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    A novel application for determination of germin gene products in situ
    (2003) Çalişkan, M.; Ozcan, B.; Cuming, A.C.
    In situ nucleic acid hybridization is one of the most powerful techniques developed for localizing the expression site of a particular gene at the cell, tissue and organ levels. This method is especially useful in understanding the function of specific gene products in particular tissues and the relation between tissue function and its localization in the whole structure of an organ. In this system, labeled anti-sense RNA probes were used to hybridize with desired mRNA while labeled sense probes were used as negative control. In general, each of these labeled probes applied to successive sections rather then the same section. That might cause some uncertainty if there are any spatial differences in the expression of gene products. In the current study, in our knowledge for the first time, anti-sense and sense probes were applied to the same sample to overcome any spatial differentiation of gene expression. The technique was successfully used to localize a plant gene product by applying anti-sense probes to the one site of the section while the other site of the same section was reacted with sense probes for control hybridization. © 2003 Taylor and Francis Group, LLC.
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    Spatial localization of germin-like oxalate oxidase genes in developing wheat shoots
    (2004) Çalişkan, M.; Ozcan, B.; Turan, C.; Cuming, A.C.
    When quiescent embryos in dry grains of ripened wheat are isolated and provided with ample water and oxygen at an appropriate temperature, they begin to grow very rapidly. The onset of the growth is signaled by nascent synthesis of germin genes which encodes a relatively rare, water-soluble homop entameric glycoprotein. Germin is resistant to pepsin digestion under conditions that lead to hydrolysis of virtually all other proteins in wheat embryos. Germin proteins have oxalate oxidase activity, an activity that degrades oxalic acid to generate hydrogen peroxide which involves in many aspects of plant development. Following 48-hour imbibition on water, wheat embryos give rise to distinguishable shoot and roots. Shoots comprise coleoptile, leaf primordium and shoot apex. In the current study, non-radioactively labeled germin riboprobes were prepared by in vitro transcription. The riboprobes were used to search and localize germin mRNAs in sections taken from throughout of shoots. The results revealed that although leaf primordium and shoot apex did not show any signals of the presence of germin mRNAs, coleoptiles as a whole tissue displayed germin gene expression on epidermal cells and vascular bundle sheath cells. Among the sections taken from different parts of shoots, the sections from middle part gave the strongest signals on coleoptile cells. © 2004 Taylor and Francis Group, LLC.