Browsing by Author "Walford, Sally-Ann"

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    Activation of seed-specific genes in leaves and roots of the desiccation tolerant plant, Xerophyta humilis
    (2008) Walford, Sally-Ann; Illing, Nicola; Farrant, Jill M; Denby, Katherine J
    The ability of tissues to survive almost complete loss of cellular water is a trait found throughout the plant kingdom. While this desiccation tolerance is common in seeds of most angiosperms it is rare in their vegetative tissues. Xerophyta humilis (Bak.) Dur and Schintz belongs to a small group of resurrection angiosperms and it possesses the ability to withstand extreme desiccation of greater than 90% in both its seeds and vegetative tissues and return to active metabolism upon rehydration. We have tested the hypothesis that vegetative desiccation tolerance in angiosperms has evolved as an adaptation of seed desiccation tolerance.
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    A framework for the informed normalization of printed microarrays
    (Academy of Science of South Africa, 2007) van Heerden, Johan; Walford, Sally-Ann; Shen, Arthur; Illing, Nicola
    Microarray technology has become an essential part of contemporary molecular biological research. An aspect central to any microarray experiment is that of normalization, a form of data processing directed at removing technical noise while preserving biological meaning, thereby allowing for more accurate interpretations of data. The statistics underlying many normalization methods can appear overwhelming to microarray newcomers, a situation which is further compounded by a lack of accessible, non-statistical descriptions of common approaches to normalization. Normalization strategies significantly affect the analytical outcome of a microarray experiment, and consequently it is important that the statistical assumptions underlying normalization algorithms are understood and met before researchers embark upon the processing of raw microarray data. Many of these assumptions pertain only to whole-genome arrays, and are not valid for custom or directed microarrays. A thorough diagnostic evaluation of the nature and extent to which technical noise affects individual arrays is paramount to the success of any chosen normalization strategy. Here we suggest an approach to normalization based on extensive stepwise exploration and diagnostic assessment of data prior to, and after, normalization. Common data visualization and diagnostic approaches are highlighted, followed by descriptions of popular normalization methods, and the underlying assumptions they are based on, within the context of removing general technical artefacts associated with microarray data.
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    Isolation and characterisation of an Hsp90 homologue from the resurrection plant Xerophyta viscosa
    (2002) Walford, Sally-Ann; Mundree, Sagadevan G; Thomson, Jennifer Ann; Farrant, Jill M
    Prior to this study, a eDNA library of dehydrated Xerophyta viscosa was differentially screened and several genes were found to be upregulated during dehydration. One of these cDNAs was found to share a high degree of sequence identity with the ER-Iocated Hsp90 or Grp94 family of proteins (hereafter referred to as XVGrp94) and forms the basis of this work. The XVGrp94 eDNA was found to be truncated at the 5· terminus and a full length eDNA was isolated using SMART-RACE™ (§witching Mechanism gt 5' end of RNA Iranscript- Random ~mplification of Complementary .!;rids). This eDNA was sequenced and appeared to be a representative of the Hsp90 family of genes. The putative gene contained an ORF (Open Reading frame) potentially coding for an 812 amino acid protein with a calculated size of 92.83 kDa. It shares 85% homology with other Hsp90s from plants and it contains several characteristic features of these proteins. Additionally, it contains the ER (endoplasmic reticulum) targeting and retention signals. Southern blot analysis confirmed the presence of the gene in the X. viscosa genome possibly as a member of a family of closely related genes. Northern blot analysis revealed a transcript size of 2.8 kb, however, expression patterns of the transcript could not be established. Western blot analysis showed that the XVGrp94 concentration increased significantly in response to heat and dehydration, and a slight increase was observed in response to conditions of high salt, but no response was seen in response to high light, cold or exogenous ABA (abscisic acid) application. The XVGrp94 open reading frame was cloned into the pProEX HTa expression vector and expressed in E. coli, but purification of the recombinant protein was not successful.