- S. Ducra (1)
- L.J.W.J. Gilissen (1)
- Z. Housley (1)
- A.R. Iorio (1)
- D. Kolarich (1)
- M. Laimer (1)
- Y. Ma (1)
- A. Martinelli (1)
- G. Marzban (1)
- G. Pagliarani (2)
- R. Paris (2)
- S. Peters (1)
- H. Puehringer (1)
- G. Ricci (1)
- M.F. Schenk (1)
- M.J.M. Smulders (2)
- S. Tartarini (2)
- W.E. Weg van de (3)
- M. Zaccarini (1)
A qRT-PCR assay for the expression of all Mal d 1 isoallergen genes
Pagliarani, G. ; Paris, R. ; Arens, P. ; Tartarini, S. ; Ricci, G. ; Smulders, M.J.M. ; Weg, W.E. van de - \ 2013
BMC Plant Biology 13 (2013). - ISSN 1471-2229 - 25 p.
apple malus-domestica - bet v 1 - major allergen mal-d-1 - ige-binding epitopes - birch pollen - l. borkh - in-vivo - fruit - cultivars - food
Background - A considerable number of individuals suffer from oral allergy syndrome (OAS) to apple, resulting in the avoidance of apple consumption. Apple cultivars differ greatly in their allergenic properties, but knowledge of the causes for such differences is incomplete. Mal d 1 is considered the major apple allergen. For Mal d 1, a wide range of isoallergens and variants exist, and they are encoded by a large gene family. To identify the specific proteins/genes that are potentially involved in the allergy, we developed a PCR assay to monitor the expression of each individual Mal d 1 gene. Gene-specific primer pairs were designed for the exploitation of sequence differences among Mal d 1 genes. The specificity of these primers was validated using both in silico and in vitro techniques. Subsequently, this assay was applied to the peel and flesh of fruits from the two cultivars 'Florina' and 'Gala'. Results - We successfully developed gene-specific primer pairs for each of the 31 Mal d 1 genes and incorporated them into a qRT-PCR assay. The results from the application of the assay showed that 11 genes were not expressed in fruit. In addition, differential expression was observed among the Mal d 1 genes that were expressed in the fruit. Moreover, the expression levels were tissue and cultivar dependent. Conclusion - The assay developed in this study facilitated the first characterisation of the expression levels of all known Mal d 1 genes in a gene-specific manner. Using this assay on different fruit tissues and cultivars, we obtained knowledge concerning gene relevance in allergenicity. This study provides new perspectives for research on both plant breeding and immunotherapy.
Genomic organisation of the Mal d 1 gene cluster on linkage group 16 in apple
Pagliarani, G. ; Paris, R. ; Iorio, A.R. ; Tartarini, S. ; Ducra, S. ; Arens, P. ; Peters, S. ; Weg, W.E. van de - \ 2012
Molecular Breeding 29 (2012)3. - ISSN 1380-3743 - p. 759 - 778.
bet v 1 - birch pollen allergen - x-domestica borkh. - major allergen - duplicate genes - resistance locus - scab resistance - arabidopsis - expression - cloning
European populations exhibit progressive sensitisation to food allergens, and apples are one of the foods for which sensitisation is observed most frequently. Apple cultivars vary greatly in their allergenic characteristics, and a better understanding of the genetic basis of low allergenicity may therefore allow allergic individuals to increase their fruit intake. Mal d 1 is considered to be a major apple allergen, and this protein is encoded by the most complex allergen gene family. Not all Mal d 1 members are likely to be involved in allergenicity. Therefore, additional knowledge about the existence and characteristics of the different Mal d 1 genes is required. In the present study, we investigated the genomic organisation of the Mal d 1 gene cluster in linkage group 16 of apple through the sequencing of two bacterial artificial chromosome clones. The results provided new information on the composition of this family with respect to the number and orientation of functional and pseudogenes and their physical distances. The results were compared with the apple and peach genome sequences that have recently been made available. A broad analysis of the whole apple genome revealed the presence of new genes in this family, and a complete list of the observed Mal d 1 genes is supplied. Thus, this study provides an important contribution towards a better understanding of the genetics of the Mal d 1 family and establishes the basis for further research on allelic diversity among cultivars in relation to variation in allergenicity
Mass spectrometry and pollen allergies
Schenk, M.F. ; Gilissen, L.J.W.J. ; Smulders, M.J.M. ; America, A.H.P. - \ 2010
Expert Review of Proteomics 7 (2010)5. - ISSN 1478-9450 - p. 627 - 630.
bet v 1 - birch pollen - recombinant allergens - cross-reactivity - major allergens - potential use - food allergy - expression - proteomics - isoforms
Localisation and distribution of the major allergens in apple fruits
Marzban, G. ; Puehringer, H. ; Dey, R. ; Brynda, S. ; Ma, Y. ; Martinelli, A. ; Zaccarini, M. ; Weg, W.E. van de; Housley, Z. ; Kolarich, D. ; Altmann, F. ; Laimer, M. - \ 2005
Plant Science 169 (2005)2. - ISSN 0168-9452 - p. 387 - 394.
lipid transfer protein - birch pollen allergen - bet v 1 - malus-domestica - genes - food - positions - mal-d-1 - cloning - family
The importance of apple allergens, in particular Mal d 1, a Bet v 1 homologue for the pollen-fruit syndrome in Northern Europe, and Mal d 3, responsible for true fruit allergy in Southern Europe, has been repeatedly emphasized. However, little is known about the distribution pattern of major allergens in fruits and whether differences exist among different cultivars. Transcript expression of Mal d 1 isoforms and Mal d 3 was examined by RealTime-PCR and Northern analysis, respectively. An immuno-tissue-print (ITP) assay was developed to localise major allergens in apple fruit tissue and a Mal d 1 specific, patient independent ELISA was established. ITP analyses show that Mal d 1 and Mal d 2 are distributed throughout the apple pulp and peel, while Mal d 3 is restricted to the peel. Data obtained by ELISA reveal a variation of Mal d 1 content ranging from 0.84 to 33.17 ¿g/g fresh weight in 39 selected cultivars. Different apple cultivars show a markedly different expression of major allergens; this finding will influence the development of diagnostic tools as well as the dietary management of allergic individuals