Summary for non-specialists Archives - Page 5 of 6

Orange sweet potato champions biofortified foods in Africa

by Charis Cook

Categories: biofortified foods, Summary for non-specialists, teaching resources

Tags: agriculture, biofortified foods, Gates Foundation, nutrition, synthetic biology, vitamin A

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Published on: August 22, 2012

Two Ugandan children dig in to a plate of orange sweet potato (Credit: HarvestPlus)

According to a study published in the Journal of Nutrition this month, eating orange sweet potato reduces the prevelance of vitamin A deficiency in children in Uganda and Mozambique. Vitamin A is critical for the development of good vision as it is an essential component of rhodopsin, a pigment in photoreceptor cells in the eye. Consequently in poor communities in Africa and south-east Asia, where diets poor in vitamin A are widespread, vitamin A deficiency is the leading cause of preventable blindness. Healthy levels of vitamin A are also necessary for normal organ formation and maintenance. Orange-fleshed sweet potato varieties contain more than 50-fold more β-carotene, which is converted to vitamin A after ingestion, than the yellow or white varieties commonly eaten in African countries.

The study monitored the effects of the Orange Sweet Potato (OSP) project, which was funded by the Bill and Melinda Gates foundation and coordinated by HarvestPlus. The conclusions predict a promising future for the use of biofortified foods bred for increased nutritional value. It was the first large-scale study of its kind, involving 24 000 households from Uganda and Mozambique. Nutritionists and farmers educated communities on the health benefits of orange sweet potato and on growing, storing, and commercialising orange sweet potato crops. Local women were also given recipes and information about hygiene practices. (more…)

Resource: Dataset of Arabidopsis genes with a loss-of-function mutant phenotype

by Charis Cook

Categories: methods, resource, Summary for non-specialists

Tags: gene, methods, phenotype, resources

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Published on: August 21, 2012

Highlighted article: J. Lloyd and D. Meinke (2012) A Comprehensive Dataset of Genes with a Loss-of-Function Mutant Phenotype in Arabidopsis thaliana. Plant Physiology, January 2012 pp.111.192393.

In plant science, many published papers involve at least one loss-of-function mutant. A huge number of mutant Arabidopsis lines exist in labs all around the world, detailed in as many journal articles. Now however the genotype and phenotype information for loss-of-function Arabidopsis mutants is stored one place: a dataset assembled by Johnny Lloyd and David Meinke of Oklahoma State University.

Lloyd and Meinke painstakingly went through TAIR, their own database SeedGenes.org, and PubMed to find 2400 Arabidopsis thaliana genes with a loss-of-function mutant phenotype. Out of necessity, the database excludes the effects of under- or over- expression of genes.

The phenotypic effects of gene knock-outs were classified into four groups: essential, morphological, cellular-biochemical, and conditional. The groups were divided into classes reflecting the developmental stage or organ where the phenotype manifests itself, and further divided into subsets which specify the characteristic affected by the phenotype, for example ‘pigmentation’, ‘gamerophyte defective’, and ‘stomata, trichomes’.

The dataset is found in the supplementary data of the paper. Supplemental Table 2 is the complete dataset. On tab 1 the dataset is sorted by locus number and includes 19 columns of information on the gene and the mutant phenotype. This information encompasses the classification of the phenotype, a description of the phenotype, and a reference to the lab in which the research was carried out. (more…)

Synthetic enzyme reduces lignin content

by Charis Cook

Categories: Summary for non-specialists, synthetic biology, teaching resources

Tags: biochemistry, biofuel, cell wall, lignin, synthetic biology

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Published on: August 13, 2012

Public domain image. Source: Glazer, A. W., and Nikaido, H. (1995). Microbial Biotechnology: fundamentals of applied microbiology. San Francisco: W. H. Freeman, p. 340. ISBN 0-71672608-4

Highlighted article: Kewwi Zhang, Mohammad-Wadud Bhuiya, Jorge Rencoret Pazo, Yuchen Miao, Hoon Kim, John Ralph, and Chang-Jun Liu (2012) An Engineered Monolignol 4-O-Methyltransferase Depresses Lignin Biosynthesis and Confers Novel Metabolic Capability in Arabidopsis. Plant Cell Preview.

Zhang et al. reduce lignin content by introducing an artificial enzyme to the cell wall biosynthesis pathway. This is the first time synthetic biology has been used to change cell wall structure, which is usually modified by changing the expression of endogenous enzymes or introducing a protein from another organism. In fact at the moment, synthetic biology is not a common method of manipulating any plant pathway.

