engleski

Optimisation of a viroid RNA binding/elution conditions to CIM monolithic supports

Viroids are subviral entities known as causal agents of a number of agriculturally important plant diseases. Their single-stranded, circular and uncapsidated RNA is only 246-401 nucleotides long and does not translate into proteins. Without coding potential, all their functions, as well as remarkable stability, depend on their secondary/tertiary structure. Potato spindle tuber viroid (PSTVd) was the first viroid discovered and ever since, it has been recognized as highly infectious plant pathogen causing severe damage in potato crops. Lately, it has also been used as a model in a number of studies investigating non-coding RNA-plant interactions. In order to analyze chemical and physical properties of viroid molecules, they need to be obtained as purified molecules in high yield. Optimized conditions of elution and binding of viroid RNA to CIM monolithic supports may serve as the basis for improving current purification protocols which are time consuming and low yield methods. Another possible application is the concentration of the viroid from large water volumes which enables its detection in otherwise unsuitable samples. Preliminary results presented at the 4th Monolith Summer School and Symposium held in 2010 in Portorož (Slovenia), showed 20% PSTVd recovery from DEAE analytical disk by ion exchange chromatography method. Here, we present results obtained by hydrophobic interactions chromatography on C4 analytical disk. Different concentrations of ammonium sulphate in the loading buffer together with different gradients were tested. The results obtained with 3M ammonium sulphate in the loading buffer showed PSTVd recovery that exceeded the one described in the previous report

Potato spindle tuber viroid (PSTVd); CIM monolith supports

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