Establishment and application of TRV-mediated virus-induced gene silencing in cotton

Abstract

Using upland cotton GhCLA1 as marker gene, tobacco rattle virus induced genes silencing (TRV-VIGS) was established in cotton. A gene fragment of TRV subgenomic RNA2 was amplified by reverse transcription-polymerase chain reaction (RT-PCR) in root, stem and leaf of cotton, demonstrating the virus can spread into various organs. The TRV induced silencing of CLA1 gene was further tested in 34 different cotton varieties (lines) originated from several ecological regions of China. The results showed that CLA1 could be silenced in all tested varieties (lines), though the levels of silencing showed a little difference, implicating wide application perspective of TRV-VIGS system in cotton. GhMAPKKK, a mitogen-activated protein kinase (MAPK) kinase kinase gene of cotton, was up-regulated in cotton at 96 hours post inoculation by Verticillium dahliae. Silencing GhMAPKKK in G. barbadense cv. Hai 7124, a cotton variety with high resistance to V. dahlia exhibited reduced resistance to V. dahliae infection, suggesting that GhMAPKKK participated in cotton resistance signaling pathway to V. dahliae. With the wide adaption without genotype selection, sensitivity and high throughput, the TVR-VIGS system will significantly promote functional gene analysis in cotton.