Genome-wide identification of internal reference genes for normalization of gene expression values during endosperm development in wheat

Mu, Junyi; Chen, Lin; Gu, Yunsong; Duan, Luning; Han, Song; Li, Yaxuan; Yan, Yueming; Li, Xiaohui

doi:10.1007/s13353-019-00503-0

Cited by 19 publications

(19 citation statements)

References 35 publications

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“…Interestingly, although the ten unigenes participated in some essential cellular processes, including regulation of chromatin states (EMSY3-1 and EMSY3-2), protein degradation (SUD1), sphingolipid synthesis (ORMDL3), and transcription regulation (CNOT11 and TAF12B), they were not classical housekeeping genes ( Robles et al, 2007 ; Breslow et al, 2010 ; Tsuchiya and Eulgem, 2011 ; Doblas et al, 2013 ; Laribee et al, 2015 ). These results were in line with the findings of transcriptomics survey of reference genes in other plants, which all identified many novel stably expressed genes ( Mu et al, 2019 ; Wang et al, 2019 ; Long et al, 2020 ).…”

Section: Discussionsupporting

confidence: 88%

Organ-Specific Transcriptome Analysis Identifies Candidate Genes Involved in the Stem Specialization of Bermudagrass (Cynodon dactylon L.)

Chen

et al. 2021

Front. Genet.

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As an important warm-season turfgrass and forage grass species with wide applications, bermudagrass (Cynodon dactylon L.) simultaneously has shoot, stolon and rhizome, three types of stems with different physiological functions. To better understand how the three types of stems differentiate and specialize, we generated an organ-specific transcriptome dataset of bermudagrass encompassing 114,169 unigenes, among which 100,878 and 65,901 could be assigned to the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the Gene Ontology (GO) terms, respectively. Using the dataset, we comprehensively analyzed the gene expression of different organs, especially the shoot, stolon and rhizome. The results indicated that six organs of bermudagrass all contained more than 52,000 significantly expressed unigenes, however, only 3,028 unigenes were enrich-expressed in different organs. Paired comparison analyses further indicated that 11,762 unigenes were differentially expressed in the three types of stems. Gene enrichment analysis revealed that 39 KEGG pathways were enriched with the differentially expressed unigenes (DEGs). Specifically, 401 DEGs were involved in plant hormone signal transduction, whereas 1,978 DEGs were transcription factors involved in gene expression regulation. Furthermore, in agreement with the starch content and starch synthase assay results, DEGs encoding starch synthesis-related enzymes all showed the highest expression level in the rhizome. These results not only provided new insights into the specialization of stems in bermudagrass but also made solid foundation for future gene functional studies in this important grass species and other stoloniferous/rhizomatous plants.

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Section: Discussionsupporting

confidence: 88%

Organ-Specific Transcriptome Analysis Identifies Candidate Genes Involved in the Stem Specialization of Bermudagrass (Cynodon dactylon L.)

Chen

et al. 2021

Front. Genet.

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“…But the criteria for choosing RGs in different wheat varieties or under different conditions remain similar. Basically, any candidate genes with more than two isoforms were excluded and the coincident regions among genes with less than two homologs were used for primer designing [30,52]. In this study, due to the high estimated percentage of duplicated genes in oat, one to four copy numbers of RGs were all considered as candidates, among which 18S, GAPDH1, TUV6 and UBC21 are duplicated genes ( Table 1).…”

Section: Discussionmentioning

confidence: 99%

“…Polyploids such as tobacco, potato, rapeseed, camelina and wheat are widely cultivated and economically important. Single-copy genes are usually used as RGs, although they only account for a small proportion in the genomes of polyploids [30,31]. In fact, it is worth noting that most researches on RG selection in polyploids neither display nor discuss the copy number of candidate RGs [32][33][34][35][36].…”

Section: Introductionmentioning

confidence: 99%

Evaluation of duplicated reference genes for quantitative real-time PCR analysis in genome unknown hexaploid oat (Avena sativa L.)

