eChapters

Summary

Researchers face a series of similar-yet unique problems to isolate proteins. To solve these they must have dip knowledge to select an appropriate technique. The mechanism, to study the way in which a living system works, is necessary to dismantle the machine and to isolate the component parts so that they may be studied, separately and in their interaction with other parts. The knowledge that is gained in this way may be put to practical use, for example, in the design of medicines, diagnostics, pesticides, or industrial processes. In every case a pure protein is desirable as impurities may either be misleading, dangerous or unproductive, respectively. Protein isolation is, therefore, a very common, almost central, procedure in biotechnology. The purpose of this chapter is thus to fill the beginners with techniques relevant to protein isolation, hopefully to improve their knowledge.

Abstract

The field of bioinformatics emerged as a tool to facilitate biological discoveries more than 10 years ago. The ability to capture, manage, process, analyze and interpret data became more important than ever. Bioinformatics in agricultural science has many complex aspects and problems that must be resolved due to the involvement of a wide variety of fields and subjects. Consequently, the establishment of interdisciplinary research areas is required, which should include collaboration among experimental and informatics researchers in different fields within industry, academia, and government. Thus, the Agricultural Bioinformatics Special Interest Group was established to promote interaction among researchers and provide an opportunity to report on bioinformatics studies in agricultural science: design and analysis of functional foods, scientific testing of food safety, breeding of agricultural organisms, development of environmental friendly agricultural chemicals, bioremediation and production of useful chemical materials using microorganisms, measuring and monitoring biodiversity, statistical genetics in resource ecology, and so on. The activities of the Agricultural Bioinformatics Special Interest Group include (1) Studies of applied bioinformatics in the agricultural science (2) Development of new research areas in agricultural science using bioinformatics (3) Practical application of bioinformatics technologies in the field of agricultural science (4) Development of new bioinformatics technologies characteristic of agricultural science.

Abstract

Proteins are complex molecules composed of one or more polypeptide chains made up of amino acid residues. These molecules perform a variety of essential functions in living organisms, including DNA replication, molecule transport, catalyzing metabolic processes, and providing structural support to cells. In this chapter, we explore the four levels of protein structure: primary, secondary, tertiary, and quaternary. Primary structure refers to the specific sequence of amino acids in a polypeptide chain, while secondary structure describes the folding of the chain into alpha-helices, beta-sheets, or other patterns. Tertiary structure is the three-dimensional shape of a single polypeptide chain, and quaternary structure refers to the interactions between multiple polypeptide chains to form a larger protein complex. Additionally, this chapter covers various methods used to predict protein structure, including homology modeling, ab initio prediction, and threading algorithms.

Introduction
World food security has become largely dependent only on a handful of crops like; wheat, rice, and corn which are the major cereals that support more than 50% of the global calorie demand. While these grains are an essential part of various diets, however they are devoid of essential micronutrients. As a result, it is estimated that micronutrient deficiency affects an estimated 2 billion people worldwide raising health concerns over our high dependence on cereal crops. As agriculture sector is considered as backbone of nation and therefore utilizing a handful of crops have placed the global food security at risk. In this context, Food and Agriculture Organization (FAO) has identified many plants as under-utilized, which can significantly contribute for improving nutrition and health, enhance food basket and livelihoods, future food security and sustainable development (FAOSTAT, 2023). These underutilized crops offer an immense potential in the functional food sector to combat hidden hunger crisis and offer the options of income generation.
Cereals such as rice, maize and wheat are monocots and belong to family Poaceae, however buckwheat, chia, quinoa and amaranthus although resembles with cereals are dicots and are categorized as Pseudocereals. These crops have been placed under the category of important crops by UNESCO due to dwindling cultivation and exploitation in the wild besides, these pseudocereals has gained a worldwide importance in the nutraceutical industry due to the rich nutritional profile compared to cereals and are considered as crops of the 21st century (Pirzadah et al., 2020). These are enriched with various active principals such as; polyphenols, flavonoids, amino acids, dietary fibre, lignans, vitamins, minerals, antioxidants, unsaturated fatty acids and other essential components like fagopyritols. The protein quality and quantity in pseudocereals is far better compared to cereals and this property qualifies their entry into the functional food industry (Rodriguez et al., 2020). Pseudocereals are rich in amino acids such as; arginine, tryptophan, lysine and histidine that proved essential for infant and child health thus, projected pseudocereals as an appropriate food supplement for child nutrition. The food production agro-systems must be adapted to climate change to ensure food security and stability. However, pseudocereals are climate resilient crops and can be grown even on marginal lands that are not fit for cereals thus, are considered as future crops to tackle mal-nutrition and future food crises. To explore these neglected or lost crops, there is an increasing interest in research and development that needs heightened direction and focus (Mabhaudhi et al., 2019). In this context, there is a need to adopt recent biotechnological interventions for the crop improvement and domestication of pseudocereal crops. This chapter focuses on nutritional profile of emerging pseudocereals viz., Buckwheat, Chia, Quinoa and Grain amaranth, phylogeny, genetic resources, breeding behavior and advanced breeding methods and their integration into the global food system

