Efficient High-throughput Techniques for the Analysis of Disease-Resistant Plant Varieties and Detection of Food Adulteration

Over the past two decades, the advances in the next generation sequencing (NGS) platforms have led to the identification of numerous genes/QTLs at high-resolution for their potential use in crop improvement. The genomic resources generated through these high-throughput sequencing techniques have been efficiently used in screening of particular gene of interest particularly for numerous types of plant stresses and quality traits. Subsequently, the identified-markers linked to a particular trait have been used in marker-assisted backcross breeding (MABB) activities. Besides, these markers are also being used to catalogue the food crops for detection of adulteration to improve the quality of food. With the advancement of technologies, the genomic resources are originating with new markers; however, to use these markers efficiently in crop breeding, high-throughput techniques (HTT) such as multiplex PCR and capillary electrophoresis (CE) can be exploited. Robustness, ease of operation, good reproducibility and low cost are the main advantages of multiplex PCR and CE. The CE is capable of separating and characterizing proteins with simplicity, speed and small sample requirements. Keeping in view the availability of vast data generated through NGS techniques and development of numerous markers, there is a need to use these resources efficiently in crop improvement programmes. In summary, this review describes the use of molecular markers in the screening of resistance genes in breeding programmes and detection of adulterations in food crops using high-throughput techniques.

An overview of databases and bioinformatics tools for plant antimicrobial peptides

: Antimicrobial peptides (AMPs) are small, ribosomally synthesized proteins found in nearly all forms of life. In plants, AMPs play a central role in plant defense due to their distinct physicochemical properties. Due to their broad-spectrum antimicrobial activity and rapid killing action, plant AMPs have become important candidates for the development of new drugs to control plant and animal pathogens that are resistant to multiple drugs. Further research is required to explore the potential uses of these natural compounds. Computational strategies have been increasingly used to understand key aspects of antimicrobial peptides. These strategies will help to minimize the time and cost of "wet-lab" experimentation. Researchers have developed various tools and databases to provide updated information on AMPs. However, despite the increased availability of antimicrobial peptide resources in biological databases, finding AMPs from plants can still be a difficult task. The number of plant AMP sequences in current databases is still small and yet often redundant. To facilitate further characterization of plant AMPs, we have summarized information on the location, distribution, and annotations of plant AMPs available in the most relevant databases for AMPs research. We also mapped and categorized the bioinformatics tools available in these databases. We expect that this will allow researchers to advance in the discovery and development of new plant AMPs with potent biological properties. We hope to provide insights to further expand the application of AMPs in the fields of biotechnology, pharmacy, and agriculture.

Overview of Enzyme Engineering Towards the Production of Hyaluronic Acid with Tailored Molecular Weight

: Hyaluronic acid or hyaluronan (HA) is a natural biopolymer composed of D-glucuronic acid and N-acetylglucosamine units, distributed as a non-sulfated and anionic glycosaminoglycan in important tissues of the body, and is commercially and biologically important. Its biological properties are determined by the molecular weight and dispersity which are suitable for particular medical and cosmetic applications. The synthesis of well-defined and monodisperse HA is still a significant obstacle and an impressive research field for advanced medical applications. High polydispersity by bacterial fermentation, the lack of knowledge of the mechanism required to start and continue the synthesis process, increased cost of raw materials to produce HA, clarification and explanation of factors limiting synthesis in bacterial systems are among the important challenges of hyaluronic acid synthesis. Hyaluronan synthase plays a critical role in HA molecular mass by producing a wide range of HA involved in various biological processes. Hyaluronan biosynthesis has been considered extensively; however, the control of its size and weight during the synthesis process is poorly investigated. This review focuses on these uncharted biochemical details to obtain the uniform chain lengths of Hyaluronan by protein engineering and regulating the function of Hyaluronan synthase.