Relevant background

public domain image, courtesy of Chino

Lignin is one of three components of secondary cell walls. It is the part which makes extracting sugar from the cell wall, for example for second generation biofuel production, difficult.

Lignin is made up of three monolignols: coniferyl, sinapyl, and p-coumaryl.

They are synthesised in the cytosol and transported to the cell wall. At the cell wall, the monolignols are oxidised, causing their phenol group to become radicalised. The phenoxy radicals polymerise to form the lignin macromolecule.

The Liu lab had the idea of preventing monolignol oxidation by methylation of the phenol group so that the phenoxy radicals were prevented from forming. Their first attempt was to synthesise a selection of monolignol 4-O-methyltransferases (MOMTs). The artificial MOMTS were fusions of two naturally occurring enzymes: lignin biosynthesis pathway methyltransferase COMT, which does not have any 4-O-methyltransferase activity; and fairy fan enzyme isoeugenol O-methyltransferase, which catalyzes 4-O-methylation of isoeugenol and eugenol, but doesn’t affect monolignols. Although several of these artificial enzymes were able to 4-O-methylate monolignols as expected in vitro, they had no activity in vivo.

Results

Zhang et al. used MOMT3, a promising enzyme from their earlier work, as a starting point. (more…)

Transcription factor-like effectors (TALEs)

by Charis Cook

Categories: methods, plant pathogens, Summary for non-specialists, synthetic biology

Tags: methods, plant pathogens, synthetic biology, TALEs

Comments: 2 Comments

Published on: August 8, 2012

Ubud, Bali by Mee Lin Woon; DNA Sequence by schulergd. Via stock.chng

Background

Xanthomonas spp. are plant pathogens that modulate their host’s gene expression in order to facilitate infection. They do this using transcription activator-like effectors (TALEs). Two domains are conserved in TALEs: an N-terminus, required for type III secretion into the plant cells; and a C-terminus with transcription factor activity. In the middle is a set of tandem repeats of amino acids, which mediates binding to host DNA.

As the binding and effector domains of TALEs can be customised, the possibility of using them for molecular and synthetic biology has been explored for some time. They have been used to change gene expression in plants, yeast, and even human cells.

TALEs have been adapted by researchers to make TALE nucleases (TALENs) – TALEs attached to a FOK1 nuclease domain. TALENs work in pairs that flank either side of the target site so that the nuclease domains meet at the point of cleavage. The nucleases cause a double-stranded DNA break, which is fixed imperfectly, causing an insertion or deletions.

In May this year, a paper was published demonstrating the huge impact TALEs could have on agriculture. Li et al. prove that transcription activator-like effector nucleases (TALENs) can be used to render rice resistant to the major agricultural pathogen, Xanthomonas oryzae pv. Oryzae (Xoo). (more…)

Programmed Cell Death in action

by Charis Cook

Categories: Summary for non-specialists

Tags: BMC Series, Lace plant, PCD

Comments: No Comments

Published on: August 7, 2012

Madagascar lace plant. Credit: Jim Yupangco

The lace plant, Aponogeton madagascariensis, is an aquatic plant native to Madagascar. It has characteristic lacey leaves which are formed by programmed cell death (PCD) in the areoles between leaf veins. Wertman et al., from Dalhousie University, used the predictable nature of PCD in lace plants to monitor the process using light, confocal and standing electron microscopy. They were able to discern the precise order of organelle disruption and activity during PCD. They could even video the process using live cell imaging on a light microscope – you can see the cells rupture in front of your eyes in the video below.

Highlighted article: Jaime Wertman, Christina CEN Lord, Adrian N Dauphinee and Arunika HLAN Gunawardena (2012) The pathway of cell dismantling during programmed cell death in lace plant (Aponogeton madagascariensis) leaves. BMC Plant Biology 2012, 12:115

The paper is quite straightforward and a nice one to read so I won’t go in to detail here, but I will quickly go over their findings. After initial anthocyanin reduction, which is how the team identified accurately where and when PCD would take place, the first phase of activity included the bundling of actin microfilaments, and transvacuolar strands increased in number. Then the tonoplast membrane began to fold and mitochondrial aggregates and chloroplasts moved randomly. Nuclear DNA, actin microfilaments and the cell wall broke up. The vacuole swelled and its membrane ruptured while the nucleus shrank and the plasma membrane collapsed. The whole process took roughly 48 hours, and it took more than 24 more hours before the cell wall completely disappeared.

