Step-by-step protocols for rice gamete isolation

  • Chenxin Li
  • Hengping Xu
  • Scott D. RussellEmail author
  • Venkatesan SundaresanEmail author
Methods Paper
Part of the following topical collections:
  1. Cellular Omics Methods in Plant Reproduction Research


Key message

A detailed, step-by-step protocol for isolation of rice gametes for transcriptional profiling, with a general workflow that includes controls for RNA contamination from surrounding cells and tissues is presented.


Characterization of the transcriptome and other -omics studies of flowering plant gametes are challenging as a consequence of the small sizes and relative inaccessibility of these cells. Collecting such poorly represented cells is also complicated by potential contamination from surrounding sporophytic, adjacent gametophytic tissues and difficulties in extracting high-quality intact cells. Here we present detailed, step-by-step procedures for collecting intact, unfixed rice (Oryza sativa) egg cells and sperm cells without enzymatic treatments. In addition, we also present a general workflow for assessing sample purity by RT-PCR, using primers specific for marker genes preferentially expressed in surrounding cells and tissues. These protocols should facilitate future studies of genome-scale characterization of gametes in this important model crop.


Oryza sativa Egg cell Sperm cell Gametes Transcriptomes 



We thank Imtiyaz Khanday, Jonathan Gent, Sarah Anderson and Daniel Jones for helpful advice for optimizing the experimental methods. We thank Debra Skinner for assistance in artwork. This research was funded by the National Science Foundation (Award No. IOS-1547760) and the USDA Agricultural Experiment Station (Project No. CA-D-XXX-6973-H).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

497_2019_363_MOESM1_ESM.pdf (476 kb)
Fig. 1: Comparison of egg synergid morphology after isolation. Figure 2: Bioanalyzer traces of egg and sperm total RNA. Table 1: Primers sequences and amplicon lengths (PDF 475 kb)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Plant BiologyUniversity of CaliforniaDavisUSA
  2. 2.Department of Microbiology and Plant BiologyUniversity of OklahomaNormanUSA
  3. 3.Department of Plant SciencesUniversity of CaliforniaDavisUSA

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