A protocol for in vivo RNA labeling and visualization in tobacco pollen tubes

Abstract

Summary: Here, we present a protocol for labeling and live visualization of RNA-protein complexes in the form of ribonucleoprotein particles (RNPs) in tobacco pollen tubes. We describe steps for constructing RNA-pp7/MS2 tag and biolistic gene-gun-mediated pollen transformation. We then provide detailed procedures for RNA labeling using PP7 aptamer nascent RNA tagging and a fluorescently labeled Pseudomonas aeruginosa PP7 bacteriophage coat protein (PCP) reporter that binds to PP7 RNA stem loops. This protocol is adaptable to other cell types by employing tissue-specific promoters. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• cell biology
• developmental biology
• microscopy
• plant sciences