eChapters

DNA can be easily isolated and purified after size selection on an agarose gel. The fragment of interest is simply cut out of the gel with a razor blade and purified by a number of different methods.The easiest is to use a method that involves first dissolving the agarose slice in a solution at 50°C, then binding the DNA from the melted agarose to a silica-gel membrane.

Principle

In this method following gel electrophoresis using low gelling/ melting temperature agarose, the band of interest is removed from the gel. The agarose slice is then melted and subjected to phenol extraction.

here are many methods for extracting DNA from gel apart from using commercial kits, some methods however yield cleaner DNA than others.

1. Freeze squeeze

Excise band and place pieces of it in a home-made spin-column (a piece of filter placed in a PCR 0.5ml eppendorf, pierced the bottom of the PCR tube with a needle). Place the PCR tube at -20 °C for 15-20 mins or freeze in liquid N2. Then place the PCR tube into the 1.5 ml eppendorf and spin at top speed in a microfuge for 15 mins, collect DNA solution which can be EtOH ppt or use directly.

2. Powdered silica

Melt excised bands at 70°C cool and add equal volume of TE-buffered phenol and mix. Spin for 5 min, remove aqueous phase, repeat and EtOh ppt. the DNA.

3. Electroelution

Nanomaterials are a rapidly growing technological sector with diverse applications. These nanomaterials are categorized into distinct groups according to their shape, size, and materials used in their fabrication. The synthesis of nanomaterials follows different strategies that are tailored to their specific applications. This chapter describes the different approaches used for the synthesis of nanomaterials, including top-down, bottom-up, and hybrid approaches. Furthermore, this chapter discusses the advantages and disadvantages of each fabrication method and explores the diverse applications of nanomaterials across multiple fields.

Factors which influence the colonization and life cycle of VAM fungi have important bearing on the extent to which they may benefit their host. As we move closer to using mycorrhizal fungi to increase growth and productivity of different plant communities, a thorough understanding of the factors which influence their survival, germination, root colonization an d sporulation will be critical. Seasons, physico-chemical characters of the soil, agronomic practices, plant protection chemicals, pollutants and genotypic characters of the host may affect VAM fungi and their potentiality to colonize and provide benefit to the host. In view of this, the effect of such factors on the VA mycorrhizal symbiosis and its benefit to diverse plant communities has been extensively investigated worldover.

Start with a 1:2 molar ratio of vector: insert DNA when cloning a fragment into a plasmid vector. This ratio will vary with other types of vectors, for example, cDNA and genomic cloning vectors. The following example illustrates the conversion of molar ratios to mass ratios for a 3.0kb plasmid and a 0.5kb insert DNA fragment.

Introduction
The integration of bioinformatics into plant breeding has been shaped by several key milestones over the past few decades. Concepts like Breeding 4.0 (Wallace et al.,2018) and, 5G Breeding (Varshney et al., 2020) have gained global prominence, strengthened by the availability of extensive omics data. The advent of DNA sequencing technologies marked a key milestone in this field, catalyzing numerous genome sequencing projects worldwide such as Arabidopsis, rice, tomato, maize, wheat, etc. Next-generation sequencing (NGS) platforms, such as Illumina, PacBio, and Oxford Nanopore, have enabled rapid sequencing of entire plant genomes, facilitating the identification of genetic variations, including single nucleotide polymorphisms (SNPs), insertions, deletions, and structural variants (Deschamps & Campbell, 2010; Fuentes-Pardo et al.,2017). These advancements have significantly improved comparative genomics, enabling researchers to compare the genetic makeup of different plant species or varieties and identify genes associated with agronomic traits, such as yield, disease resistance, and environmental stress tolerance (Khan et al., 2024). As a result, these developments have revolutionized plant genetic research and crop improvement strategies, providing unprecedented opportunities to enhance agricultural productivity (Bellare et al., 2018; Shendure and Ji, 2008).
Bioinformatics has revolutionized plant biology by combining computational and statistical tools to analyze genomes, transcriptomes, proteomes, and metabolomes facilitating insights into genome structure, function, and evolution (Yadav et al., 2015; Saha et al., 2023; Roychowdhury et al., 2023). Genomics, emerged in the 1980s primarily focussing on mapping and sequencing genomes and there after laid the groundwork for functional genomics, which systematically explored gene functions (Hieter and Boguski, 1997; (Jackson et al., 2011). Further, advances in sequencing technologies, such as long-read platforms, have enabled detailed analyses of structural variants in crop genomes, crucial for identifying agronomic traits.
The large volumes of genomic sequence data have allowed researchers to identify genetic markers for crop improvement and candidate genes for trait development. However, the full potential of these genomic sequencing efforts depends on assigning functions to the thousands of genes with unknown roles. Model organisms such as humans, mice, Arabidopsis, rice, tomato, Medicago, and wheat have been instrumental in advancing genomic investigations (Bult, 2006). A notable example is the model organism Arabidopsis thaliana, which has provided a foundation for exploring fundamental biological phenomena using resources such as the TAIR database (Lamesch et al., 2012). Leveraging advanced bioinformatics techniques, researchers have enhanced crop productivity, mitigated diseases, and optimized resource management in agriculture. Advanced approaches like genome-wide association studies (GWAS) and transcriptome analysis continue to refine the understanding of genetic underpinnings, revolutionizing modern agricultural strategies (Mai et al., 2023).

