In the vast and versatile world of plant science, understanding gene expression is a cornerstone for researchers worldwide. However, one of the greatest challenges in this endeavor is selecting appropriate reference genes for normalization in quantitative real-time PCR (qPCR) experiments. The selection of suitable internal control genes is crucial for obtaining reliable and reproducible results. Buffalobur (Solanum rostratum Dunal), a notorious invasive species that wreaks havoc on ecosystems, agriculture, human health, and livestock, has recently been the subject of one such research endeavor. A team of scientists has made significant strides in this direction, their findings now published in the journal Scientific Reports, under the DOI: 10.1038/s41598-019-43438-6.
Delving into the world of buffalobur, a plant covered in spines and brimming with toxins, Zhao Dandan and colleagues embarked on a meticulous search for the most stable reference genes to use in gene expression studies involving this species. Studying buffalobur is paramount as it stands as one of the top ten alien invasive species in China, causing extensive damage to local flora and fauna, including economic crops.
The research, supported by non-U.S. government funding, was executed at the Institute of Plant Protection, Chinese Academy of Agricultural Sciences in Beijing, China, and the Mid-Florida Research and Education Center, part of the University of Florida Institute of Food and Agricultural Sciences. The cross-institutional team studied fourteen candidate genes and evaluated their expression stability across various buffalobur tissues, developmental stages, and several stress conditions, using renowned algorithms such as geNorm, NormFinder, BestKeeper, and RefFinder.
In a groundbreaking revelation, the study identified elongation factor 1-alpha (EF1α), actin (ACT), and SAND family protein as the triumvirate of reference genes that exhibited remarkable stability, making them excellent candidates for normalization in buffalobur. This scientific triumph is particularly noteworthy as it is the first report of its kind concerning this invasive species.
The validation of these findings came through the assessment of relative expression levels of two target genes, P5CS and GI, involved respectively in proline biosynthesis—a response to stress—and flowering time regulation. The implications are profound, paving the way for more accurate genetic analyses and potentially aiding in formulating strategies to combat this invasive weed.
This study not only contributes significantly to the scientific understanding of buffalobur but has broader implications for the study of gene expression in plants subjected to various stressors.
References
1. Wei, S., et al. (2010). “Rapid and effective methods for breaking seed dormancy in buffalobur (Solanum rostratum)” Weed Sci., 58, 141–146. DOI: 10.1614/WS-D-09-00005.1
2. Bustin, S., et al. (2005). “Quantitative real-time RT-PCR–a perspective.” J. Mol. Endocrinol., 34, 597–601. DOI: 10.1677/jme.1.01755
3. Huggett, J., et al. (2005). “Real-time RT-PCR normalisation; strategies and considerations.” Genes Immun., 6, 279. DOI: 10.1038/sj.gene.6364190
4. Pfaffl, M. W., et al. (2004). “Determination of stable housekeeping genes, differentially regulated target genes and sample integrity: BestKeeper–Excel-based tool using pair-wise correlations.” Biotechnol. Lett., 26, 509–515. DOI: 10.1023/B:BILE.0000019559.84305.47
5. Vandesompele, J., et al. (2002). “Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes.” Genome Biol., 3, research0034. DOI: 10.1186/1471-2164-3-7-34
Keywords
1. Buffalobur gene expression
2. Reference genes for qPCR
3. Solanum rostratum research
4. qPCR normalization in plants
5. Invasive weed gene studies