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Sheng Dongping, Zhang Xiaoyang, Feng Mengting, Ye Jianren, Qiu Xiuwen. Construction and its function analysis of RNA interference vector of cytochrome cyp-13A11 gene in Bursaphelenchus xylophilus[J]. Journal of Beijing Forestry University, 2021, 43(1): 96-102. DOI: 10.12171/j.1000-1522.20200025
Citation: Sheng Dongping, Zhang Xiaoyang, Feng Mengting, Ye Jianren, Qiu Xiuwen. Construction and its function analysis of RNA interference vector of cytochrome cyp-13A11 gene in Bursaphelenchus xylophilus[J]. Journal of Beijing Forestry University, 2021, 43(1): 96-102. DOI: 10.12171/j.1000-1522.20200025

Construction and its function analysis of RNA interference vector of cytochrome cyp-13A11 gene in Bursaphelenchus xylophilus

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  • Received Date: January 19, 2020
  • Revised Date: April 08, 2020
  • Available Online: November 23, 2020
  • Published Date: February 04, 2021
  •   Objective  In order to reveal the function of cyp-13A11 gene in Bursaphelenchus xylophilus, the RNA interference vector was constructed and the function of cyp-13A11 gene of Bursaphelenchus xylophilus was analyzed to provide theoretical basis for biological control of pine wilt diseases.
      Method  Specific primers were designed to amplify the cyp-13A11 gene fragment of Bursaphelenchus xylophilus. The products of amplification were constructed into pEASY-T1 interference vector, which was further transferred into Trans1-T1 escherichia coli strain. RNA interference efficiency was determined by qRT-PCR. The feeding reproduction and pathogenicity of pine wood nematodes were investigated after the nematodes soaked in cyp-13A11 dsRNA solution were inoculated on Botrytis cinerea and pine trees, respectively.
      Result  Trans1-T1 strain with pEASY-T1 interference vector was successfully constructed and cyp-13A11 dsRNA was synthesized. RNA interference inhibited the expression of cyp-13A11 gene in Bursaphelenchus xylophilus. In addition, the feeding area was less in cyp-13A11 dsRNA treatment than that in ddH2O treatment, as two thirds of the Botrytis cinerea were fed by nematodes in ddH2O treatment on day 3. After 6 days, one third of Botrytis cinerea was fed by nematodes in cyp-13A11 dsRNA treatment, while almost all of the Botrytis cinerea were exhausted by nematodes in ddH2O treatment. The reproduction number of nematodes in ddH2O treatment was 4.28 times higher than that in cyp-13A11 dsRNA treatment. Furthermore, the wilting rates in both ddH2O and cyp-13A11 dsRNA treatments were 22.2% and 5.6% after Bursaphelenchus xylophilus being inoculated on pine trees for 10 days, respectively. The wilting rates of pine trees in both ddH2O and cyp-13A11 dsRNA treatments were 44.4% and 33.3% after 20 days, respectively. Obviously, the wilting rates were 100% in both treatments after 30 days.
      Conclusion  The RNA interference vector of cyp-13A11 gene in pine wood nematode was successfully constructed. Silencing of cyp-13A11 gene exhibited an important effect on feeding of Bursaphelenchus xylophilus, as well as reduced the reproduction ability and pathogenicity of Bursaphelenchus xylophilus.
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