Abstract Text: Virus-induced genome silencing (VIGS) is a valuable research tool that leverages the natural antiviral defense mechanisms of plants to silence or downregulate specific targeted genes. However, studying gene functionality in cotton poses challenges due to its large diploid or allotetraploid genomes and low transformation efficiency. Interestingly, an infectious clone of the cotton leafroll dwarf virus (CLRDV) from Alabama has previously exhibited over 50% infection rates in Gossypium hirsutum L. without inducing any viral symptoms. This study aims to convert the CLRDV clone into a silencing vector and assess the levels of downregulation. Given that CLRDV is a low-titer, phloem-limited virus, it becomes crucial to compare the levels and locations of silencing with an established VIGS system previously used in cotton, tobacco rattle virus (TRV). In designing the CLRDV-VIGS vector, a multicloning site was integrated into the 5’ untranslated region of the virus, allowing the insertion of target gene sequences. The chloroplast-associated genes CLA1 and PDS, known to induce tissue photobleaching when silenced, are utilized to verify the functionality of the CLRDV silencing vector in two host plants: cotton and Nicotiana benthamiana. Currently, the two virus vectors and the two target genes are being tested across all possible combinations in both host plants. This research aims to leverage a novel virus clone to introduce a new method for conducting studies in cotton, potentially offering advantages for certain applications due to its phloem limitation.