A line homozygous at all loci, ordinarily obtained by successive self-fertilizations in plant breeding is called a pure line. This concept was proposed by Danish biologist, W.L. Johannsen in 1903 while conducting a series of selection experiments for seed weight in common bean, which is strictly self-pollinated species. This is applied in self-pollinated crops, like wheat, rice, pulses, several vegetable crops, namely, tomato, chilli, eggplant, okra, cucurbits, etc. This is also applied to develop inbred parental lines in maize and several other cross-pollinated crops. However, ultimately these parental lines are used in development of new hybrids. This could be summarized as follows:

• Lines that are genetically different may be successfully isolated from within a population of mixed genetic types.
• Any variation that occurs within a pure line is not heritable but due to environmental factors only. Consequently, as Johansen’s bean study showed, further selection within the line is not effective.

Alinehomozygousatallloci, ordinarilyobtainedbysuccessive self-fertilization in plant breeding is known as pure-line. W. L. Johannsen (1903), a Danish biologist provided a sound scientific basis for selection in self-pollinated crops when he defined ‘pure-lines’ and described the genetic mechanism by which they are established through a series of experiments for seed weight selection in bean, variety Princess. Pure-lines in principle are supposed to be 100% homozygous but that hardly happens in practical plant breeding programmes and usually the so called pure-lines are ‘near pure-lines. Pure-lines tend to be uniform in the traits of interest. Genetic mutations, admixtures and limited out-crossing are the main causes of variation within pure-lines. This is a rapid and most in-expensive method. Pure-line theory may be summarized as follows:

QTL Traits

Polygenic inheritance, also known as quantitative or multifactorial inheritance refers to inheritance of a phenotypic characteristic (trait) that is attributable to two or more genes and their interaction with the environment. Unlike monogenic traits, polygenic traits do not follow patterns of Mendelian inheritance (qualitative traits). Instead, their phenotypes typically vary along a continuous gradient depicted by a bell curve. Though not necessarily genes themselves, quantitative trait loci (QTLs) are stretches of DNA that are closely linked to the genes that underlie the trait in question. QTLs can be molecularly identified (for example, with PCR or AFLP) to help map regions of the genome that contain genes involved in specifying a quantitative trait. This can be an early step in identifying and sequencing these genes.

Qualitative characters are those characters which show discontinuous or discrete differences and are counted into different classes and there is no need to measure them. Typically speaking, characters studied by Mendel in garden pea whose results were published in 1966 as Mendel’s Law were the typical characters that fitted into this class of characters in those days. To recall, these characters were (i) tall vs dwarf plants, (ii) round vs wrinkled seeds, (iii) yellow vs green pods, (iv) violet vs white flowers, (v) flat pods vs constricted pods, (vi), green pods vs yellow pods and (vii) axial flowers vs terminal flowers. Mendel postulated “Factors” responsible for these differences and he could explain his results of F₂ in ratio of 3:1 while dealing with one trait at a time and 9:3:3:1 while dealing with two traits simultaneously and proposed (i) law of segregation and (ii) principles of independent assortment to explain these ratios. Later on the term “factor” was called as gene.

