Biomaterial Database

Identification and Validation of Reference Genes for RT-qPCR Analysis in Switchgrass under Heavy Metal Stresses. 2020

Junming Zhao, and Man Zhou, and Yu Meng

Department of Grassland Science, Sichuan Agricultural University, Chengdu 611130, China.

Switchgrass (Panicum Virgatum L.) has been recognized as the new energy plant, which makes it ideal for the development of phytoremediation on heavy metal contamination in soils with great potential. This study aimed to screen the best internal reference genes for the real-time quantitative PCR (RT-qPCR) in leaves and roots of switchgrass for investigating its response to various heavy metals, such as cadmium (Cd), lead (Pb), mercury (Hg), chromium (Cr), and arsenic (As). The stability of fourteen candidate reference genes was evaluated by BestKeeper, GeNorm, NormFinder, and RefFinder software. Our results identified U2AF as the best reference gene in Cd, Hg, Cr, and As treated leaves as well as in Hg, Pb, As, and Cr stressed root tissues. In Pb treated leaf tissues, 18S rRNA was demonstrated to be the best reference gene. CYP5 was determined to be the optimal reference gene in Cd treated root tissues. The least stable reference gene was identified to be CYP2 in all tested samples except for root tissues stressed by Pb. To further validate the initial screening results, we used the different sets of combinatory internal reference genes to analyze the expression of two metal transport associated genes (PvZIP4 and PvPDB8) in young leaves and roots of switchgrass. Our results demonstrated that the relative expression of the target genes consistently changed during the treatment when CYP5/UBQ1, U2AF/ACT12, eEF1a/U2AF, or 18S rRNA/ACT12 were combined as the internal reference genes. However, the time-dependent change pattern of the target genes was significantly altered when CYP2 was used as the internal reference gene. Therefore, the selection of the internal reference genes appropriate for specific experimental conditions is critical to ensure the accuracy and reliability of RT-qPCR. Our findings established a solid foundation to further study the gene regulatory network of switchgrass in response to heavy metal stress.

UI	MeSH Term	Description	Entries
D008897	Panicum	A plant genus of the family POACEAE. The seed is one of the EDIBLE GRAINS used in millet cereals and in feed for birds and livestock (ANIMAL FEED). It contains diosgenin (SAPONINS).	Millet, Broomcorn,Millet, Common,Millet, Little,Panicgrass,Panicum miliaceum,Panicum miliare,Panicum virgatum,Proso Millet,Switchgrass,Broomcorn Millet,Common Millet,Little Millet,Millet, Proso,Millets, Proso,Proso Millets,Switchgrasses,miliaceums, Panicum
D010940	Plant Proteins	Proteins found in plants (flowers, herbs, shrubs, trees, etc.). The concept does not include proteins found in vegetables for which PLANT PROTEINS, DIETARY is available.	Plant Protein,Protein, Plant,Proteins, Plant
D012015	Reference Standards	A basis of value established for the measure of quantity, weight, extent or quality, e.g. weight standards, standard solutions, methods, techniques, and procedures used in diagnosis and therapy.	Standard Preparations,Standards, Reference,Preparations, Standard,Standardization,Standards,Preparation, Standard,Reference Standard,Standard Preparation,Standard, Reference
D002352	Carrier Proteins	Proteins that bind or transport specific substances in the blood, within the cell, or across cell membranes.	Binding Proteins,Carrier Protein,Transport Protein,Transport Proteins,Binding Protein,Protein, Carrier,Proteins, Carrier
D001692	Biological Transport	The movement of materials (including biochemical substances and drugs) through a biological system at the cellular level. The transport can be across cell membranes and epithelial layers. It also can occur within intracellular compartments and extracellular compartments.	Transport, Biological,Biologic Transport,Transport, Biologic
D012333	RNA, Messenger	RNA sequences that serve as templates for protein synthesis. Bacterial mRNAs are generally primary transcripts in that they do not require post-transcriptional processing. Eukaryotic mRNA is synthesized in the nucleus and must be exported to the cytoplasm for translation. Most eukaryotic mRNAs have a sequence of polyadenylic acid at the 3' end, referred to as the poly(A) tail. The function of this tail is not known for certain, but it may play a role in the export of mature mRNA from the nucleus as well as in helping stabilize some mRNA molecules by retarding their degradation in the cytoplasm.	Messenger RNA,Messenger RNA, Polyadenylated,Poly(A) Tail,Poly(A)+ RNA,Poly(A)+ mRNA,RNA, Messenger, Polyadenylated,RNA, Polyadenylated,mRNA,mRNA, Non-Polyadenylated,mRNA, Polyadenylated,Non-Polyadenylated mRNA,Poly(A) RNA,Polyadenylated mRNA,Non Polyadenylated mRNA,Polyadenylated Messenger RNA,Polyadenylated RNA,RNA, Polyadenylated Messenger,mRNA, Non Polyadenylated
D012989	Soil Pollutants	Substances which pollute the soil. Use for soil pollutants in general or for which there is no specific heading.	Soil Pollutant,Pollutant, Soil,Pollutants, Soil
D013312	Stress, Physiological	The unfavorable effect of environmental factors (stressors) on the physiological functions of an organism. Prolonged unresolved physiological stress can affect HOMEOSTASIS of the organism, and may lead to damaging or pathological conditions.	Biotic Stress,Metabolic Stress,Physiological Stress,Abiotic Stress,Abiotic Stress Reaction,Abiotic Stress Response,Biological Stress,Metabolic Stress Response,Physiological Stress Reaction,Physiological Stress Reactivity,Physiological Stress Response,Abiotic Stress Reactions,Abiotic Stress Responses,Abiotic Stresses,Biological Stresses,Biotic Stresses,Metabolic Stress Responses,Metabolic Stresses,Physiological Stress Reactions,Physiological Stress Responses,Physiological Stresses,Reaction, Abiotic Stress,Reactions, Abiotic Stress,Response, Abiotic Stress,Response, Metabolic Stress,Stress Reaction, Physiological,Stress Response, Metabolic,Stress Response, Physiological,Stress, Abiotic,Stress, Biological,Stress, Biotic,Stress, Metabolic
D017343	Genes, Plant	The functional hereditary units of PLANTS.	Plant Genes,Gene, Plant,Plant Gene
D053263	Gene Regulatory Networks	Interacting DNA-encoded regulatory subsystems in the GENOME that coordinate input from activator and repressor TRANSCRIPTION FACTORS during development, cell differentiation, or in response to environmental cues. The networks function to ultimately specify expression of particular sets of GENES for specific conditions, times, or locations.	Gene Circuits,Gene Modules,Gene Networks,Transcriptional Networks,Gene Module,Circuit, Gene,Circuits, Gene,Gene Circuit,Gene Network,Gene Regulatory Network,Module, Gene,Modules, Gene,Network, Gene,Network, Gene Regulatory,Network, Transcriptional,Networks, Gene,Networks, Gene Regulatory,Networks, Transcriptional,Regulatory Network, Gene,Regulatory Networks, Gene,Transcriptional Network