Abstract

FASTA, which stands for “Fast All”, is a sequence alignment tool developed by William R. Pearson and David J. Lipman in 1985. It is commonly used to identify similarities between DNA and protein sequences. FASTA employs a “hashing” strategy to locate matches for short stretches of identical residues with a length of k, referred to as k-tuples or k-tups. This method enables FASTA to quickly search large databases for sequence similarities, making it a popular tool for identifying homologous sequences, predicting gene functions, and analyzing evolutionary relationships. This chapter elaborates some basics of FASTA.

9.1 Introduction

Nanoparticles (NPs) are particulate matter with at least one dimension less than 100 nm (Christian et al., 2008). Although NPs are naturally present in the environment, manufactured NPs may have distinctive surface properties and chemistry in comparison with natural NPs (Handy et al., 2008). Manufactured NPs have a large area-to-volume ratio and size-dependent properties, which impart novel properties and behaviors that make them suitable for nanotechnology applications (Auffan et al., 2009). Range of applications for NPs has brought about a rapid development of the nanotechnology industry.

Plant breeding activities basically involve development of new and superior crop varieties to replace the old ones and this naturally needs comparison of several lines/cultivars for agronomically important traits which are often quantitative in nature. Therefore, lots of environmental influences are there on the mean performance and mere numerical comparison of mean data is not sufficient to draw valid conclusions regarding the superiority of one variety over the other. Great advances have been made during the last about 100 years in the area of experimental designs and analytical procedures to facilitate such comparisons. Most commonly used designs by the plant breeders are included here for better understanding by the students and the breeders and to handle the experimental data on their own when confronted with such issues. Before applying statistical procedures on the experimental designs, it must be well understood that proper use of field plot technique is of great value to have meaningful data for further statistical analysis and use.

Minimum field standards for seed production have been prescribed  for each crop. The major components to be looked into for meeting the field standards are: 
   Preceding crops 
   Isolation distance 
   Number of field inspections 

The minimum proceding crop requirement has been specified to minimize genetic contamination on account of volunteer plants germinating from the left over seeds in the previous season. The disease contamination is also possible from soil borne disease organisms if the previous crop was not certified. The preceding crop requirement would mean that for a prescribed minimum period the same crop or any other contaminating crop should not have been grown on the same field where seed production is to be taken up unless the preceding crop was a seed crop and had met the field/the seed standards.

13.1 Biosafety Research Trials: Scope and Implementation
Introduction
Biosafety research trials serve as a critical phase in the evaluation and regulation of genetically modified organisms (GMOs), particularly transgenic crops and microorganisms. These trials assess potential environmental and health impacts under confined and controlled conditions before the organism can be considered for commercial release. With the expansion of biotechnology applications, especially in agriculture, pharmaceuticals, and industrial processes, the need for scientifically rigorous and ethically sound biosafety research trials has become increasingly important.
13.1.1. Scope of Biosafety Research Trials
1.1. Definition and Objectives
Biosafety research trials are confined, stepwise, and monitored field or laboratory-based studies designed to evaluate the safety, stability, performance, and unintended effects of GMOs. The overarching objectives include:
• Assessment of potential risks to human health, non-target organisms, and the environment.
• Evaluation of gene stability, trait expression, and environmental adaptability.
• Monitoring of gene flow and potential invasiveness or weediness.
• Determination of agronomic efficacy under field conditions.
• Data generation for regulatory dossiers and risk assessment reports.

