IC4R010-Epigenomic-2013-23539454

Project Title

• Plants regenerated from tissue culture contain stable epigenome changes in rice

The Background of This Project

• The rice genome is DNA methylated in all three cytosine contexts (CG, CHG, CHH, where H=A, T,or C), with high levels of CG and CHG methylation and very low levels of CHH methylation (Feng et al., 2010; Zemach et al., 2010). Whole genome bisulfite sequencing (BS-seq) enables measurement of DNA methylation at single nucleotide resolution and thus allows one to distinguish DNA methylation in different cytosine contexts (Cokus et al., 2008; Lister et al., 2008).

Plant Culture & Treatment

• Wild-type rice (Oryza sativa ssp japonica cv Nipponbare) and regenerated rice lines (in Nipponbare background) were used in this study (Zhou et al., 2006; Vega-Sanchez et al., 2008). Hygromycin wasused as the selection marker in rice transformation. All the resistant plants were selfed for indicated generations (Table 1). Homozygosity was confirmed by PCR analysis of the transgene. Rice seeds were surface-sterilized and transferred to 1/2 MS medium. After germination, rice seedlings were transplanted into soil and kept in a growth chamber at 26/20°C under a 14-hr light/10-hr dark cycle. The rice plants regenerated from untransformed rice callus induced from Nipponbare seeds (WT2011)were prepared as previously described (Zhou et al., 2006; Vega-Sanchez et al., 2008). Rice leaf samples were collected at 3 weeks after transplanted into soil and the rice callus were harvested from the callus inducing media.

Research Findings

• The researchers performed deep BS-seq to map DNA methylation in nine regenerated rice lines in the Nipponbare ecotype background that were transformed by various transgenes and were at various stages of inbreeding after transformation: rice blast resistance lines PiZ-t, PiZ-t-839 (a non-functional PiZ-t), Pi9, and an RNAi line for flowering time regulator Spin1 (Zhou et al., 2006; Vega-Sanchez et al., 2008; Table 1). For the PiZ-t line, both transgenic and non-transgenic T2 and T4 plants were available by genetic segregation of the PiZ-t transgene (Table 1).

Labs working on this Project

• Department of Molecular, Cell and Developmental Biology, University of California,Los Angeles, Los Angeles, United States;
• Department of Plant Pathology, Ohio State University, Columbus, United States;
• Department of Plant and Soil Sciences,Delaware Biotechnology Institute, University of Delaware, Newark, United States;
• Howard Hughes Medical Institute, University of California, Los Angeles,Los Angeles, United States.

Corresponding Author

• Steven E Jacobsen:jacobsen@ucla.edu