Hybrid Control Systems: a Design Case Study

This paper presents a modification to UML to improve the modelling and analysis of discrete-event dynamic system (DEDS) representations of manufacturing systems. It shows how Petri nets can be used to improve the representation and analysis of the dynamic model of a system specified using UML. Finally the technique is illustrated by its application to a simplified production line

γ-Zeins are essential for endosperm modification in quality protein maize

Essential amino acids like lysine and tryptophan are deficient in corn meal because of the abundance of zein storage proteins that lack these amino acids. A naturalmutant, opaque 2 (o2) causes reduction of zeins,anincreaseofnonzeinproteins,andas a consequence, adoubling of lysine levels.However, o2’s soft inferior kernels precluded its commercial use. Breeders subsequently overcame kernel softness, selectingseveral quantitative loci (QTLs), called o2modifiers,without losing the high-lysine trait. These maize lines are known as “quality protein maize” (QPM). One of the QTLs is linked to the 27-kDa γ-zein locus on chromosome 7S. Moreover, QPM lines have 2- to 3-fold higher levels of the 27-kDa γ-zein, but the physiological significance of this increase is not known. Because the 27- and 16-kDa γ-zein genes are highly conserved in DNA sequence, we introduced a dominant RNAi transgene into a QPM line (CM105Mo2) to eliminate expression of them both. Elimination of γ-zeins disrupts endosperm modification by o2 modifiers, indicating their hypostatic action to γ-zeins. Abnormalities in protein body structure and their interaction with starch granules in the F1 with Mo2/+; o2/o2; γRNAi/+ genotype suggests that γ-zeins are essential for restoring protein body density and starch grain interaction in QPM. To eliminate pleiotropic effects caused by o2, the 22-kDa α-zein, γ-zein, and β-zein RNAis were stacked, resulting in protein bodies forming as honeycomb-like structures. We are unique in presenting clear demonstration that γ-zeins play a mechanistic role in QPM, providing a previously unexplored rationale for molecular breeding

An Exome-seq Based Tool for Mapping and Selection of Candidate Genes in Maize Deletion Mutants

Despite the large number of genomic and transcriptomic resources in maize, there is still much to learn about the function of genes in developmental and biochemical processes. Some maize mutants that were generated by gamma-irradiation showed clear segregation for the kernel phenotypes in B73 X Mo17 F2 ears. To better understand the functional genomics of kernel development, we developed a mapping and gene identification pipeline, bulked segregant exome sequencing (BSEx-seq), to map mutants with kernel phenotypes including opaque endosperm and reduced kernel size. BSEx-seq generates and compares the sequence of the exon fraction from mutant and normal plant F2 DNA pools. The comparison can derive mapping peaks, identify deletions within the mapping peak, and suggest candidate genes within the deleted regions. We then used the public kernel-specific expression data to narrow down the list of candidate genes/mutations and identified deletions ranging from several kb to more than 1 Mb. A full deletion allele of the Opaque-2 gene was identified in mutant 531, which occurs within a ~200-kb deletion. Opaque mutant 1486 has a 6248-bp deletion in the mapping interval containing two candidate genes encoding RNA-directed DNA methylation 4 (RdDM4) and AMP-binding protein, respectively. This study demonstrates the efficiency and cost-effectiveness of BSEx-seq for causal mutation mapping and candidate gene selection, providing a new option in mapping-by-sequencing for maize functional genomics studies

The megaton mission : the role of the nuclear arms race in 1950s British imperialism

In August 1945, the Pacific theatre of the Second World War came to a dramatic close with the United States dropped two atomic bombs on Hiroshima and Nagasaki. Following this, tensions between the United States and the Soviet Union grew tremendously and the two superpowers set about developing even more destructive bombs in case these tensions reached a breaking point. The nuclear bomb came to symbolise power in the post-war era. However, they were not the only powers with nuclear programmes in the 1950s. Churchill, and successive Conservative governments, were keen in joining the nuclear arms race. Operation Grapple was conducted secretly from 1954-58 on Christmas Island, then part of the Gilbert and Ellice Islands in the Southern Pacific. The Gilbert and Ellice Islands were a Crown Colony at the time, and this paper shall examine the connection between the nuclear experiments and the British Empire, and geopolitical power by extension. This thesis is divided into three thematic chapters. The first chapter explores how space and place shaped aspirations for power and vice versa. Not only did Operation Grapple bring about colossal environmental damage but major demographic changes with both the arrival of the British and Fijian military personnel as well as a new transient population who brought the resources necessary to sustain such a population. As well as the functions of the different islands being altered, they came to symbolise the long-term health issues which affected the Fijian veterans long after the experiments were over. The second chapter examines how colonial rule in the Gilbert and Ellice islands was shaped by the nuclear experiments. It explores the rapid change in infrastructure needed to accommodate the new population and the friction it caused when the issue of financial responsibility was raised. This chapter also explores how colonial rule became increasingly reliant on the military and how the developing military complex at Britain extended to remote colonies. Finally, the limits of ‘East of Suez’ and various governmental departments’ visions for the future are discussed as the South Pacific was useful for further developments of nuclear weaponry. The third and final chapter gauges Britain’s place in the world through their relationships with other nations. This is achieved by examining how Britain interacted with the United States, France, Japan, Australia, New Zealand, and Fiji. It concludes that Britain was in a weaker position to negotiate with other nations but were still able to wield some power in a softer way, despite their label as a ‘superpower’ being called into question. This thesis shall reach the overall conclusion that the experiments elevated the status of the islands in the minds of the British government to one of high importance as well as offering the British the means of maintaining geopolitical relevance as well as recalibrating attention to the South Pacific, albeit briefly