Identifying mutations in Arabidopsis – a faster, cheaper method

by Charis Cook

Categories: methods, Summary for non-specialists

Tags: forward genetics, genetics, methods, mutations, phenotype, Plant Methods

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Published on: July 30, 2012

Highlighted paper: Liu, McCormack and Sheen (2012) Targeted parallel sequencing of large genonmic regions for identifying mutations in Arabidopsis. Plant Methods 8:12

Kun-hsiang Liu, Matthew McCormack and Jen Sheen from Harvard have developed a PCR-based method of identifying mutations in Arabidopsis. It is cheaper and easier than traditional methods of identifying mutations, using bench-top PCR and a new user-friendly method of bioinformatics analysis using web-based resource Galaxy. Liu et al. estimate that using their method to identify a mutation mapped to a 550kb genomic region will cost roughly US$500, a fraction of the usual ten thousand dollar cost of currently used methods of mutant identification.

Liu et al. tested the new method of identifying mutations by searching for new nitrogen response genes. They generated an Arabidopsis thaliana line in which LUCIFERASE was driven by the promoter for nitrogen response marker NIR. Using EMS-mutagenesis, the team made 25 000 mutant NIR:LUC lines and identified seedlings that were nitrate insensitive (nis) or showed nitrate constitutive response (ncr).

When the lines were made, the phenotypes were identified and a second generation was grown. Three second generation lines – ncr1, nis1 and nis2 – were selected for further investigation.

Liu et al. used their novel TPSeq method to locate the mutations causing the ncr1, nis1 and nis2 phenotypes. (more…)

From bench to bountiful harvest … MASC roadmap summarised in current Plant Cell Paper

by Charis Cook

Categories: Summary for non-specialists, teaching resources

Tags: Arabidopsis, MASC, Plant Cell, Ruth Bastow, summary for non-specialists

Comments: No Comments

Published on: July 19, 2012

Highlighted article: Lavagi I., Estelle M., Weckwerth W., Beynon J., and Bastow R. (2012) From Bench to Bountiful Harvests: A Road Map for the Next Decade of Arabidopsis Research. Plant Cell Advance Online Publication.

The Multinational Arabidopsis Steering Committee (MASC), as you might expect from their title, is an international group of Arabidopsis researchers who steer research in a productive direction. MASC reduces redundancy in research and encourages collaborations. Over the last 20 years, MASC has neatly guided the Arabidopsis community to achievements in genome sequencing, understanding of plant hormones, development of open access bioinformatics resources and much more. Now MASC has planned a roadmap for the next ten years of Arabidopsis research entitled From Bench to Bountiful Harvest.

The roadmap consists of five broad objectives: (more…)

New TAIR10-compatible CDF files and review of RNA labelling methods

by Charis Cook

Categories: Summary for non-specialists

Tags: agronomics, methods, microarray, summary for non-specialists, TAIR

Comments: No Comments

Published on: July 17, 2012

A current paper in Plant Methods assessed the pros and cons of two RNA labeling methods for AGRONOMICS1 tiling arrays, concluding that random priming is more suitable for organelle transcriptome analysis as it can label non-polyadenylated transcripts effectively. They also generated new TAIR-10 based CDF files, which can be used to re-analyse existing AGRANOMICS1 CEL files. The new CDFs can be accessed here.

First of all, the authors gave an overview of the AGRONOMICS1 tiling array. It contains all the probes from the traditionally used Affymetrix ATH1 array, but has additional probes which mean the AGRONOMICS1 array yields expression data for over 7000 more genes, around a third of the genome. 90% of annotated genes on the TAIR9 database are on the array. Mitochondrial and chloroplast genomes are completely represented, and sRNA, tRNA and miRNA can also be detected. The AGRONOMICS1 array has probes that represent both strands of the entire Arabidopsis genome, allowing epigenetic profiling. The quality is comparable to that of the ATH1 array.

Müller et al. compared the GeneChip© IVT express kit, an oligo-dT based RNA labeling technique, with the GeneChip© whole transcript (WT) Sense Target Labeling Assay which uses random hexamers tagged with T7 promotor sequences. Both kits are from Affymetrix, Santa Carla, CA. (more…)

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