title: Method: low-cost delivery of the cotton leaf crumple virus-induced gene silencing system.
abstract: BACKGROUND
  We previously developed a virus-induced gene silencing (VIGS) vector for cotton from the bipartite geminivirusCotton leaf crumple virus (CLCrV). The original CLCrV VIGS vector was designed for biolistic delivery by a gene gun. This prerequisite limited the use of the system to labs with access to biolistic equipment. Here we describe the adaptation of this system for delivery by Agrobacterium (Agrobacterium tumefaciens). We also describe the construction of two low-cost particle inflow guns.
  
  
  RESULTS
  The biolistic CLCrV vector was transferred into two Agrobacterium  binary plasmids. Agroinoculation of the binary plasmids into cotton resulted in silencing and GFP expression comparable to the biolistic vector. Two homemade low-cost gene guns were used to successfully inoculate cotton (G. hirsutum)  and N. benthamiana with either the CLCrV VIGS vector or the Tomato golden mosaic virus (TGMV) VIGS vector respectively.
  
  
  CONCLUSIONS
  These innovations extend the versatility of CLCrV-based VIGS for analyzing gene function in cotton. The two low-cost gene guns make VIGS experiments affordable for both research and teaching labs by providing a working alternative to expensive commercial gene guns.
authors.affiliation: Department of Crop Science, North Carolina State University, Raleigh, NC 27695, USA. jrtuttle@ncsu.edu.
authors.forename: John Richard
authors.initials: JR
authors.lastname: Tuttle
authors.forename: Candace H
authors.initials: CH
authors.lastname: Haigler
authors.forename: Dominique
authors.initials: D
authors.lastname: Robertson