mRNA transcription profile of potato (Solanum tuberosum L.) in response to explant cutting

Abstract

The explant is an important parameter in plant tissue culture. Most plant tissue culture studies in the literature have focused almost exclusively on the use of explants as a tool for organogenesis or (micro)propagation, and there is limited information that documents the changes that occur in explants in response to cutting during explant preparation just prior to or during culture. To better understand the molecular changes occurring during the wounding process that accompanies the preparation of an explant, this study examined the mRNA transcription profile of 4-week-old potato (Solanum tuberosum L. cv. Desirée) single-node stem segments. To achieve this, RNA-seq analysis was performed to identify significantly differentially expressed genes (DEGs) that existed between uncut in vitro plantlets and cut explants. DEGs corresponding to specific enzyme profiles were broadly connected to cellular processes in the extracellular region, nucleus, and plasma membrane, and were associated with biosynthesis, carbohydrate metabolism and catabolism, cellular protein modification, growth and development, and response to stress. The RNA-seq data of eight highly up-or down-regulated DEGs was validated by RT-qPCR, showing high positive Spearman and Pearson correlation coefficients (0.73 and 0.95, respectively) between SeqMonk LFC and RT-qPCR LFC. These results provide a novel perspective of the changes taking place within explants when they are cut and serve as a valuable basis for the study of explant-related stress in plant tissue culture in other species. In addition, our findings may contribute to an understanding of the success or failure of the tissue culture protocol and subsequent establishment of in vitro cultures by better appreciating the stress-related molecular changes taking place. Key message mRNA transcription profiles were used to show how differentially expressed genes in cut stem explants of potato during preparation for plant tissue culture constitutes an abiotic stress, although not fatal.

mRNA transcription profile of potato (Solanum tuberosum L.) exposed to ultrasound during different stages of in vitro plantlet development

“…The application of the same analyses to other plant systems would allow for an understanding if these patterns exist in other plant species. A recent study (Teixeira da Silva et al 2019) that essentially employed identical methodology as was used in this study, found that, as in this study, several stress-related DEGs were both up- and down-regulated in cut potato explants relative to intact 4 w plantlets.…”

“…The entire methodology (between “Total RNA isolation and removal of rRNA” and “Protein analysis by two-dimensional (2-D) gel electrophoresis”), save for minor differences, has been reported verbatim in a paper that recently examined the effect of cutting on explant transcriptomic profile (Teixeira da Silva et al 2019).…”

Transcriptomic Response of In Vitro Potato (Solanum tuberosum L.) to Piezoelectric Ultrasound

“…Using the methodology reported earlier Teixeira da Silva et al 2019), total protein was extracted from the five samples (0 h, 24 h, 48 h, 1 week, 4 weeks) in three biological replicates from each sample and analyzed using the Agilent Protein 230 Kit (Agilent, Santa Clara, CA, USA) on an Agilent Bioanalyzer 2100 (Agilent) both under reducing (dithiothreitol) and nonreducing (distilled water) conditions.…”

Genome-wide screening of novel RT-qPCR reference genes for study of GLRaV-3 infection in wine grapes and refinement of an RNA isolation protocol for grape berries

“…5A, B). Actin has been routinely used by many research groups as a reference gene in gene expression studies [40,41,[105][106][107][108][109][110][111][112][113]. Our results raised questions on the validity of conclusions of earlier reports where actin was used as a reference gene in analysis involving grapevine and viral infections.…”