1. Introduction:- Overview of Good Laboratory Practices (GLP)
1.1 Definition and Scope of GLP
GLP refers to a set of standards that govern the organizational processes, personnel responsibilities, and conditions under which laboratory studies are conducted. It aims to regulate research environments and ensure the generation of high-quality, traceable, and reproducible results. The scope of GLP encompasses a wide range of scientific fields including pharmaceuticals, agrochemicals, veterinary products, and environmental safety assessments. Good Laboratory Practices (GLP) regulations were developed by the Organization for Economic Co-operation and Development (OECD) and well adopted by regulatory agencies world-wide. The primary objective of GLP is to promote high standards of accuracy and integrity in non-clinical research to protect human health and the environment.
1.2 Principles of GLP
The fundamental principles of GLP include:
• Study Planning and Protocol Development: Every study must have a clear plan and defined objectives, which should be outlined in a comprehensive protocol.
• Personnel Responsibilities: The roles of study directors, scientists, quality assurance personnel, and support staff are clearly defined.
• Test Systems and Facilities: Laboratories must maintain appropriate facilities and control systems for conducting studies. These include properly calibrated equipment, controlled environments, and standardized procedures.
• Data Integrity and Reporting: Data must be meticulously recorded, verified, and archived, ensuring it is traceable and reproducible.
• Study Documentation and Archiving: All records, raw data, and reports must be stored and accessible for a period defined by regulations, ensuring transparency and reproducibility.

Seed in agriculture recognized as one of the most vital and crucial input in agriculture ever since the human kind has started tilling the land to produce the food for his existences. Seed technologist defined seed as matured fertilized ovule consisting of an intact embryo, endosperm and or cotyledon with protective covering (seed coat). A quality hybrid maize seed production system ensures design of the production technology which enables to maintain the healthy embryo, endosperm and /or cotyledon with protective covering. In fact the search of quality seed had acted as a major force to evolve the science of plant breeding. Ever since the farmers understood the importance of seed, they started trading the quality seed within community and now between geographies. Self-pollinated crops possess the natural mechanisms to maintain the intactness of genetic constitution of a seed during multiplication as there is very minimal chance of natural crossings. On the contrary, cross pollinated crops like maize requires special techniques in seed multiplication to maintain the genetic constitution of a F₁ hybrid. Hybrid Maize seed production techniques are well defined and are used by public and private sectors to produce the required hybrid seed in India. In this chapter, I will be highlighting the process, potential agro-climatic zones, hybrid maize seed production capacity and its future perspective in India. In addition, it also reviews the current system and possible strategic move in order to support the maize seed production system and steps required to meet future demand.

To Estimate the DNA Using Diphenylamine

Introduction

DNA estimation is helpful to know about the total amount of DNA of prokaryotic as well as eukaryotic system. One method used for DNA estimation is Diphenylamine assay, is that of Burton who modified the original procedure of Dische. This procedure is based on treatment of nucleic acid with hot perchloric acid to hydrolyze DNA nucleotides. Diphenylamine reacts specifically with deoxyribose in DNA to form a blue-colored complex. Since diphenylamine does not react with ribose, one can assay for DNA in the presence of contaminating RNA.

Principle

The DPA method is based on the colorimetry measurement of products formed by the reaction of ù- hydroxylevulinaldehyde (from the deoxyribose released after depurination of DNA) with diphenylamine in 2.5 N perchloric acid.

When DNA is treated with diphenylamine under acidic condition (PCA) a blue complex is formed with a sharp absorption maximum at 595 nm (600 nm). In acid solution, the straight chain form of the deoxypentose is converted to highly reactive ω – hydroxylevulinaldehyde that reacts with diphenylamine to give blue complex. The reaction yields two mixture of product which together have absorbance maximum at 595 nm (600 nm). Interferences include proteins, lipids, saccharides and RNA. In DNA the only deoxyribose of the purine nucleotides reacts, so that the value obtained represents half of the total deoxyribose present.

Quantitative Traits

• Quantitative traits exhibit a continuum of phenotypic variation that may follow a normal distribution
• Statistical methods are used to evaluate a frequency distribution quantitatively
• Some statistical methods compare two variables with each other

Polygenic Inheritance

Polygenic inheritance and environmental factors create overlaps between genotypes and phenotypes

Polygenic inheritance explains DDT-resistance in Drosophila

Quantitative trait loci (QTLs) are now mapped by linkage to molecular markers

15.1. Cereals

15.1.1. Oat (Avena sativa)

A. Multiple Choice Questions

1. While studying the history of domestication of various cultivated plants were recognized earlier:
   1. Centers of origin
   2. Centers of domestication
   3. Centres of hybrid
   4. Centres of variation
2. The quickest method of plant breeding is:
   1. Mutation breeding
   2. Hybridization
   3. Selection
   4. Introduction

Reagents

Restriction enzymes of choice, such as BamH1 and EcoRI

Restriction enzyme reaction buffer,

70 % Ethanol

100 % Ethanol

3 M Sodium acetate (pH 5.2)

Distilled water

Plasmid DNA isolated from bacteria, such as pGFP(R) and pBC(R)

An atom consists of a positively charged nucleus surrounded by a cloud of negatively charged electrons. The positive nuclear charge is due to the presence of protons which carry a mass of 1 and a charge of +1. Besides protons in the nucleus, there are neutrons which are heavier than protons. They carry no charge. Comparatively electrons have no mass (1836 times lesser than that of a proton) but they carry a charge of –1. Hence, the whole mass of an atom is likely to be concentrated in the nucleus.

