title: A rapid and robust method of identifying transformed Arabidopsis thaliana seedlings following floral dip transformation.
abstract: BACKGROUND
  The floral dip method of transformation by immersion of inflorescences in a suspension of Agrobacterium is the method of choice for Arabidopsis transformation. The presence of a marker, usually antibiotic- or herbicide-resistance, allows identification of transformed seedlings from untransformed seedlings. Seedling selection is a lengthy process which does not always lead to easily identifiable transformants. Selection for kanamycin-, phosphinothricin- and hygromycin B-resistance commonly takes 7-10 d and high seedling density and fungal contamination may result in failure to recover transformants.
  
  
  RESULTS
  A method for identifying transformed seedlings in as little as 3.25 d has been developed. Arabidopsis T1 seeds obtained after floral dip transformation are plated on 1% agar containing MS medium and kanamycin, phosphinothricin or hygromycin B, as appropriate. After a 2-d stratification period, seeds are subjected to a regime of 4-6 h light, 48 h dark and 24 h light (3.25 d). Kanamycin-resistant and phosphinothricin-resistant seedlings are easily distinguished from non-resistant seedlings by green expanded cotyledons whereas non-resistant seedlings have pale unexpanded cotyledons. Seedlings grown on hygromycin B differ from those grown on kanamycin and phosphinothricin as both resistant and non-resistant seedlings are green. However, hygromycin B-resistant seedlings are easily identified as they have long hypocotyls (0.8-1.0 cm) whereas non-resistant seedlings have short hypocotyls (0.2-0.4 cm).
  
  
  CONCLUSIONS
  The method presented here is an improvement on current selection methods as it allows quicker identification of transformed seedlings: transformed seedlings are easily discernable from non-transformants in as little as 3.25 d in comparison to the 7-10 d required for selection using current protocols.
authors.affiliation: Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge, CB2 3EA, UK. Sam_Harrison@cantab.net
authors.forename: Samuel J
authors.initials: SJ
authors.lastname: Harrison
authors.forename: Ellie K
authors.initials: EK
authors.lastname: Mott
authors.forename: Kate
authors.initials: K
authors.lastname: Parsley
authors.forename: Sue
authors.initials: S
authors.lastname: Aspinall
authors.forename: John C
authors.initials: JC
authors.lastname: Gray
authors.forename: Amanda
authors.initials: A
authors.lastname: Cottage