Measurement of high protein concentrations by optical rotation: a case study for monitoring of monoclonal antibody drug downstream processes

Background: Recent advancements in cell engineering and bioreactor engineering have enabled high monoclonal antibody (mAb) concentrations in harvested solutions for the downstream process (DSP). Methods: As many unit operations such as capture chromatography, polish chromatography, membrane filtration, virus inactivation, virus filtration, and concentration by ultrafiltration are involved in DSP, it is crucial to monitor the process carefully in order to perform reliable and stable DSP operations. One of the most important signals (process parameter) to be monitored is the protein concentration CP. Although various methods are available, most of them are not suited for measuring high CP. In this paper, we have developed a method for measuring very high CP by optical rotation (OR). Result: Linear correlations were confirmed between OR and CP in the range CP = 0 to 80 g/L for mAbs with high repeatability and small variation coefficients. This method was applied to the monitoring of CP in the opaque (colored) solution during the cell culture. The CP by OR was in good agreement with those by the standard Protein A HPLC method. Conclusion: Monitoring of high CP by OR is expected to be an efficient process analytical tool (PAT) for DSP.

Advanced protein drugs and formulations

: For a long time, proteins were a subset of molecules rarely applied as therapeutically active molecules. Some of the first applications of proteins as drugs have been insulin and vaccines for supply a physiological deficiency and the prevention of diseases, respectively. Nowadays, proteins have increased their range of application, not only as drugs but also as drug delivery systems to be administered by different routes. Due to their nature, proteins show different behavior while the conditions of the environment are modified. For this reason, it has been necessary to study their behavior for predicting the correct manufacturing, storing, or combination with other possible molecules in a formulation or into the body. The application of techniques for predicting the behavior of proteins in different environments has led to associate this type of behavior into the body with the occurrence of diseases such as celiac disease or Alzheimer's disease. Thus, this work shows an overview of the main types of proteins applied as active therapeutically molecules, proteins-based drug delivery systems, and techniques for predicting their stability into the storing container and the body.

Soybean β-conglycinin: structure characteristic, allergenicity, plasma lipid-controlling, prevention of obesity and non-alcoholic fatty liver disease

: Soybean has become an important world commodity because of its low price, good nutritional value and recognized functional health benefits in recent years. β-conglycinin is one of the major storage proteins in soybean. It has captured a growing interest recently because of its allergenicity and potential health benefits, which continues to drive the research and commercial development of β-conglycinin-based food products and ingredients. In this review, the structure, the amino acid composition, extraction methods and electrophoretic pattern of β-conglycinin are briefly summarized. Studies on β-conglycinin by allergenicity, plasma lipid-controlling, obesity and non-alcoholic fatty liver disease are highlighted, critically discussing their main shortcomings and challenges and identifying the research gaps. Studies to date have demonstrated the cultivation of β-conglycinin with health benefits as functional ingredients and foodstuffs. The current research focuses on proteins, mainly challenging the mechanisms of subunit/peptide effects interaction and identifying and characterizing the hidden biological activities in the polypeptide chains. There is much scope for further exploration into various aspects of β-conglycinin, such as the selection of mutant strains and genetic engineering and prospects on targeted β-conglycinin exploitation in the nutraceutical area. In addition, the safety evaluation of β-conglycinin and its stabilized emulsions deserve more attention to food-related applications.

Novel Sequence Features of Leucine-rich Repeats in Proteins from Nucleo-cytoplasmic Large DNA Viruses

: Leucine-rich repeats (LRRs) occurring in tandem are 20-29 amino acids long. Eleven LRR types have been recognized. Sequence features of LRRs from viruses were investigated using over 600 LRR proteins from 89 species. Before, metagenome data of nucleo-cytoplasmic large dsDNA viruses (NCLDVs) have been published; the 2,074 NCLDVs encode 199,021 proteins. From the NCLDVs, 549 LRR proteins were identified and analyzed. A comprehensive analysis of TpLRR and FNIP that belong to an LRR class was first performed. The repeating unit lengths (RULs) in five types are 19 residues, which are the shortest among all LRRs. Some RULs are one to five residues shorter than those of the known, corresponding LRR types. The shrinking of RUL is also observed in FNIP. The conserved hydrophobic residues, such as Leu, Val or Ile, in the consensus sequences are frequently substituted by cysteine at one or two positions. Some unique LRR types that are different from those identified previously have been observed. The present study confirms the previous result that the sequence novelty is a general feature of viral LRR proteins.