The Use of Induced Somatic Sector Analysis (ISSA) for Studying Genes and Promoters Involved in Wood Formation and Secondary Stem Development.

Secondary stem growth in trees and associated wood formation are significant both from biological and commercial perspectives. However, relatively little is known about the molecular control that governs their development. This is in part due to physical, resource and time limitations often associated with the study of secondary growth processes. A number of in vitro techniques have been used involving either plant part or whole plant system in both woody and non-woody plant species. However, questions about their applicability for the study of secondary stem growth processes, the recalcitrance of certain species and labor intensity are often prohibitive for medium to high throughput applications. Also, when looking at secondary stem development and wood formation the specific traits under investigation might only become measurable late in a tree's lifecycle after several years of growth. In addressing these challenges alternative in vivo protocols have been developed, named Induced Somatic Sector Analysis, which involve the creation of transgenic somatic tissue sectors directly in the plant's secondary stem. The aim of this protocol is to provide an efficient, easy and relatively fast means to create transgenic secondary plant tissue for gene and promoter functional characterization that can be utilized in a range of tree species. Results presented here show that transgenic secondary stem sectors can be created in all live tissues and cell types in secondary stems of a variety of tree species and that wood morphological traits as well as promoter expression patterns in secondary stems can be readily assessed facilitating medium to high throughput functional characterization.

The fasciclin-like arabinogalactan protein family of Eucalyptus grandis contains members that impact wood biology and biomechanics.

Fasciclin-like arabinogalactan protein (FLA) families have been identified and characterised in key plant species, with some members exhibiting functional specialization. Here we identify the FLA family of Eucalyptus grandis, and investigate the roles of three single-FAS domain FLAs, with particular focus on secondary cell-wall formation and wood properties. We use various in-silico approaches to identify and characterise E. grandis genome FLAs, and perform phylogenetic comparisons with other species. For three key FLAs, we perform functional testing including promoter-reporter and overexpression transgenic approaches using eucalypts, poplar and tobacco. Of the 18 eucalypt FLAs identified, several were specifically and highly expressed in stems. The specificity to stem xylem vessel and fibre development was demonstrated with EniFLA1promoter:GUS studies in several species. Testing of select eucalypt FLAs resulted in altered wood development and properties, for example 35S:EgrFLA2 led to a 3 degree reduction in cellulose microfibril angle in eucalypt xylem fibres, and 35S:EgrFLA3 to a reduction in tobacco stem flexural strength. These results indicate that the eucalypt FLA family contains diverse members, and particular members with single FAS domains that are functionally specialized for secondary cell wall growth and properties.

Coloured overlays in schools: orthoptic and optometric findings.

In two studies, the first in a school in Peterborough and the second in a school in Norwich, more than 233 children aged 8-12 years received either an orthoptic examination, or an optometric examination, together with an examination using coloured overlays and a test of reading fluency. In both studies more than one-third of the children reported visual symptoms. More than one-third of the children chose to use an overlay, and they read more quickly with it than without. The colour of the overlay chosen was weakly related to the binocular amplitude of accommodation: overlays reflecting greater energy at long wavelengths were chosen more frequently by children with a higher amplitude of accommodation. Although the visual symptoms were strongly related to the use of an overlay, in neither study was the benefit from an overlay strongly related to the orthoptic or optometric findings. Nevertheless, children who used an overlay had slightly, but significantly, reduced mean binocular amplitude of accommodation and fusional reserves. On average, children with 'sensory' or 'motor' instability of the nonius strips of the Mallett unit read more slowly than others, as did those with poor stereopsis. However, 60% of those demonstrating sustained overlay use gave a normal response on the Mallett aligning prism test, compared with 80% of those who did not use an overlay for a sustained period. Another indicator of decompensated heterophoria, Sheard's criterion, did not differentiate subjects who used overlays from those who did not. Although binocular and accommodative anomalies do not appear to be the underlying mechanism for the benefit from coloured filters in most cases, there may be some individuals who respond to coloured filters and in whom these ocular motor factors require treatment. Children with visually precipitated symptoms and/or reading difficulties need both a careful evaluation of their accommodative and binocular status, and an investigation of the effect of coloured filters.