Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum)
Abstract Background Celiac disease (CD) is caused by specific sequences of gluten proteins found in cereals such as bread wheat (Triticum aestivum ssp. aestivum) and spelt (T. aestivum ssp. spelta). Among them, the α-gliadins display the highest immunogenicity, with four T-cell stimulatory epitopes....
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2017-09-01
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| Series: | Plant Methods |
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| Online Access: | http://link.springer.com/article/10.1186/s13007-017-0222-2 |
| _version_ | 1818543793773740032 |
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| author | Benjamin Dubois Pierre Bertin Yordan Muhovski Emmanuelle Escarnot Dominique Mingeot |
| author_facet | Benjamin Dubois Pierre Bertin Yordan Muhovski Emmanuelle Escarnot Dominique Mingeot |
| author_sort | Benjamin Dubois |
| collection | DOAJ |
| description | Abstract Background Celiac disease (CD) is caused by specific sequences of gluten proteins found in cereals such as bread wheat (Triticum aestivum ssp. aestivum) and spelt (T. aestivum ssp. spelta). Among them, the α-gliadins display the highest immunogenicity, with four T-cell stimulatory epitopes. The toxicity of each epitope sequence can be reduced or even suppressed according to the allelic form of each sequence. One way to address the CD problem would be to make use of this allelic variability in breeding programs to develop safe varieties, but tools to track the presence of toxic epitopes are required. The objective of this study was to develop a tool to accurately detect and quantify the immunogenic content of expressed α-gliadins of spelt and bread wheat. Results Four TaqMan probes that only hybridize to the canonical—i.e. toxic—form of each of the four epitopes were developed and their specificity was demonstrated. Six TaqMan probes targeting stable reference genes were also developed and constitute a tool to normalize qPCR data. The probes were used to measure the epitope expression levels of 11 contrasted spelt accessions and three ancestral diploid accessions of bread wheat and spelt. A high expression variability was highlighted among epitopes and among accessions, especially in Asian spelts, which showed lower epitope expression levels than the other spelts. Some discrepancies were identified between the canonical epitope expression level and the global amount of expressed α-gliadins, which makes the designed TaqMan probes a useful tool to quantify the immunogenic potential independently of the global amount of expressed α-gliadins. Conclusions The results obtained in this study provide useful tools to study the immunogenic potential of expressed α-gliadin sequences from Triticeae accessions such as spelt and bread wheat. The application of the designed probes to contrasted spelt accessions revealed a high variability and interesting low canonical epitope expression levels in the Asian spelt accessions studied. |
| first_indexed | 2024-12-11T22:40:02Z |
| format | Article |
| id | doaj.art-039db3332700471b914c8e1e35f2db16 |
| institution | Directory Open Access Journal |
| issn | 1746-4811 |
| language | English |
| last_indexed | 2024-12-11T22:40:02Z |
| publishDate | 2017-09-01 |
| publisher | BMC |
| record_format | Article |
| series | Plant Methods |
| spelling | doaj.art-039db3332700471b914c8e1e35f2db162022-12-22T00:47:50ZengBMCPlant Methods1746-48112017-09-0113111410.1186/s13007-017-0222-2Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum)Benjamin Dubois0Pierre Bertin1Yordan Muhovski2Emmanuelle Escarnot3Dominique Mingeot4Département Sciences du vivant, Centre wallon de Recherches agronomiques (CRA-W)Earth and Life Institute – Agronomy, Université catholique de Louvain (UCL)Département Sciences du vivant, Centre wallon de Recherches agronomiques (CRA-W)Département Sciences du vivant, Centre wallon de Recherches agronomiques (CRA-W)Département Sciences du vivant, Centre wallon de Recherches agronomiques (CRA-W)Abstract Background Celiac disease (CD) is caused by specific sequences of gluten proteins found in cereals such as bread wheat (Triticum aestivum ssp. aestivum) and spelt (T. aestivum ssp. spelta). Among them, the α-gliadins display the highest immunogenicity, with four T-cell stimulatory epitopes. The toxicity of each epitope sequence can be reduced or even suppressed according to the allelic form of each sequence. One way to address the CD problem would be to make use of this allelic variability in breeding programs to develop safe varieties, but tools to track the presence of toxic epitopes are required. The objective of this study was to develop a tool to accurately detect and quantify the immunogenic content of expressed α-gliadins of spelt and bread wheat. Results Four TaqMan probes that only hybridize to the canonical—i.e. toxic—form of each of the four epitopes were developed and their specificity was demonstrated. Six TaqMan probes targeting stable reference genes were also developed and constitute a tool to normalize qPCR data. The probes were used to measure the epitope expression levels of 11 contrasted spelt accessions and three ancestral diploid accessions of bread wheat and spelt. A high expression variability was highlighted among epitopes and among accessions, especially in Asian spelts, which showed lower epitope expression levels than the other spelts. Some discrepancies were identified between the canonical epitope expression level and the global amount of expressed α-gliadins, which makes the designed TaqMan probes a useful tool to quantify the immunogenic potential independently of the global amount of expressed α-gliadins. Conclusions The results obtained in this study provide useful tools to study the immunogenic potential of expressed α-gliadin sequences from Triticeae accessions such as spelt and bread wheat. The application of the designed probes to contrasted spelt accessions revealed a high variability and interesting low canonical epitope expression levels in the Asian spelt accessions studied.http://link.springer.com/article/10.1186/s13007-017-0222-2Speltα-gliadinCeliac diseaseTaqMan probeReference genesGluten |
| spellingShingle | Benjamin Dubois Pierre Bertin Yordan Muhovski Emmanuelle Escarnot Dominique Mingeot Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum) Plant Methods Spelt α-gliadin Celiac disease TaqMan probe Reference genes Gluten |
| title | Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum) |
| title_full | Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum) |
| title_fullStr | Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum) |
| title_full_unstemmed | Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum) |
| title_short | Development of TaqMan probes targeting the four major celiac disease epitopes found in α-gliadin sequences of spelt (Triticum aestivum ssp. spelta) and bread wheat (Triticum aestivum ssp. aestivum) |
| title_sort | development of taqman probes targeting the four major celiac disease epitopes found in α gliadin sequences of spelt triticum aestivum ssp spelta and bread wheat triticum aestivum ssp aestivum |
| topic | Spelt α-gliadin Celiac disease TaqMan probe Reference genes Gluten |
| url | http://link.springer.com/article/10.1186/s13007-017-0222-2 |
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