Identification and characterization of the maize arogenate dehydrogenase gene family

In plants, the amino acids tyrosine and phenylalanine are synthesized from arogenate by arogenate dehydrogenase and arogenate dehydratase, respectively, with the relative flux to each being tightly controlled. Here the characterization of a maize opaque endosperm mutant (mto140), which also shows retarded vegetative growth, is described The opaque phenotype co-segregates with a Mutator transposon insertion in an arogenate dehydrogenase gene (zmAroDH-1) and this led to the characterization of the four-member family of maize arogenate dehydrogenase genes (zmAroDH-1–zmAroDH-4) which share highly similar sequences. A Mutator insertion at an equivalent position in AroDH-3, the most closely related family member to AroDH-1, is also associated with opaque endosperm and stunted vegetative growth phenotypes. Overlapping but differential expression patterns as well as subtle mutant effects on the accumulation of tyrosine and phenylalanine in endosperm, embryo, and leaf tissues suggest that the functional redundancy of this gene family provides metabolic plasticity for the synthesis of these important amino acids. mto140/arodh-1 seeds shows a general reduction in zein storage protein accumulation and an elevated lysine phenotype typical of other opaque endosperm mutants, but it is distinct because it does not result from quantitative or qualitative defects in the accumulation of specific zeins but rather from a disruption in amino acid biosynthesis

Generation and Evaluation of Modified \u3ci\u3eOpaque\u3c/i\u3e-2 Popcorn Suggests a Route to Quality Protein Popcorn

Introducing traits from dent corn to popcorn is challenging because it is difficult to recover adequate popping characteristics. QPM (Quality Protein Maize) is a dent corn variety carrying the opaque-2 (o2) mutation, specifying increased amounts of normally limiting essential amino acids, and modifier genes which restore the wild type vitreous kernel phenotype. In this study, we introgressed o2 and selected for endosperm modification using vitreousness and high 27-kD gamma zein content. In this way, we recovered high-lysine, fully poppable Quality Protein Popcorn (QPP). BC2F4 individuals with vitreous kernels were confirmed to be o2 mutants by both genotyping and SDSPAGE. Amino acid profiling of BC2F4 individuals showed that they all have significantly increased lysine compared with popcorn parental lines. Principal Component Analysis of the amino acid profiles showed that all introgressions were grouped with corresponding QPM parental lines. Popping analysis of the BC2F5 individuals showed that while there is variability in popping volume between lines, some lines show equivalent popping to the popcorn parent. In this proof-of-concept study for QPP, we have shown that it is possible to rapidly recover sufficient popcorn characteristics in a modified o2 background using simple phenotypic, biochemical and genetic selection. Furthermore, this shows increased y-zein is an acceptable substitute for a-zein for full poppability. Since we have developed multiple QPP introgressions, this gives good scope for ongoing hybrid production and future evaluation of agronomic performance and selection of elite hybrids. In a wider context, this study shows the potential for breeding beneficial traits into popcorn for agronomic improvement

Generation and Evaluation of Modified Opaque-2 Popcorn Suggests a Route to Quality Protein Popcorn

Introducing traits from dent corn to popcorn is challenging because it is difficult to recover adequate popping characteristics. QPM (Quality Protein Maize) is a dent corn variety carrying the opaque-2 (o2) mutation, specifying increased amounts of normally limiting essential amino acids, and modifier genes which restore the wild type vitreous kernel phenotype. In this study, we introgressed o2 and selected for endosperm modification using vitreousness and high 27-kD gamma zein content. In this way, we recovered high-lysine, fully poppable Quality Protein Popcorn (QPP). BC2F4 individuals with vitreous kernels were confirmed to be o2 mutants by both genotyping and SDS-PAGE. Amino acid profiling of BC2F4 individuals showed that they all have significantly increased lysine compared with popcorn parental lines. Principal Component Analysis of the amino acid profiles showed that all introgressions were grouped with corresponding QPM parental lines. Popping analysis of the BC2F5 individuals showed that while there is variability in popping volume between lines, some lines show equivalent popping to the popcorn parent. In this proof-of-concept study for QPP, we have shown that it is possible to rapidly recover sufficient popcorn characteristics in a modified o2 background using simple phenotypic, biochemical and genetic selection. Furthermore, this shows increased γ-zein is an acceptable substitute for α-zein for full poppability. Since we have developed multiple QPP introgressions, this gives good scope for ongoing hybrid production and future evaluation of agronomic performance and selection of elite hybrids. In a wider context, this study shows the potential for breeding beneficial traits into popcorn for agronomic improvement

