Optimising ketocarotenoid production in potato tubers : effect of genetic background, transgene combinations and environment

Astaxanthin is a high value carotenoid produced by some bacteria, a few green algae, several fungi but only a limited number of plants from the genus Adonis. Astaxanthin has been industrially exploited as a feed supplement in poultry farming and aquaculture. Consumption of ketocarotenoids, most notably astaxanthin, is also increasingly associated with a wide range of health benefits,as demonstrated in numerous clinical studies. Currently astaxanthin is produced commercially by chemical synthesis or from algal production systems. Several studies have used a metabolic engineering approach to produce astaxanthin in transgenic plants. Previous attempts to produce transgenic potato tubers biofortified with astaxanthin have met with limited success. In this study we have investigated approaches to optimising tuber astaxanthin content. It is demonstrated that the selection of appropriate parental genotype for transgenic approaches and stacking carotenoid biosynthetic pathway genes with the cauliflower Or gene result in enhanced astaxanthin content, to give six-fold higher tuber astaxanthin content than has been achieved previously. Additionally we demonstrate the effects of growth environment on tuber carotenoid content in both wild type and astaxanthin-producing transgenic lines and describe the associated transcriptome and metabolome restructuring.

What lies beneath? : The pattern and abundance of the subterranean tuber bank of the invasive liana cat’s claw creeper, Macfadyena unguis-cati (Bignoniaceae)

Cat’s claw creeper, Macfadyena unguis-cati (L.) Gentry (Bignoniaceae) is a major environmental weed of riparian areas, rainforest communities and remnant natural vegetation in coastal Queensland and New South Wales, Australia. In densely infested areas, it smothers standing vegetation, including large trees, and causes canopy collapse. Quantitative data on the ecology of this invasive vine are generally lacking. The present study examines the underground tuber traits of M. unguis-cati and explores their links with aboveground parameters at five infested sites spanning both riparian and inland vegetation. Tubers were abundant in terms of density (~1000 per m2), although small in size and low in level of interconnectivity. M. unguis-cati also exhibits multiple stems per plant. Of all traits screened, the link between stand (stem density) and tuber density was the most significant and yielded a promising bivariate relationship for the purposes of estimation, prediction and management of what lies beneath the soil surface of a given M. unguis-cati infestation site. The study also suggests that new recruitment is primarily from seeds, not from vegetative propagation as previously thought. The results highlight the need for future biological-control efforts to focus on introducing specialist seed- and pod-feeding insects to reduce seed-output.

Tensile properties of pumpkin peel and flesh tissue and review of current testing methods

In South and Southeast Asia, postharvest loss causes material waste of up to 66% in fruits and vegetables, 30% in oilseeds and pulses, and 49% in roots and tubers. The efficiency of postharvest equipment directly affects industrial-scale food production. To enhance current processing methods and devices, it is essential to analyze the responses of food materials under loading operations. Food materials undergo different types of mechanical loading during postharvest and processing stages. Therefore, it is important to determine the properties of these materials under different types of loads, such as tensile, compression, and indentation. This study presents a comprehensive analysis of the available literature on the tensile properties of different food samples. The aim of this review was to categorize the available methods of tensile testing for agricultural crops and food materials to investigate an appropriate sample size and tensile test method. The results were then applied to perform tensile tests on pumpkin flesh and peel samples, in particular on arc-sided samples at a constant loading rate of 20 mm min-1. The results showed the maximum tensile stress of pumpkin flesh and peel samples to be 0.535 and 1.45 MPa, respectively. The elastic modulus of the flesh and peel samples was 6.82 and 25.2 MPa, respectively, while the failure modulus values were 14.51 and 30.88 MPa, respectively. The results of the tensile tests were also used to develop a finite element model of mechanical peeling of tough-skinned vegetables. However, to study the effects of deformation rate, moisture content, and texture of the tissue on the tensile responses of food materials, more investigation needs to be done in the future.

Biopharming the SimpliRED™ HIV diagnostic reagent in barley, potato and tobacco

This is the first report of an antibody-fusion protein expressed in transgenic plants for direct use in a medical diagnostic assay. By the use of gene constructs with appropriate promoters, high level expression of an anti-glycophorin single-chain antibody fused to an epitope of the HIV virus was obtained in the leaves and stems of tobacco, tubers of potato and seed of barley. This fusion protein replaces the SimpliRED™ diagnostic reagent, used for detecting the presence of HIV-1 antibodies in human blood. The reagent is expensive and laborious to produce by conventional means since chemical modifications to a monoclonal antibody are required. The plant-produced fusion protein was fully functional (by ELISA) in crude extracts and, for tobacco at least, could be used without further purification in the HIV agglutination assay. All three crop species produced sufficient reagent levels to be superior bioreactors to bacteria or mice, however barley grain was the most attractive bioreactor as it expressed the highest level (150 μg of reagent g-1), is inexpensive to produce and harvest, poses a minuscule gene flow problem in the field, and the activity of the reagent is largely undiminished in stored grain. This work suggests that barley seed will be an ideal factory for the production of antibodies, diagnostic immunoreagents, vaccines and other pharmaceutical proteins.

Product stability and sequestration mechanisms in Solanum tuberosum engineered to biosynthesize high value ketocarotenoids

To produce commercially valuable ketocarotenoids in Solanum tuberosum, the 4, 4′ β-oxygenase (crtW) and 3, 3′ β-hydroxylase (crtZ) genes from Brevundimonas spp. have been expressed in the plant host under constitutive transcriptional control. The CRTW and CRTZ enzymes are capable of modifying endogenous plant carotenoids to form a range of hydroxylated and ketolated derivatives. The host (cv. Désirée) produced significant levels of nonendogenous carotenoid products in all tissues, but at the apparent expense of the economically critical metabolite, starch. Carotenoid levels increased in both wild-type and transgenic tubers following cold storage; however, stability during heat processing varied between compounds. Subcellular fractionation of leaf tissues revealed the presence of ketocarotenoids in thylakoid membranes, but not predominantly in the photosynthetic complexes. A dramatic increase in the carotenoid content of plastoglobuli was determined. These findings were corroborated by microscopic analysis of chloroplasts. In tuber tissues, esterified carotenoids, representing 13% of the total pigment found in wild-type extracts, were sequestered in plastoglobuli. In the transgenic tubers, this proportion increased to 45%, with esterified nonendogenous carotenoids in place of endogenous compounds. Conversely, nonesterified carotenoids in both wild-type and transgenic tuber tissues were associated with amyloplast membranes and starch granules.