Filler flowers play an essential role in floral arrangements by providing volume, texture, colour balance and supporting elements around major cut flowers. This chapter presents several commercially useful filler crops and their production technologies. It covers crop establishment, climate, soil, propagation, nutrient management, irrigation, crop regulation, pest and disease management, harvesting, drying and postharvest handling. Crops such as Ammi and other seasonal fillers are discussed from a commercial production perspective. The chapter also considers yield, stem quality and market requirements, demonstrating how filler crops can diversify farm income and create profitable opportunities within modern floriculture enterprises.

A TTO needs a planned budget of five years to sustain it in the long term. This involves budgeting the infrastructure, manpower, research, and outreach programs. The budgetary needs of a TTO will be based on the size and scope of the TTO and the maturity of the parent institution it caters to. The initial years will be focused on capacity development, the appointment of experts, and the establishment of processes of intellectual property management. The mature TTO will generate income through the returns of the licensing agreements and the collaborative efforts with the industry. Nevertheless, external funding through grants and government schemes plays a vital part, particularly in the initial years. The planned budget ensures long-term stability and development of the TTO.
9.1 Budgetary heads
9.1.1 Operational expenditure
Operational expenditure refers to the ongoing costs incurred by a TTO to maintain its day-to-day operations. This includes the cost of maintaining the office, including rent, power and utility bills, computer and network infrastructure, patent management cost, and administration charges. The filing and legal charges also fall within this category. The overheads will be a heavy burden if not planned with finances properly. A TTO has to be provided with a properly planned operational budget that covers the daily operational expenses. The operational expenses help the office perform effectively, sustain the employees, and effectively manage IP. The operational expenses that the budget normally covers include:

What is FISH ?

Fluorescent in situ hybridization is a technique in which single-stranded nucleic acids (usually DNA, but RNA may also be used) are permitted to interact so that complexes, or hybrids, are formed by molecules with sufficiently similar, complementary sequences. Through nucleic acid hybridization, the degree of sequence identity can be determined, and specific sequences can be detected and located on a given chromosome. Fluorescent in situ hybridization (FISH) is a powerful technique for detecting RNA or DNA sequences in cells, tissues, and tumors. FISH provides a unique link among the studies of cell biology, cytogenetics, and molecular genetics.

Principle of FISH

FISH involves the preparation of short sequences of single-stranded DNA, called probes, which are complementary to the DNA sequences the researchers wish to paint and examine. These probes hybridize, or bind, to the complementary DNA and, because they are labeled with fluorescent tags, allow researchers to see the location of those sequences of DNA. Unlike most other techniques used to study chromosomes, which require that the cells be actively dividing, FISH can also be performed on non dividing cells, making it a highly versatile procedure.

Forage crops are plant species used to feed domestic animals (such as cattle, sheep, goat, pig, and poultry). Both cultivated and wild plants can serve as forage. The domestic animals feed on forages either by grazing or stall feeding (cut fodder eaten in stalls).

Grazing, that is pasturing of animals, is cheaper and allows production of both livestock and dairy produce at a lower cost because no expense is involved in growing, cutting, transporting or feeding. In North and South America, Australia, New Zealand, and in many other temperate countries pastures are widespread and popular because sufficient land is available. While land is cheap and plentiful, labour is scarce and expensive; hence management of natural, and in recent years sown pastures has received much attention.

Until about fifty years ago, the natural grasslands in the Western countries met the minimum needs of the farmers and grassland management was operated at a level comparable to subsistence agriculture of many developing countries. In other words, emphasis was laid on exploitation of natural productivity of the grassland, using a low level of management based on minimum use of inputs like chemical fertilizers. However, it was soon realized that productivity of native grasslands will have to be greatly improved if the high demands for meat and milk were to be adequately met.

In India, however, out of a total cultivable area of about 131.11 million ha less than 2 per cent is utilized for forage crops because feeding the large human population takes the first preference for land use to produce grain and cash crops. It is, therefore, necessary to intensify efforts, particularly in over-populated countries, for a more efficient production of high yielding forage crops to get maximum return out of minimum land.

Introduction

Tree improvement through the application of genetic principles is directed towards modifying the heredity of tree populations so that they are able to meet the needs of the forester and the wood based industry. The breeder of annual crop plants resorts to hybridisation, selection, testing of the improved strains, multiplication of improved seeds and finally their release for commercial planting in that logical sequence. But the tree breeder, whose career may be shorter than the life cycle of the tree species taken up for improvement, tends to carry out the above operations in a modified form and often some of the operations are carried out concurrently.

