Mature seed-derived callus of the model indica rice variety Kasalath is highly competent in Agrobacterium-mediated transformation.
We previously established an efficient Agrobacterium-mediated transformation system using primary calli derived from mature seeds of the model japonica rice variety Nipponbare. We expected that the shortened tissue culture period would reduce callus browning--a common problem with the indica transformation system during prolonged tissue culture in the undifferentiated state. In this study, we successfully applied our efficient transformation system to Kasalath--a model variety of indica rice. The Luc reporter system is sensitive enough to allow quantitative analysis of the competency of rice callus for Agrobacterium-mediated transformation. We unexpectedly discovered that primary callus of Kasalath exhibits a remarkably high competency for Agrobacterium-mediated transformation compared to Nipponbare. Southern blot analysis and Luc luminescence showed that independent transformation events in primary callus of Kasalath occurred successfully at ca. tenfold higher frequency than in Nipponbare, and single copy T-DNA integration was observed in ~40% of these events. We also compared the competency of secondary callus of Nipponbare and Kasalath and again found superior competency in Kasalath, although the identification and subsequent observation of independent transformation events in secondary callus is difficult due to the vigorous growth of both transformed and non-transformed cells. An efficient transformation system in Kasalath could facilitate the identification of QTL genes, since many QTL genes are analyzed in a Nipponbare × Kasalath genetic background. The higher transformation competency of Kasalath could be a useful trait in the establishment of highly efficient systems involving new transformation technologies such as gene targeting.
Saika
Hiroaki
H
Plant Genetic Engineering Research Unit, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba, Ibaraki, Japan.
Toki
Seiichi
S
D001483:Base Sequence; D015139:Blotting, Southern; D017931:DNA Primers; D005453:Fluorescence; D017343:Genes, Plant; D012275:Oryza / Q000235:genetics / Q000382:microbiology / Q000502:physiology*; D016133:Polymerase Chain Reaction; D040641:Quantitative Trait Loci; D012231:Rhizobium / Q000502:physiology*; D012639:Seeds; D014169:Transformation, Bacterial
D017931:DNA Primers