Novel TPLPV-Mediated Virus-Induced Gene Silencing System for Tea Plants (Camellia sinensis)

In the post-genomic era of tea plants, the lack of an efficient genetic transformation technology system hinders the accurate identification of gene functions. Virus-induced gene silencing (VIGS) is a post-transcriptional silencing technology building on the plant's anti-virus mechanism, which does not rely on the stable transformation and regeneration system of plants. In this study, the tripartite tea plant line pattern virus (TPLPV) isolated from tea plants was first developed into an infectious cDNA clone. The infectious cDNA clone of TPLPV, containing an additional 30-bp poly(A) tail at the 3' end of the TPLPV genome, and pCB301 as a binary expression vector were successfully used to infect Nicotiana benthamiana and tea plants and caused the line pattern phenotype of leaves. Furthermore, a VIGS vector was constructed successfully for the silencing of the phytoene desaturase (CsPDS) gene of tea plants. The copy number of TPLPV in tea plants was detected in a range from 14.0 to 1.40 × 10⁸ by a newly established real-time quantitative reverse transcription PCR analysis. The TPLPV-mediated VIGS system reduced the CsPDS expression level to 55.0% with a silencing efficiency of 40.0% and led to a chlorotic phenotype in systemic tea leaves, as demonstrated via vacuum infiltration with 0.8 kPa for 10 min at 45 days postinoculation. The optimal concentration of Agrobacterium suspension containing TPLPV-VIGS recombinant vectors was determined as OD₆₀₀ = 0.6. The newly developed TPLPV-mediated VIGS system serves as an effective tool for gene function analysis in tea plants.