LOCUS       sequence-454417-        6427 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 6427)
  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 6427)
  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: pSIG2
FEATURES             Location/Qualifiers
     source          1..6427
                     /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        115..560
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(561..647,648..1982)
                     /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(2018..2036)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      2037..2084
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_feature    2130..2154
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(2310..3458)
                     /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(3794..4382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3874..3893)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4553..5344,5345..5413))
                     /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     5176..5195
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5414..5518)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5605..6236
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    6328..6352
                     /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 tgagacgaac ccatcacata
      121 tacctgccgt tcactattat ttagtgaaat gagatattat gatattttct gaattgtgat
      181 taaaaaggca actttatgcc catgcaacag aaactataaa aaatacagag aatgaaaaga
      241 aacagataga ttttttagtt ctttaggccc gtagtctgca aatcctttta tgattttcta
      301 tcaaacaaaa gaggaaaata gaccagttgc aatccaaacg agagtctaat agaatgaggt
      361 cgaaaagtaa atcgcgcggg tttgttactg ataaagcagg caagacctaa aatgtgtaaa
      421 gggcaaagtg tatactttgg cgtcacccct tacatatttt aggtcttttt ttattgtgcg
      481 taactaactt gccatcttca aacaggaggg ctggaagaag cagaccgcta acacagtaca
      541 taaaaaagga gacatgaacg atgaacatca aaaagtttgc aaaacaagca acagtattaa
      601 cctttactac cgcactgctg gcaggaggcg caactcaagc gtttgcgaaa gaaacgaacc
      661 aaaagccata taaggaaaca tacggcattt cccatattac acgccatgat atgctgcaaa
      721 tccctgaaca gcaaaaaaat gaaaaatatc aagtttctga atttgattcg tccacaatta
      781 aaaatatctc ttctgcaaaa ggcctggacg tttgggacag ctggccatta caaaacgctg
      841 acggcactgt cgcaaactat cacggctacc acatcgtctt tgcattagcc ggagatccta
      901 aaaatgcgga tgacacatcg atttacatgt tctatcaaaa agtcggcgaa acttctattg
      961 acagctggaa aaacgctggc cgcgtcttta aagacagcga caaattcgat gcaaatgatt
     1021 ctatcctaaa agaccaaaca caagaatggt caggttcagc cacatttaca tctgacggaa
     1081 aaatccgttt attctacact gatttctccg gtaaacatta cggcaaacaa acactgacaa
     1141 ctgcacaagt taacgtatca gcatcagaca gctctttgaa catcaacggt gtagaggatt
     1201 ataaatcaat ctttgacggt gacggaaaaa cgtatcaaaa tgtacagcag ttcatcgatg
     1261 aaggcaacta cagctcaggc gacaaccata cgctgagaga tcctcactac gtagaagata
     1321 aaggccacaa atacttagta tttgaagcaa acactggaac tgaagatggc taccaaggcg
     1381 aagaatcttt atttaacaaa gcatactatg gcaaaagcac atcattcttc cgtcaagaaa
     1441 gtcaaaaact tctgcaaagc gataaaaaac gcacggctga gttagcaaac ggcgctctcg
     1501 gtatgattga gctaaacgat gattacacac tgaaaaaagt gatgaaaccg ctgattgcat
     1561 ctaacacagt aacagatgaa attgaacgcg cgaacgtctt taaaatgaac ggcaaatggt
     1621 acctgttcac tgactcccgc ggatcaaaaa tgacgattga cggcattacg tctaacgata
     1681 tttacatgct tggttatgtt tctaattctt taactggccc atacaagccg ctgaacaaaa
     1741 ctggccttgt gttaaaaatg gatcttgatc ctaacgatgt aacctttact tactcacact
     1801 tcgctgtacc tcaagcgaaa ggaaacaatg tcgtgattac aagctatatg acaaacagag
     1861 gattctacgc agacaaacaa tcaacgtttg cgccgagctt cctgctgaac atcaaaggca
     1921 agaaaacatc tgttgtcaaa gacagcatcc ttgaacaagg acaattaaca gttaacaaat
     1981 aaaaacgcaa aagaaaatgc cgatgggtac cgtctcacct atagtgagtc gtattactag
     2041 cataacccct tggggcctct aaacgggtct tgaggggttt tttgcgctgc aattgtcttc
     2101 tgcacgaagt ggtttaaact atcagtgttt gacaggatat attggcgggt aaacctaaga
     2161 gaaaagagcg tttattagaa taatcggata tttaaaaggg cgtgaaaagg tttatccgtt
     2221 cgtccatttg tatgtgcatg ccaaccacag ggttccccag atcaggcgct ggctgctgaa
     2281 cccccagccg gaactgaccc cacaaggccc tagcgtttgc aatgcaccag gtcatcattg
     2341 acccaggcgt gttccaccag gccgctgcct cgcaactctt cgcaggcttc gccgacctgc
     2401 tcgcgccact tcttcacgcg ggtggaatcc gatccgcaca tgaggcggaa ggtttccagc
     2461 ttgagcgggt acggctcccg gtgcgagctg aaatagtcga acatccgtcg ggccgtcggc
