Expression Stability of Internal Reference Gene in Response to Trichoderma Polysporum Infection in Avena Fatua L

Overview

Journal Curr Genet

Specialty Genetics

Date 2021 Jul 21

PMID 34287660

Citations 6

Authors

Haixia Zhu

Yongqiang Ma

Qingyun Guo

Affiliations

Soon will be listed here.

Abstract

Trichoderma polysporum was a pathogenic fungi which showed strong pathogenicity to Avena fatua L. in recent study. The stress response of A. fatua to T. polysporum is mediated by the regulation of gene expression. Quantification of the expression of genes requires normalizing RT-qPCR data using reference genes with stable expression in the system studied as internal standards. To construct a RT-qPCR system suitable for response of A. fatua to T. polysporum, and screen stable internal reference genes, GeNorm, NormFinder, BestKeeper and RefFinde were used to perform SYBR Green-based RT-qPCR analysis on eight candidate internal reference genes (18S, 28S, TUA, UBC, ACT, GAPDH, TBP and EF-1α) in A. fatua samples after inoculation of T. polysporum Strain HZ-31. The results showed that TBP, 18S and UBC were the most stable internal reference genes, TBP and TUA, TBP and GAPDH, 18S and TBP, UBC and 18S were the most suitable combination of the two internal reference genes, which could be used as internal reference genes for functional gene expression analysis during the interaction between T. polysporum and A. fatua. This is the first study describing a set of reference genes with a stable expression under fungi stress in A. fatua. These genes are also candidate reference genes of choice for studies seeking to identify stress-responsive genes in A. fatua.

Citing Articles

Exploring the Post Mortem Interval (PMI) Estimation Model by circRNA in Mouse Liver Tissue.

Fu J, Song B, Qian J, Cheng J, Chiampanichayakul S, Anuchapreeda S Int J Mol Sci. 2025; 26(3).

PMID: 39940814 PMC: 11817265. DOI: 10.3390/ijms26031046.

Identification of reference genes for gene expression assessment in Avena sativa under biotic stress triggered by Blumeria graminis.

Sowa S, Toporowska J, Paczos-Grzeda E Sci Rep. 2024; 14(1):30006.

PMID: 39622934 PMC: 11612269. DOI: 10.1038/s41598-024-81348-4.

Assessment of Candidate Reference Genes for Gene Expression Studies Using RT-qPCR in from Litchi.

Dong D, Huang R, Hu Y, Yang X, Xu D, Jiang Z Genes (Basel). 2023; 14(12).

PMID: 38137037 PMC: 10743022. DOI: 10.3390/genes14122216.

Stable reference gene selection for Ophiocordyceps sinensis gene expression studies under different developmental stages and light-induced conditions.

Tong C, Wei J, Mao X, Pan G, Li C, Zhou Z PLoS One. 2023; 18(4):e0284486.

PMID: 37079619 PMC: 10118168. DOI: 10.1371/journal.pone.0284486.

Genome-Wide Screening and Stability Verification of the Robust Internal Control Genes for RT-qPCR in Filamentous Fungi.

Yang Y, Xu X, Jing Z, Ye J, Li H, Li X J Fungi (Basel). 2022; 8(9).

PMID: 36135677 PMC: 9504127. DOI: 10.3390/jof8090952.

References

Bhatia P, Taylor W, Greenberg A, Wright J . Comparison of glyceraldehyde-3-phosphate dehydrogenase and 28S-ribosomal RNA gene expression as RNA loading controls for northern blot analysis of cell lines of varying malignant potential. Anal Biochem. 1994; 216(1):223-6. DOI: 10.1006/abio.1994.1028. View

Bustin S . Why the need for qPCR publication guidelines?--The case for MIQE. Methods. 2009; 50(4):217-26. DOI: 10.1016/j.ymeth.2009.12.006. View

Chao W, Dogramaci M, Foley M, Horvath D, Anderson J . Selection and validation of endogenous reference genes for qRT-PCR analysis in leafy spurge (Euphorbia esula). PLoS One. 2012; 7(8):e42839. PMC: 3419244. DOI: 10.1371/journal.pone.0042839. View

Chen Y, Tan Z, Hu B, Yang Z, Xu B, Zhuang L . Selection and validation of reference genes for target gene analysis with quantitative RT-PCR in leaves and roots of bermudagrass under four different abiotic stresses. Physiol Plant. 2014; 155(2):138-148. DOI: 10.1111/ppl.12302. View