A population of deletion mutants and an integrated mapping and exome-seq pipeline for gene discovery in maize

To better understand maize endosperm filling and maturation, we used gamma irradiation of the B73 maize reference line to generate mutants with opaque endosperm and reduced kernel fill phenotypes, and created a population of 1788 lines including 47 Mo17 × F2s showing stable, segregating and viable kernel phenotypes. For molecular characterization of the mutants, we developed a novel functional genomics platform that combined Bulked Segregant RNA and Exome sequencing (BSREx-seq) to map causative mutations and identify candidate genes within mapping intervals. To exemplify the utility of the mutants and provide proof-ofconcept for the bioinformatics platform, we present detailed characterization of line 937; an opaque mutant harboring a 6203-bp in-frame deletion covering six exons within the Opaque-1 gene. In addition, we describe mutant line 146 which contains a 4.8 Kb intra-gene deletion within the Sugary-1 gene and line 916 in which an 8.6 Kb deletion knocks out a Cyclin A2 gene. The publically available algorithm developed in this work improves the identification of causative deletions and its corresponding gaps within mapping peaks. This study demonstrates the utility of gamma irradiation for forward genetics in large non-dense genomes such as maize since deletions often affect single genes. Furthermore, we show how this classical mutagenesis method becomes applicable for functional genomics when combined with state-of-the-art genomics tools

A population of deletion mutants and an integrated mapping and exome-seq pipeline for gene discovery in maize

To better understand maize endosperm filling and maturation, we used gamma irradiation of the B73 maize reference line to generate mutants with opaque endosperm and reduced kernel fill phenotypes, and created a population of 1788 lines including 47 Mo17 × F2s showing stable, segregating and viable kernel phenotypes. For molecular characterization of the mutants, we developed a novel functional genomics platform that combined Bulked Segregant RNA and Exome sequencing (BSREx-seq) to map causative mutations and identify candidate genes within mapping intervals. To exemplify the utility of the mutants and provide proof-ofconcept for the bioinformatics platform, we present detailed characterization of line 937; an opaque mutant harboring a 6203-bp in-frame deletion covering six exons within the Opaque-1 gene. In addition, we describe mutant line 146 which contains a 4.8 Kb intra-gene deletion within the Sugary-1 gene and line 916 in which an 8.6 Kb deletion knocks out a Cyclin A2 gene. The publically available algorithm developed in this work improves the identification of causative deletions and its corresponding gaps within mapping peaks. This study demonstrates the utility of gamma irradiation for forward genetics in large non-dense genomes such as maize since deletions often affect single genes. Furthermore, we show how this classical mutagenesis method becomes applicable for functional genomics when combined with state-of-the-art genomics tools

Validation of QTL mapping and transcriptome profiling for identification of candidate genes associated with nitrogen stress tolerance in sorghum

Background: Quantitative trait loci (QTLs) detected in one mapping population may not be detected in other mapping populations at all the time. Therefore, before being used for marker assisted breeding, QTLs need to be validated in different environments and/or genetic backgrounds to rule out statistical anomalies. In this regard, we mapped the QTLs controlling various agronomic traits in a recombinant inbred line (RIL) population in response to Nitrogen (N) stress and validated these with the reported QTLs in our earlier study to find the stable and consistent QTLs across populations. Also, with Illumina RNA-sequencing we checked the differential expression of gene (DEG) transcripts between parents and pools of RILs with high and low nitrogen use efficiency (NUE) and overlaid these DEGs on to the common validated QTLs to find candidate genes associated with N-stress tolerance in sorghum. Results: An F7 RIL population derived from a cross between CK60 (N-stress sensitive) and San Chi San (N-stress tolerant) inbred sorghum lines was used to map QTLs for 11 agronomic traits tested under different N-levels. Composite interval mapping analysis detected a total of 32 QTLs for 11 agronomic traits. Validation of these QTLs revealed that of the detected, nine QTLs from this population were consistent with the reported QTLs in earlier study using CK60/China17 RIL population. The validated QTLs were located on chromosomes 1, 6, 7, 8, and 9. In addition, root transcriptomic profiling detected 55 and 20 differentially expressed gene (DEG) transcripts between parents and pools of RILs with high and low NUE respectively. Also, overlay of these DEG transcripts on to the validated QTLs found candidate genes transcripts for NUE and also showed the expected differential expression. For example, DEG transcripts encoding Lysine histidine transporter 1 (LHT1) had abundant expression in San Chi San and the tolerant RIL pool, whereas DEG transcripts encoding seed storage albumin, transcription factor IIIC (TFIIIC) and dwarfing gene (DW2) encoding multidrug resistance-associated protein-9 homolog showed abundant expression in CK60 parent, similar to earlier study. Conclusions: The validated QTLs among different mapping populations would be the most reliable and stable QTLs across germplasm. The DEG transcripts found in the validated QTL regions will serve as future candidate genes for enhancing NUE in sorghum using molecular approaches