 tggcacgccg cgccgcaggc aaggcagaag ccagatggtt gttcaagacg atctacgaac
     4081 gcagtggcag cgccggagag ttcaagaagt tctgtttcac cgtgcgcaag ctgatcgggt
     4141 caaatgacct gccggagtac gatttgaagg aggaggcggg gcaggctggc ccgatcctag
     4201 tcatgcgcta ccgcaacctg atcgagggcg aagcatccgc cggttcctaa tgtacggagc
     4261 agatgctagg gcaaattgcc ctagcagggg aaaaaggtcg aaaaggactc tttcctgtgg
     4321 atagcacgta cattgggaac ccaaagccgt acattgggaa ccggaacccg tacattggga
     4381 acccaaagcc gtacattggg aaccggtcac acatgtaagt gactgatata aaagagaaaa
     4441 aaggcgattt ttccgcctaa aactctttaa aacttattaa aactcttaaa acccgcctgg
     4501 cctgtgcata actgtctggc cagcgcacag ccgaagagct gcaaaaagcg cctacccttc
     4561 ggtcgctgcg ctccctacgc cccgccgctt cgcgtcggcc tatcgcggcc gctggccgct
     4621 caaaaatggc tggcctacgg ccaggcaatc taccagggcg cggacaagcc gcgccgtcgc
     4681 cactcgaccg ccggcgccca catcaaggca ccctgcctcg cgcgtttcgg tgatgacggt
     4741 gaaaacctct gacacatgca gctcccggtg acggtcacag cttgtctgta agcggatgcc
     4801 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc
     4861 atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg gcatcagagc
     4921 agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa
     4981 aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5041 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5101 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5161 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5221 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5281 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5341 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5401 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5461 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5521 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5581 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg
     5641 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5701 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5761 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5821 cacgttaagg gattttggtc atgcattcta ggtgattatt tgccgactac cttggtgatc
     5881 tcgcctttca cgtagtggac aaattcttcc aactgatctg cgcgcgaggc caagcgatct
     5941 tcttcttgtc caagataagc ctgtctagct tcaagtatga cgggctgata ctgggccggc
     6001 aggcgctcca ttgcccagtc ggcagcgaca tccttcggcg cgattttgcc ggttactgcg
     6061 ctgtaccaaa tgcgggacaa cgtaagcact acatttcgct catcaccagc ccagtcgggc
     6121 ggcgagttcc atagcgttaa ggtttcattt agcgcctcaa atagatcctg ttcaggaacc
     6181 ggatcaaaga gttcctccgc cgctggacct accaaggcaa cgctatgttc tcttgctttt
     6241 gtcagcaaga tagccagatc aatgtcgatc gtggctggct cgaagatacc tgcaagaatg
     6301 tcattgcgct gccattctcc aaattgcagt tcgcgcttag ctggataacg ccacggaatg
     6361 atgtcgtcgt gcacaacaat ggtgacttct acagcgcgga gaatctcgct ctctccaggg
     6421 gaagccgaag tttccaaaag gtcgttgatc aaagctcgcc gcgttgtttc atcaagcctt
     6481 acggtcaccg taaccagcaa atcaatatca ctgtgtggct tcaggccgcc atccactgcg
     6541 gagccgtaca aatgtacggc cagcaacgtc ggttcgagat ggcgctcgat gacgccaact
     6601 acctctgata gttgagtcga tacttcggcg atcaccgctt ccctcataat gtttaacttt
     6661 gttttagggc gactgccctg ctgcgtaaca tcgttgctgc tccataacat caaacatcga
     6721 cccacggcgt aacgcgcttg ctgcttggat gcccgaggca tagactgtac cccaaaaaaa
     6781 cagtcataac aagccatgaa aaccgccact gcgccgttac caccgctgcg ttcggtcaag
     6841 gttctggacc agttgcgtga gcgcatacgc tacttgcatt acagcttacg aaccgaacag
     6901 gcttatgtcc actgggttcg tgccttcatc cgtttccacg gtgtgcgtca cccggcaacc
     6961 ttgggtagca gcgaagtcga ggcatttctg tcctggctgg aacagaactt attatttcct
     7021 tcctcttttc tacagtattt aaagataccc caagaagcta attataacaa gacgaactcc
     7081 aattcactgt tccttgcatt ctaaaacctt aaataccaga aaacagcttt ttcaaagttg
     7141 ttttcaaagt tggcgtataa catagtatcg acggagccga ttttgaaacc gcggtgatca
     7201 caggcagcaa cgctctgtca tcgttacaat caacatgcta ccctccgcga gatcatccgt
     7261 gtttcaaacc cggcagctta gttgccgttc ttccgaatag catcggtaac atgagcaaag
     7321 tctgccgcct tacaacggct ctcccgctga cgccgtcccg gactgatggg ctgcctgtat
     7381 cgagtggtga ttttgtgccg agctgccggt cggggagctg ttggctggct gg
//