     2521 gacagcttgc ggtacttctc ccatatgaat ttcgtgtagt ggtcgccagc aaacagcacg
     2581 acgatttcct cgtcgatcag gacctggcaa cgggacgttt tcttgccacg gtccaggacg
     2641 cggaagcggt gcagcagcga caccgattcc aggtgcccaa cgcggtcgga cgtgaagccc
     2701 atcgccgtcg cctgtaggcg cgacaggcat tcctcggcct tcgtgtaata ccggccattg
     2761 atcgaccagc ccaggtcctg gcaaagctcg tagaacgtga aggtgatcgg ctcgccgata
     2821 ggggtgcgct tcgcgtactc caacacctgc tgccacacca gttcgtcatc gtcggcccgc
     2881 agctcgacgc cggtgtaggt gatcttcacg tccttgttga cgtggaaaat gaccttgttt
     2941 tgcagcgcct cgcgcgggat tttcttgttg cgcgtggtga acagggcaga gcgggccgtg
     3001 tcgtttggca tcgctcgcat cgtgtccggc cacggcgcaa tatcgaacaa ggaaagctgc
     3061 atttccttga tctgctgctt cgtgtgtttc agcaacgcgg cctgcttggc ctcgctgacc
     3121 tgttttgcca ggtcctcgcc ggcggttttt cgcttcttgg tcgtcatagt tcctcgcgtg
     3181 tcgatggtca tcgacttcgc caaacctgcc gcctcctgtt caagacgacg cgaacgctcc
     3241 acggcggccg atggcgcggg cagggcaggg ggagccagtt gcacgctgtc gcgctcgatc
     3301 ttggccgtag cttgctggac catcgagccg acggactgga aggtttcgcg gggcgcacgc
     3361 atgacggtgc ggcttgcgat ggtttcggca tcctcggcgg aaaaccccgc gtcgatcagt
     3421 tcttgcctgt atgccttccg gtcaaacgtc cgattcattc accctccttg cgggattgcc
     3481 ccgactcacg ccggggcaat gtgcccttat tcctgatttg acccgcctgg tgccttggtg
     3541 tccagataat ccaccttatc ggcaatgaag tcggtcccgt agaccgtctg gccgtccttc
     3601 tcgtacttgg tattccgaat cttgccctgc acgaatacca gcgacccctt gcccaaatac
     3661 ttgccgtggg cctcggcctg agagccaaaa cacttgatgc ggaagaagtc ggtgcgctcc
     3721 tgcttgtcgc cggcatcgtt gcgccacatc taggatctgc caggaaccgt aaaaaggccg
     3781 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3841 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3901 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3961 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4021 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4081 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4141 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4201 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4261 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4321 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4381 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4441 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4501 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4561 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4621 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4681 caatgatacc gcgagaacca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4741 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4801 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4861 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4921 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4981 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5041 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5101 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5161 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5221 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5281 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5341 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5401 gttgaatact catactcttc ctttttcaat attattgaag catttatcaa ggttattgtc
     5461 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5521 cgaattggcc agcgctgcca tttttggggt gaggccgttc gcggccgagg ggcgcagccc
     5581 ctggggggat gggaggcccg cgttagcggg ccgggagggt tcgagaaggg ggggcacccc
     5641 ccttcggcgt gcgcggtcac gcgcacaggg cgcagccctg gttaaaaaca aggtttataa
     5701 atattggttt aaaagcaggt taaaagacag gttagcggtg gccgaaaaac gggcggaaac
     5761 ccttgcaaat gctggatttt ctgcctgtgg acagcccctc aaatgtcaat aggtgcgccc
     5821 ctcatctgtc agcactctgc ccctcaagtg tcaaggatcg cgcccctcat ctgtcagtag
     5881 tcgcgcccct caagtgtcaa taccgcaggg cacttatccc caggcttgtc cacatcatct
     5941 gtgggaaact cgcgtaaaat caggcgtttt cgccgatttg cgaggctggc cagctccacg
     6001 tcgccggccg aaatcgagcc tgcccctcat ctgtcaacgc cgcgccgggt gagtcggccc
     6061 ctcaagtgtc aacgtccgcc cctcatctgt cagtgagggc caagttttcc gcgaggtatc
     6121 cacaacgccg gcggccgcgg tgtctcgcac acggcttcga cggcgtttct ggcgcgtttg
     6181 cagggccata gacggccgcc agcccagcgg cgagggcaac cagcccggtg agcgtcgcaa
     6241 aggagatcct gatctgactg atgggctgcc tgtatcgagt ggtgattttg tgccgagctg
     6301 ccggtcgggg agctgttggc tggctggtgg caggatatat tgtggtgtaa acaaattgac
     6361 gcttagacaa cttaataaca cattgcggac gtttttaatg tactggggtg gatgcagtgg
     6421 gccccac
//
