Background

Multicellular organisms, such as animals and plants, are made up of many cells, which are grouped together to form different tissues and organs. Proper development requires complex molecular regulation, ranging from changes in gene expression and signaling events to hormonal activity. Despite the efforts made to understand these mechanisms that regulate development, the great morphological complexity of tissues or organs represents a limitation to creating tissue-specific expression profiles [1–4]. However, in recent years, several techniques have been successfully developed and implemented to isolate and characterize the transcriptomic profiles of specific cell populations, such as fluorescent activated cell sorting (FACS), single-cell isolation, single-nuclei isolation, and laser-assisted microdissection (LAM) [5]. LAM is a microscopy-based technique that has established itself as a good technique for isolating and collecting specific cell populations from their tissue context [6–11]. The LAM technique is distinguished from others for not requiring the generation of marker lines or protoplasts or previous information on a large number of marker genes; it only requires knowing the morphology of the tissues to be collected. These characteristics have allowed expanding LAM implementation to different organs of different plant species such as tomato fruit [12, 13], rice [14], California poppy [15], and Arabidopsis [16–18]. Here, we describe a protocol, built on previous work by Chávez Montes et al., 2016 [10], to perform LAM and RNA-seq of four tissue types from the Arabidopsis gynoecium: carpel marginal meristem (CMM) and septum (SEP), both tissues of the medial domain of the gynoecium, and the presumptive carpel wall (PC) and valves (VV), two tissues that make up the lateral domain of the gynoecium [19]. The aim was to identify the transcriptomic profiles that regulate their development. In the implementation of this protocol, the use of slides without the membrane was chosen instead of the commercially available slides (MembraneSlides) for LAM, which was better for the collection of the tissues of interest. Furthermore, we briefly describe the data analysis and gene expression visualization. This protocol should also be applicable for other plant species.