et al. 2020

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Background Oat (Avena sativa L.), a hexaploid crop with unknown genome, has valuable nutritional, medicinal and pharmaceutical uses. However, no suitable RGs (reference genes) for qPCR (quantitative real-time PCR) has been documented for oat yet. Single-copy gene is often selected as RG, which is challengeable or impactable in unexplored polyploids. Results In this study, eleven candidate RGs, including four duplicated genes, were selected from oat transcriptome. The stability and the optimal combination of these candidate RGs were assessed in 18 oat samples by using four statistical algorithms including the ΔCt method, geNorm, NormFinder and BestKeeper. The most stable RGs for “all samples”, “shoots and roots of seedlings”, “developing seeds” and “developing endosperms” were EIF4A (Eukaryotic initiation factor 4A-3), UBC21 (Ubiquitin-Conjugating Enzyme 21), EP (Expressed protein) and EIF4A respectively. Among these RGs, UBC21 was a four-copy duplicated gene. The reliability was validated by the expression patterns of four various genes normalized to the most and the least stable RGs in different sample sets. Conclusions Results provide a proof of concept that the duplicated RG is feasible for qPCR in polyploids. To our knowledge, this study is the first systematic research on the optimal RGs for accurate qPCR normalization of gene expression in different organs and tissues of oat.

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“…But the criteria for choosing RGs in different wheat varieties or under different conditions remain similar. Basically, any candidate genes with more than two isoforms were excluded and the coincident regions among genes with less than two homologs were used for primer designing [30,61]. In this study, due to the high estimated percentage of duplicated genes in oat, one to four copy numbers of RGs were all considered as candidates, among which 18S, GAPDH1, TUV6 and UBC21 are duplicated genes ( Table 1).…”

Section: Discussionmentioning

confidence: 99%

“…Polyploids such as tobacco, potato, rapeseed, camelina and wheat are widely cultivated and economically important. Single-copy genes are usually used as RGs, although they only account for a small proportion in the genomes [30,31] of polyploids. In fact, it is worth noting that most researches on RG selection in polyploids neither display nor discuss the copy number of candidate RGs [32][33][34][35][36].…”

mentioning

confidence: 99%

Evaluation of duplicated reference genes for quantitative real-time PCR analysis in genome unknown hexaploid oat (Avena sativa L.)

Yang

Wang

Aziz

et al. 2020

Preprint

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Background: Oat (Avena sativa L.), a hexaploid crop with unknown genome, has valuable nutritional, medicinal and pharmaceutical uses. However, no suitable RGs (reference genes) for qPCR (quantitative real-time PCR) has been documented for oat yet. Single-copy gene is often selected as RG, which is challengeable or impactable in unexplored polyploids.Results: In this study, eleven candidate RGs, including four duplicated genes, were selected from oat transcriptome. The stability and the optimal combination of these candidate RGs were assessed in 18 oat samples by using four statistical algorithms including the ΔCt method, geNorm, NormFinder and BestKeeper. The most stable RGs for “all samples”, “shoots and roots of seedlings”, “developing seeds” and “developing endosperms” were EIF4A (Eukaryotic initiation factor 4A-3), UBC21 (Ubiquitin-Conjugating Enzyme 21), EP (Expressed protein) and EIF4A respectively. Among these RGs, UBC21 was a four-copy duplicated gene. The reliability was validated by the expression patterns of four various genes normalized to the most and the least stable RGs in different sample sets.Conclusions: Results provide a proof of concept that the duplicated RG is feasible for qPCR in polyploids. To our knowledge, this study is the first systematic research on the optimal RGs for accurate qPCR normalization of gene expression in different organs and tissues of oat.

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Genome-wide identification of internal reference genes for normalization of gene expression values during endosperm development in wheat

Cited by 19 publications

References 35 publications

Organ-Specific Transcriptome Analysis Identifies Candidate Genes Involved in the Stem Specialization of Bermudagrass (Cynodon dactylon L.)

Organ-Specific Transcriptome Analysis Identifies Candidate Genes Involved in the Stem Specialization of Bermudagrass (Cynodon dactylon L.)

Evaluation of duplicated reference genes for quantitative real-time PCR analysis in genome unknown hexaploid oat (Avena sativa L.)

Evaluation of duplicated reference genes for quantitative real-time PCR analysis in genome unknown hexaploid oat (Avena sativa L.)

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