Abstract Detail

Comparative Genomics/Transcriptomics

Marchant, Daniel Blaine [1], Nelms, Brad [2], Walbot, Virginia [2].

Beyond the protoplast: isolating and analyzing fixed cells for single-cell research.

The digestion of the cell wall is a major hurdle for single-cell research in plants as it requires substantial protocol optimization and produces morphologically homogenous and highly fragile protoplasts. Here we provide a protocol for single-cell research using maize anthers fixed prior to cell wall digestion. We found that fixed cells could be subjected to harsher conditions than protoplasts while maintaining RNA of sufficient quality for short read sequencing.  Fixation also preserved cellular morphology – a distinguishing feature of many plant cell types useful for automated cell sorting or manual isolation. In addition, the fraction of cellular release from fixed tissue samples was much higher than that of popular protoplasting protocols. We found that the fixed cell protocol can be broadly applied to a variety of taxa and tissues and should thus facilitate single-cell research across systems and fields of plant biology.

1 - Stanford University, Biology, Gilbert Hall, RM 206, Stanford, CA, 94305, United States
2 - Stanford University, Gilbert Hall, Stanford, CA, 94311, United States

Keywords:
single-cell
transcriptomics
maize
anther
reproduction.

Presentation Type: Oral Paper
Session: CG3, Functional & Comparative Genetics/Genomics III
Location: Virtual/Virtual
Date: Friday, July 31st, 2020
Time: 3:00 PM
Number: CG3001
Abstract ID:207
Candidate for Awards:Margaret Menzel Award