DEVELOPMENT OF cDNA NORMALIZATION SYSTEM AND PRELIMINARY TRANSCRIPTION ANALYSIS OF KCS GENES IN APPLE TISSUES

Abstract ALBERT, ZS., DEÁK, CS., MISKÓ, A., TÓTH, M., PAPP, I.: Development of cDNA normalization system and preliminary transcription analysis of KCS genes in apple tissues. Acta univ. agric. et silvic. Mendel. Brun., 2011, LIX, No. 3, pp. 9-12 Wax production is an important aspect of apple (Malus domestica Borkh.) fruit development from both theoretical and practical point of views. The complex molecular mechanism that controls wax biosynthesis is still widely unknown but many studies focused on this topic. We aimed to develop further the experimental framework of these eff orts with a description of an improved reference genes expression system. Results in the literature show that similarities exist among the expression of some housekeeping genes of diff erent plant species. Based on these considerations and on gene expression data from Arabidopsis thaliana, some genes in apple were assigned for analysis. EST sequences of apple were used to design specifi c primers for RT-PCR experiments. Isolation of intact RNA from diff erent apple tissues and performing RT-PCR reaction were also key point in obtaining expression patterns. To monitor DNA contamination of the RNA samples, specifi c primers were used that amplify intron-containing sequences from the cDNA. We found that actin primers can be used for the detection of intron containing genomic DNA, and tubulin primers are good internal controls in RT-PCR experiments. We were able to make a diff erence between tissue-specifi c and tissue-independent gene-expression, furthermore we found tissue specifi c diff erences between the expression patterns of candidate genes, that are potentially involved in wax-biosynthesis. Our results show that KCS1 and KCS4 are overexpressed in the skin tissue, this could mean that these genes have skin-specifi c expression in apple fruit. wax, cuticle, RT-PCR, cDNA, gene expression, apple (Malus domestica

Development of cDNA normalization system and preliminary transcription analysis of KCS genes in apple tissues

Wax production is an important aspect of apple (Malus domestica Borkh.) fruit development from both theoretical and practical point of views. The complex molecular mechanism that controls wax biosynthesis is still widely unknown but many studies focused on this topic. We aimed to develop further the experimental framework of these efforts with a description of an improved reference genes expression system. Results in the literature show that similarities exist among the expression of some housekeeping genes of different plant species. Based on these considerations and on gene expression data from Arabidopsis thaliana, some genes in apple were assigned for analysis. EST sequences of apple were used to design specific primers for RT-PCR experiments. Isolation of intact RNA from different apple tissues and performing RT-PCR reaction were also key point in obtaining expression patterns. To monitor DNA contamination of the RNA samples, specific primers were used that amplify intron-containing sequences from the cDNA. We found that actin primers can be used for the detection of intron containing genomic DNA, and tubulin primers are good internal controls in RT-PCR experiments. We were able to make a difference between tissue-specific and tissue-independent gene-expression, furthermore we found tissue specific differences between the expression patterns of candidate genes, that are potentially involved in wax-biosynthesis. Our results show that KCS1 and KCS4 are overexpressed in the skin tissue, this could mean that these genes have skin-specific expression in apple fruit

Epidermal and exodermal tissue structures are characteristic for the long shelf-life ‘Kárpia’ pepper cultivar

In this study two widely consumed Hungarian bell pepper cultivars ‘Hó’ and ‘Kárpia’, were examined. Aims of the investigations were to describe the postharvest behaviour of the cultivars and assign associated traits to the largely different shelf-life that is well known for these cultivars. ‘Hó’ is a white fruit flesh cultivar with average postharvest storability, while ‘Kárpia’ is of Capia type, with extended shelf-life. Postharvest water loss rates were measured, and differences between the cultivars were found. Interestingly, however, the longer shelf-life ‘Kárpia’ fruits showed higher water loss rate. Bright field microscopy was used to study the tissue structure of the outer pericarp of pepper fruits to find explanation for the good storability of ‘Kárpia’. Significantly more cell layers of the hypodermal collenchyma were found in ‘Kárpia’, than in ‘Hó’. Thicker cell walls were also found in this tissue but also in the epidermis. Fluorescent imaging confirmed these observations and also supported differences in the cuticular thickness between the cultivars. Our results highlight epidermal and hypodermal tissue structures as potential factors to influence the shelf-life of bell pepper fruits