LOCUS       sequence-456311-        6443 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MoClo-compatible vectors for dsRNA production in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6443)
  AUTHORS   Wu CF, Chang LP, Lee C, Stergiopoulos I, Chen LH
  TITLE     pSIG plasmids, MoClo-compatible vectors for efficient production of 
            chimeric double-stranded RNAs in Escherichia coli HT115 (DE3) 
            strain.
  JOURNAL   Plant Methods. 2025 Jul 11;21(1):96. doi: 
            10.1186/s13007-025-01413-5.
  PUBMED    40646633
REFERENCE   2  (bases 1 to 6443)
  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 
            Methods."; date: "2025-07-11"; volume: "21(1):96. doi"; pages: " 
            10.1186/s13007-025-01413-5"
COMMENT     Sequence Label: pSIG1
FEATURES             Location/Qualifiers
     source          1..6443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(34..81)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        82..100
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        123..568
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(569..655,656..1990)
                     /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
                     ITRHDMLQIPEQQKNEKYQVSEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        complement(2034..2052)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      2053..2100
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_feature    2146..2170
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(2326..3474)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      complement(3810..4398)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3890..3909)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4569..5360,5361..5429))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5192..5211
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5430..5534)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5621..6252
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    6344..6368
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 tctgtgaaga caatgccgaa ttcggatccg gagcaaaaaa cccctcaaga cccgtttaga
       61 ggccccaagg ggttatgcta gtaatacgac tcactatagg tactctagtg agacgaaccc
      121 atcacatata cctgccgttc actattattt agtgaaatga gatattatga tattttctga
      181 attgtgatta aaaaggcaac tttatgccca tgcaacagaa actataaaaa atacagagaa
      241 tgaaaagaaa cagatagatt ttttagttct ttaggcccgt agtctgcaaa tccttttatg
      301 attttctatc aaacaaaaga ggaaaataga ccagttgcaa tccaaacgag agtctaatag
      361 aatgaggtcg aaaagtaaat cgcgcgggtt tgttactgat aaagcaggca agacctaaaa
      421 tgtgtaaagg gcaaagtgta tactttggcg tcacccctta catattttag gtcttttttt
      481 attgtgcgta actaacttgc catcttcaaa caggagggct ggaagaagca gaccgctaac
      541 acagtacata aaaaaggaga catgaacgat gaacatcaaa aagtttgcaa aacaagcaac
      601 agtattaacc tttactaccg cactgctggc aggaggcgca actcaagcgt ttgcgaaaga
      661 aacgaaccaa aagccatata aggaaacata cggcatttcc catattacac gccatgatat
      721 gctgcaaatc cctgaacagc aaaaaaatga aaaatatcaa gtttctgaat ttgattcgtc
      781 cacaattaaa aatatctctt ctgcaaaagg cctggacgtt tgggacagct ggccattaca
      841 aaacgctgac ggcactgtcg caaactatca cggctaccac atcgtctttg cattagccgg
      901 agatcctaaa aatgcggatg acacatcgat ttacatgttc tatcaaaaag tcggcgaaac
      961 ttctattgac agctggaaaa acgctggccg cgtctttaaa gacagcgaca aattcgatgc
     1021 aaatgattct atcctaaaag accaaacaca agaatggtca ggttcagcca catttacatc
     1081 tgacggaaaa atccgtttat tctacactga tttctccggt aaacattacg gcaaacaaac
     1141 actgacaact gcacaagtta acgtatcagc atcagacagc tctttgaaca tcaacggtgt
     1201 agaggattat aaatcaatct ttgacggtga cggaaaaacg tatcaaaatg tacagcagtt
     1261 catcgatgaa ggcaactaca gctcaggcga caaccatacg ctgagagatc ctcactacgt
     1321 agaagataaa ggccacaaat acttagtatt tgaagcaaac actggaactg aagatggcta
     1381 ccaaggcgaa gaatctttat ttaacaaagc atactatggc aaaagcacat cattcttccg
     1441 tcaagaaagt caaaaacttc tgcaaagcga taaaaaacgc acggctgagt tagcaaacgg
     1501 cgctctcggt atgattgagc taaacgatga ttacacactg aaaaaagtga tgaaaccgct
     1561 gattgcatct aacacagtaa cagatgaaat tgaacgcgcg aacgtcttta aaatgaacgg
     1621 caaatggtac ctgttcactg actcccgcgg atcaaaaatg acgattgacg gcattacgtc
     1681 taacgatatt tacatgcttg gttatgtttc taattcttta actggcccat acaagccgct
     1741 gaacaaaact ggccttgtgt taaaaatgga tcttgatcct aacgatgtaa cctttactta
     1801 ctcacacttc gctgtacctc aagcgaaagg aaacaatgtc gtgattacaa gctatatgac
     1861 aaacagagga ttctacgcag acaaacaatc aacgtttgcg ccgagcttcc tgctgaacat
     1921 caaaggcaag aaaacatctg ttgtcaaaga cagcatcctt gaacaaggac aattaacagt
     1981 taacaaataa aaacgcaaaa gaaaatgccg atgggtaccg tctcaaacgg cttcctatag
     2041 tgagtcgtat tactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     2101 cgctgcaatt gtcttctgca cgaagtggtt taaactatca gtgtttgaca ggatatattg
     2161 gcgggtaaac ctaagagaaa agagcgttta ttagaataat cggatattta aaagggcgtg