Many of our tree species are wild unselected populations. They are, therefore, expected to exhibit considerable natural variability for many characters and also to respond well to selection for further improvement. Further, altitudinal and latitudinal variation in many characters between populations are expected for those tree species which have a wide geographical distribution. So, the selection of desirable seed sources or provenances is an essential step that is basic to all tree improvement programmes. The right provenance is identified through well laid out provenance trials at a number of locations in the area where the improved selection is to be grown. Further improvement is then planned and executed for exploiting the naturally available intrapopulation variation. The variation normally measured is the phenotypic variation which is the result of interaction of the genotype of the individual with the environment. However, what is essential for selection to be effective is genetic variation for the character under consideration. Genetic variation is assessed by appropriate progeny trials. Since many of the economically valuable characters of tree species are polygenically controlled and exbibit continuous variation, measurements have to be in metric units. The use of biometrical techniques for assessing and analysing this variation, therefore, becomes an essential requirement.

Introduction
The identification of crop varieties is crucial in improving agricultural productivity, ensuring quality control, and supporting breeding programs. The correct identification helps improve food security, conserve biodiversity, and manage pests and diseases. Traditionally, farmers were relying on visual traits and phenotypic characteristics, but these methods had their own limitations. Modern molecular techniques, such as DNA fingerprinting and genotyping, have revolutionized variety identification, rendering it both more accurate and efficient. This has allowed for early identification of varieties, assisting in the selection of desired traits and enhancement of crop resilience, thus contributing to more sustainable and productive agricultural practices (Azizi et al., 2021; Khan et al., 2024).
Importance of Crop Variety Identification in Agriculture
Crop variety identification remains one of the most important components in agriculture since it can improve crop production, enhance food security, and improve adaptability of the crops to diverse environmental conditions (Hasan et al., 2021). The major aspects related to correct identification of crop varieties include:
• Quality control and standardization: It is often to measure the quality of crops that could meet market demand, for optimum consumption, and for export.
• Biodiversity conservation: This helps to conserve native crop varieties and regards the preservation of genetic diversity that is key for breeding and in future agricultural resilience.
• Breeding program: It allows the identification of a variety for selecting traits such as disease resistance, drought tolerance, and yield potential for improving agricultural productivity through its effectiveness.
• Pest and disease management: It contributes to pest and disease management too since different crop varieties may express varying levels of resistance which also reflects in pest control strategy.
• Market preferences and economic value: This helps the farmers to supply some specific markets depending on the market demand of certain varieties so as to gain a profit (Hussain & Nisar, 2020; Sharma & Singh, 2021; Tazeb et al., 2018; Dheer et al., 2020; Jamali et al., 2019).

The frequency distribution of the populations of M₂ and M₃ generation for each genotype was extracted according to Panse and Sukhatme (1961). Skewness and Kurtosis for all the characters were calculated as per the standard method. Skewness refers to the symmetry of the frequency distribution. A measure of skewness is obtained as making use of the second and their moments about the mean. It is defined as,

Maize or corn (Zea mays L.), which was first domesticated in Mexico, is the most widely grown cereal crop in the world. Maize is consumed as feed and food, and also has myriads industrial applications, including the production of bioethanol and starch. Due to its highest genetic potential, production and productivity among the cereal crops, maize is called as “Queen of the Cereals”. Noble Laureate Dr. Norman E. Borlaug, the Father of Green revolution stated 15 years ago that “The past two decades saw the revolution in rice and wheat, the next few decades will be known as maize era”. In India, maize is the third largest grain crop and its production and demand is increasing rapidly. In India, it is primarily a non-food cereal crop, with only 10% being consumed as direct human food, while the rest being used for feed, industrial and export purposes. The effective deployment of various frontier technologies viz. genetic engineering, doubled haploids, molecular markers, phenomics etc.has greatly helped in raising productivity of maize crop worldwide in the last decade.The present chapter provides an overview on role of frontier technologies in maize crop improvement.

Most varieties of fruit plants have originated as chance seedlings or through somatic mutation and have been maintained and multiplied through vegetative propagation by fruit growers. The improvement of fruit plants through breeding has not yet attracted the serious attention of the research workers. Unlike field crops, fruit plant breeding is time consuming and laborious. Studies on the mode of inheritance of important economic characters of fruit plants are difficult because of the heterozygous nature of the plants. Moreover, varying degrees of sexual incompatibility have complicated the situation. Due to a long juvenile phase, it takes a long time to identify a useful hybrid. This factor has become a serious handicap because most research workers leave or shift from the programme before tangible results are obtained.

During the last two decades, considerable useful information has been generated on breeding methodology for the improvement of some of the important fruit crops. In this chapter fruits like mango, banana and papaya have been included.