De Spiegelaere W, Dern-Wieloch J, Weigel R, Schumacher V, Schorle H, Nettersheim D . Reference gene validation for RT-qPCR, a note on different available software packages. PLoS One. 2015; 10(3):e0122515. PMC: 4380439. DOI: 10.1371/journal.pone.0122515. View

Delye C . Unravelling the genetic bases of non-target-site-based resistance (NTSR) to herbicides: a major challenge for weed science in the forthcoming decade. Pest Manag Sci. 2012; 69(2):176-87. DOI: 10.1002/ps.3318. View

Derveaux S, Vandesompele J, Hellemans J . How to do successful gene expression analysis using real-time PCR. Methods. 2009; 50(4):227-30. DOI: 10.1016/j.ymeth.2009.11.001. View

Dheda K, Huggett J, Chang J, Kim L, Bustin S, Johnson M . The implications of using an inappropriate reference gene for real-time reverse transcription PCR data normalization. Anal Biochem. 2005; 344(1):141-3. DOI: 10.1016/j.ab.2005.05.022. View

Berliere M, Duhoux F, Dalenc F, Baurain J, Dellevigne L, Galant C . Tamoxifen and ovarian function. PLoS One. 2013; 8(6):e66616. PMC: 3695994. DOI: 10.1371/journal.pone.0066616. View

10.

Duhoux A, Carrere S, Gouzy J, Bonin L, Delye C . RNA-Seq analysis of rye-grass transcriptomic response to an herbicide inhibiting acetolactate-synthase identifies transcripts linked to non-target-site-based resistance. Plant Mol Biol. 2015; 87(4-5):473-87. DOI: 10.1007/s11103-015-0292-3. View

11.

Faccioli P, Ciceri G, Provero P, Stanca A, Morcia C, Terzi V . A combined strategy of "in silico" transcriptome analysis and web search engine optimization allows an agile identification of reference genes suitable for normalization in gene expression studies. Plant Mol Biol. 2006; 63(5):679-88. DOI: 10.1007/s11103-006-9116-9. View

12.

Gachon C, Mingam A, Charrier B . Real-time PCR: what relevance to plant studies?. J Exp Bot. 2004; 55(402):1445-54. DOI: 10.1093/jxb/erh181. View

13.

Gaines T, Lorentz L, Figge A, Herrmann J, Maiwald F, Ott M . RNA-Seq transcriptome analysis to identify genes involved in metabolism-based diclofop resistance in Lolium rigidum. Plant J. 2014; 78(5):865-76. DOI: 10.1111/tpj.12514. View

14.

Du Y, Yan L, Wang J, Zhan W, Song K, Han X . β1-Adrenoceptor autoantibodies from DCM patients enhance the proliferation of T lymphocytes through the β1-AR/cAMP/PKA and p38 MAPK pathways. PLoS One. 2013; 7(12):e52911. PMC: 3534136. DOI: 10.1371/journal.pone.0052911. View

15.

Jarosova J, Kundu J . Validation of reference genes as internal control for studying viral infections in cereals by quantitative real-time RT-PCR. BMC Plant Biol. 2010; 10:146. PMC: 3095291. DOI: 10.1186/1471-2229-10-146. View

16.

Kozera B, Rapacz M . Reference genes in real-time PCR. J Appl Genet. 2013; 54(4):391-406. PMC: 3825189. DOI: 10.1007/s13353-013-0173-x. View

17.

Lovdal T, Lillo C . Reference gene selection for quantitative real-time PCR normalization in tomato subjected to nitrogen, cold, and light stress. Anal Biochem. 2009; 387(2):238-42. DOI: 10.1016/j.ab.2009.01.024. View

18.

Mallona I, Lischewski S, Weiss J, Hause B, Egea-Cortines M . Validation of reference genes for quantitative real-time PCR during leaf and flower development in Petunia hybrida. BMC Plant Biol. 2010; 10:4. PMC: 2827423. DOI: 10.1186/1471-2229-10-4. View

19.

Petit C, Pernin F, Heydel J, Delye C . Validation of a set of reference genes to study response to herbicide stress in grasses. BMC Res Notes. 2012; 5:18. PMC: 3292489. DOI: 10.1186/1756-0500-5-18. View

20.

Pfaffl M . A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 2001; 29(9):e45. PMC: 55695. DOI: 10.1093/nar/29.9.e45. View