     2221 aaaaggttta tccgttcgtc catttgtatg tgcatgccaa ccacagggtt ccccagatca
     2281 ggcgctggct gctgaacccc cagccggaac tgaccccaca aggccctagc gtttgcaatg
     2341 caccaggtca tcattgaccc aggcgtgttc caccaggccg ctgcctcgca actcttcgca
     2401 ggcttcgccg acctgctcgc gccacttctt cacgcgggtg gaatccgatc cgcacatgag
     2461 gcggaaggtt tccagcttga gcgggtacgg ctcccggtgc gagctgaaat agtcgaacat
     2521 ccgtcgggcc gtcggcgaca gcttgcggta cttctcccat atgaatttcg tgtagtggtc
     2581 gccagcaaac agcacgacga tttcctcgtc gatcaggacc tggcaacggg acgttttctt
     2641 gccacggtcc aggacgcgga agcggtgcag cagcgacacc gattccaggt gcccaacgcg
     2701 gtcggacgtg aagcccatcg ccgtcgcctg taggcgcgac aggcattcct cggccttcgt
     2761 gtaataccgg ccattgatcg accagcccag gtcctggcaa agctcgtaga acgtgaaggt
     2821 gatcggctcg ccgatagggg tgcgcttcgc gtactccaac acctgctgcc acaccagttc
     2881 gtcatcgtcg gcccgcagct cgacgccggt gtaggtgatc ttcacgtcct tgttgacgtg
     2941 gaaaatgacc ttgttttgca gcgcctcgcg cgggattttc ttgttgcgcg tggtgaacag
     3001 ggcagagcgg gccgtgtcgt ttggcatcgc tcgcatcgtg tccggccacg gcgcaatatc
     3061 gaacaaggaa agctgcattt ccttgatctg ctgcttcgtg tgtttcagca acgcggcctg
     3121 cttggcctcg ctgacctgtt ttgccaggtc ctcgccggcg gtttttcgct tcttggtcgt
     3181 catagttcct cgcgtgtcga tggtcatcga cttcgccaaa cctgccgcct cctgttcaag
     3241 acgacgcgaa cgctccacgg cggccgatgg cgcgggcagg gcagggggag ccagttgcac
     3301 gctgtcgcgc tcgatcttgg ccgtagcttg ctggaccatc gagccgacgg actggaaggt
     3361 ttcgcggggc gcacgcatga cggtgcggct tgcgatggtt tcggcatcct cggcggaaaa
     3421 ccccgcgtcg atcagttctt gcctgtatgc cttccggtca aacgtccgat tcattcaccc
     3481 tccttgcggg attgccccga ctcacgccgg ggcaatgtgc ccttattcct gatttgaccc
     3541 gcctggtgcc ttggtgtcca gataatccac cttatcggca atgaagtcgg tcccgtagac
     3601 cgtctggccg tccttctcgt acttggtatt ccgaatcttg ccctgcacga ataccagcga
     3661 ccccttgccc aaatacttgc cgtgggcctc ggcctgagag ccaaaacact tgatgcggaa
     3721 gaagtcggtg cgctcctgct tgtcgccggc atcgttgcgc cacatctagg atctgccagg
     3781 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3841 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     3901 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     3961 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4021 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4081 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4141 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4201 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4261 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4321 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4381 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4441 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4501 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4561 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4621 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4681 tctggcccca gtgctgcaat gataccgcga gaaccacgct caccggctcc agatttatca
     4741 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4801 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4861 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     4921 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     4981 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5041 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5101 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5161 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5221 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5281 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5341 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5401 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5461 tatcaaggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     5521 ataggggttc cgcgcacgaa ttggccagcg ctgccatttt tggggtgagg ccgttcgcgg
     5581 ccgaggggcg cagcccctgg ggggatggga ggcccgcgtt agcgggccgg gagggttcga
     5641 gaaggggggg cacccccctt cggcgtgcgc ggtcacgcgc acagggcgca gccctggtta
     5701 aaaacaaggt ttataaatat tggtttaaaa gcaggttaaa agacaggtta gcggtggccg
     5761 aaaaacgggc ggaaaccctt gcaaatgctg gattttctgc ctgtggacag cccctcaaat
     5821 gtcaataggt gcgcccctca tctgtcagca ctctgcccct caagtgtcaa ggatcgcgcc
     5881 cctcatctgt cagtagtcgc gcccctcaag tgtcaatacc gcagggcact tatccccagg
     5941 cttgtccaca tcatctgtgg gaaactcgcg taaaatcagg cgttttcgcc gatttgcgag
     6001 gctggccagc tccacgtcgc cggccgaaat cgagcctgcc cctcatctgt caacgccgcg
     6061 ccgggtgagt cggcccctca agtgtcaacg tccgcccctc atctgtcagt gagggccaag
     6121 ttttccgcga ggtatccaca acgccggcgg ccgcggtgtc tcgcacacgg cttcgacggc
     6181 gtttctggcg cgtttgcagg gccatagacg gccgccagcc cagcggcgag ggcaaccagc
     6241 ccggtgagcg tcgcaaagga gatcctgatc tgactgatgg gctgcctgta tcgagtggtg
     6301 attttgtgcc gagctgccgg tcggggagct gttggctggc tggtggcagg atatattgtg
     6361 gtgtaaacaa attgacgctt agacaactta ataacacatt gcggacgttt ttaatgtact
     6421 ggggtggatg cagtgggccc cac
//
