High Throughput RNA extraction from corn utilizing the Omni Bead Ruptor Elite bead mill homogenizer and chemagic 360 nucleic acid extractor

Staple crop transcriptomics holds the key to deciphering the intricate language of gene expression in plants, shaping the future of agricultural science and sustainable food production. In a world grappling with the ever-increasing challenges of a growing 8 billion person population, climate change, and diminishing arable land, understanding the molecular mechanisms that underlie crop growth, development, and response to environmental stressors has become paramount. The elucidation of transcriptomes opens new avenues for advancing crop breeding, improving crop yield, and enhancing resilience to environmental fluctuations or disease. The advent of high-throughput sequencing technologies has revolutionized the field of plant transcriptomics, enabling researchers to comprehensively analyze the transcriptome with unprecedented precision and depth. These methods allow us to not only quantify gene expression levels but also provide insights into alternative splicing, post-transcriptional regulation, and non-coding RNA-mediated gene regulation. As a result, it has become increasingly evident that the transcriptome of crops is far more intricate and dynamic than previously envisioned. This ultimately results in the need for more transcriptomes to be analyzed.

However, the transcriptome cannot be analyzed without high quality RNA to sequence. Part of the difficulty in performing RNA purifications from plant tissue comes from the challenging nature of the tissue itself. Unlike animal tissue, plants are surrounded by thick polysaccharide cell walls that makes exposing the genetic material contained within more challenging. Typical methods to resolve this problem involve lengthy and manual steps utilizing techniques such as enzymatic lysis and manual disruption. While effective, these methods can impose considerable time and labor cost to the researcher, as well as increasing the risk for RNA degradation due to exposure to nucleases and heat. A solution to this problem can be found by introducing semiautomated homogenization techniques which resolve issues with reproducibility and scalability of common nucleic acid extractions. Bead mills and other similar devices provide a high shear environment where physical lysis of the tough plant matrix allows for the soluble RNA to be released into a protective lysis buffer where denaturing of the molecule is minimized. Along with ensuring the integrity of the RNA, the lysis buffers provide a liquid medium conducive to maximally remove downstream inhibitors ensuring that both library preparations and other amplification techniques are not impeded when analysis is ultimately preformed.

Herein, we demonstrate integration of the Omni Bead Ruptor Elite bead mill homogenizer into a semi-automated total plant RNA purification workflow using the chemagic™ 360 instrument. This workflow solution not only provides a rapid nucleic acid extraction, but the resulting product is high quality, high quantity and compatible for many RNA analysis methods.

Table 1. Nucleic acid yield and quality results from 12 RNA extractions.