Development of mite (Tetranychus urticae)-resistant transgenic Chinese cabbage using plant-mediated RNA interference

摘要

The two-spotted spider mite [Tetranychus urticae (Koch)] is a well-known omnivorous pest with approximately 3800 host crops, including Chinese cabbage (Brassica rapa ssp. pekinesis), one of the most important vegetable crops in Korea. We used plant-mediated RNA interference (RNAi) to produce mite-resistant Chinese cabbage lines and analyzed the molecular and phenotypic characteristics of the transgenic plants. A RNAi vector was constructed containing complementary fragments of the gene encoding coatomer protein complex subunit 2 (COPB2) derived from T. urticae, with both sense- and antisense-orientation, and the phosphinothricin acetyltransferase (Bar) gene, which was used as a selection marker. After Agrobacterium-mediated transformation, T-1 transgenic lines, selected from 24 T-0 transgenic lines, were generated by bud-pollination. The bioassay and reverse transcription quantitative PCR results indicated that the 'CO 17-24' T-1 transgenic line showed high mite resistance with nearly a 100% mortality rate, and expressed approximately 160 times more COPB2 hairpin RNA than the other T-1 transgenic lines. A phenotypic analysis showed no difference in the 26 examined characteristics between non-transgenic and T-1 transgenic lines. We confirmed that the 'CO 17-24' T-1 transgenic line contained one copy of the T-DNA inserted into the intergenic region between Bra039383 and Bra039384. And the inserted T-DNA of the 'CO 17' T-0 transgenic line was stably inherited in 'CO 17-24' without any variation in the internal structure of the T-DNA. In conclusion, 'CO 17-24' is a candidate for an elite line in a breeding program to develop mite-resistant Chinese cabbage. This study demonstrated the effectiveness of the plant-mediated RNAi technique in developing mite-resistant Chinese cabbage, and it will provide useful information for developing pest-resistant plants.