The Ramayana written by sage Valmiki is one of the great epics of the Sanskrit language. It is most religiously regarded and popular book in Hinduism. The book is believed to be complied between 300 B.C.E. – 200 AD. However, the original story could be as old 500 B.C.E. (Goldman, 1984). There are many other versions of the epic in various regional languages found in India and South Asia (Raghunathan, 1981; Arya,1998; Dutt, 2004; Mukherjee, 2015; Sharma and Arya, 2022).

As per literature references, the name ‘Ramanaya’ is composed of two words, ‘Rama’ and ‘Aya?a’. Rama, the name of the main character of the Epic, has two contextual meanings. In the Atharvaveda, it means 'dark’, dark-coloured, ‘black' and is related to the word ‘ratri’ (night) which means 'darkness of night'. The other meaning, which can be found in the Mahabharata, is 'pleasing’, pleasant, charming, lovely, beautiful. The word ‘Ayana’ means travel or journey. Thus, Ramaya?a means ‘Rama's Journey’ from his kingdom to various places, forests during his life time (Anonymous, 1960; Tulsidas, 1966; Murthy, 2003; Kishore, 2007; Rajagopalachari, 2013).

Ramayana contains over 24,000 couplet verses, in its full form, which are divided into following seven ‘Ka??as’ (booklets) - Balaka??a, Ayodhyaka??a, Ara?yaka??a, Ki?kindhaka??a, Sundaraka??a, Yuddhaka??a, Uttaraka??a and about 500 sargas (chapters). It is regarded as one of the longest epic poems written in history (Keshavadas, 1988; Debroy, 2017; Gupta, 2021).

morally right man in the world? Narada replied that such a man is Rama, son of King Dasharatha of Ayodhya. Valmiki wanted to know more about Rama and proceeded to take bath when he saw two birds in playful mood and was happy. Suddenly, a hunter hiding in the bush shot one of the two birds. Valmiki became painful and chanted some dialogues in curse which ultimately created a new form of sloka. This was the beginning of the main narrative (Goldman, 1984; Sachithanantham, 2004).

RAPD stands for Random Amplification of Polymorphic DNA. It is a type of PCR reaction, but the segments of DNA that are amplified are random. The scientist performing RAPD creates several arbitrary, short primers (8-12 nucleotides), then proceeds with the PCR using a large template of genomic DNA, hoping that fragments will amplify. By resolving the resulting patterns, a semi-unique profile can be gleaned from a RAPD reaction.

No knowledge of the DNA sequence for the targeted gene is required, as the primers will bind somewhere in the sequence, but it is not certain exactly where. This makes the method popular for comparing the DNA of biological systems that have not had the attention of the scientific community, or in a system in which relatively few DNA sequences are compared (it is not suitable for forming a DNA databank). Because it relies on a large, intact DNA template sequence, it has some limitations in the use of degraded DNA samples. Its resolving power is much lower than targeted, species specific DNA comparison methods, such as short tandem repeats. In recent years, RAPD is used to characterize, and trace, the phylogeny of diverse plant and animal species.

RAPD stands for Random Amplification of Polymorphic DNA. It is a type of PCR reaction, but the segments of DNA that are amplified are random. The scientist performing RAPD creates several arbitrary, short primers (8-12 nucleotides), then proceeds with the PCR using a large template of genomic DNA, hoping that fragments will amplify. By resolving the resulting patterns, a semi-unique profile can be gleaned from a RAPD reaction.

No knowledge of the DNA sequence for the targeted gene is required, as the primers will bind somewhere in the sequence, but it is not certain exactly where. This makes the method popular for comparing the DNA of biological systems that have not had the attention of the scientific community, or in a system in which relatively few DNA sequences are compared (it is not suitable for forming a DNA databank). Because it relies on a large, intact DNA template sequence, it has some limitations in the use of degraded DNA samples. Its resolving power is much lower than targeted, species specific DNA comparison methods, such as short tandem repeats. In recent years, RAPD is used to characterize, and trace, the phylogeny of diverse plant and animal species.