Single Cell Type Specific RNA Isolation and Gene Expression Analysis in Rice Using Laser Capture Microdissection (LCM)-Based Method

Methods Mol Biol. 2021:2238:275-283. doi: 10.1007/978-1-0716-1068-8_18.

Abstract

The success of single cell type-specific gene expression or functional study largely depends on the efficient isolation of high-quality RNA from them. Laser capture microdissection (LCM) is an efficient technique that allows accessing and dissecting out a specific individual cell or cell type from a microscopic heterogeneous tissue in a minimally disruptive way. Here, we describe an efficient and inexpensive LCM-based method for the extraction of RNAs with high yield and integrity from laser-microdissected mesophyll and bundle sheath cells of rice leaf. The integrity of isolated RNA is assessed with bioanalyzer analysis, and the presence of mRNA of a specific gene is validated through RT-PCR. This RNA could further be used for uncovering single cell type-specific gene expression signature using next-generation transcriptome sequence or through regular RT-PCR.

Keywords: Bundle sheath cells; Gene expression; LCM; Laser microdissection; Mesophyll cells; RNA isolation; Rice.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Gene Expression Profiling
  • Gene Expression Regulation, Plant*
  • Laser Capture Microdissection / methods*
  • Oryza / genetics*
  • Oryza / metabolism
  • Plant Proteins / genetics*
  • Plant Proteins / metabolism
  • RNA, Plant / analysis*
  • RNA, Plant / genetics
  • RNA, Plant / isolation & purification
  • Single-Cell Analysis / methods*

Substances

  • Plant Proteins
  • RNA, Plant