eChapters

Introduction

Only after the discovery of restriction endonucleases it became possible to digest any DNA molecule at a specific site and any two pieces of DNA could be joined together with the help of another modifying enzyme, the DNA ligase. Consequently, a new branch of molecular biology, namely genetic engineering has been developed. With the help of genetic engineering techniques, it is now possible to identify and isolate the gene (for a protein of interest and insert it into the genome of another organism with the help of a suitable vehicle. The foreign gene thus cloned, can be amplified to produce millions of its copies. The insert gene can be expressed by the host to synthesize the gene product. The technique can also be used for understanding the regulation of expression of the cloned gene. The possibilities are endless and probably sky is the only limit. In this chapter we will briefly discuss the basic principles and protocol involved in gene cloning.

There are other groups that contribute to the reactivity of enzymes beside amino acid residues.

These groups are called cofactors - chemicals required by apoenzymes (inactive) to become holoenzymes (active).

There are two types of cofactors:

1. Essential ions - metal ions -inorganic
2. Coenzymes - organic molecules that act as group-transfer reagents (accept or donate groups)- can also be H⁺ and/or e^-

Both provide reactive groups not found on amino acids side chains.

Coenzymes can be either cosubstrates (loosely bound to enzyme; is altered, then regenerated) or prosthetic groups (tightly bound to enzyme).

Coenzymes can be classified by their source:

Biometric observations

The phenological stages i.e. germination, tasseling, silking, and physiological maturity were recorded from fixed mid three rows of each plot. A particular stage was supposed to be completed while 90% of the observed plant showed the characteristics of that phase and number of days was counted from date of sowing.

Days to emergence (DOE)
 
When the plant shoot pokes through the soil surface. Count the 75% seedlings emerged from at least 50% of the total rows starting from the centre. If 75% of the total expected seedlings emerged, then can be considered as 75% emergence. How the growth stages of maize are distinguished can be observed from the chart below.

The most important root and tuber crops cultivated on a worldwide scale are potato (Solanum tuberosum), cassava (Manihot esculenta), sweet potato (Ipomoea batatas), yams (Dioscorea spp.) and taro (Colocasia esculenta), but there are many others that are of regional, national or local importance, in a total of over a dozen dicot and monocot families. Most of these originated in tropical or subtropical areas and are mainly used as sources of carbohydrates. Many minor root and tuber crops, such as turmeric (Curcuma longa) and arrowroot (Maranta arundinacea, Tacca leontopedaloides), are also used in folk medicine and as spices (Sastrapradja et al., 1981). All these crops are vegetatively propagated. There are also vegetatively (or clonally) propagated crops that are not roots or tubers, e.g. bananas and sugarcane. A selection of the more important crops is given in Table 13.1 (Huaman et al., 1995).

For seed storage in the gene banks, the seed stored should have high quality which is greatly affected by seed handling during collection. The details on these issues have been given by Smith (1995).  In terms of seed storage characteristics seeds are of following 3 steps.

Active plant breeding programmes call for continuous search for new genes to be incorporated into advanced breeding lines to face the new problems and the challenges. The rapid technological developments in gene transfer technology in recent years have opened new avenues for introducing useful genes from wild and weedy relatives of crop plants. It is also a fact that due to genetic erosion, wild species, are getting lost. Therefore, systematic collection of germplasm of wild and weedy species have become more relevant today then ever before for the following reasons (Bothmer and Seberg, 1995).

6.1. Line × tester analysis

A successful approach of crop breeding programme depends on the proper choice of best parents for hybridization and the ideal selection adopted in the early generation. But, the selection of parents for hybridization programme is relatively tough in the case of complex traits like yield and their components as they governed by large number of quantitative genes which are influenced by environments and seasons. Thus, breeders often meet with difficulty to fix in advance the desirable parents which will give superior progenies. In such a situation, knowledge on the nature of gene action on such complex quantitative traits of economic importance is necessary to plan and adopt appropriate selection techniques (Simmonds, 1979) and breeding methodology.

Combining ability analysis gives useful information regarding the selection of parents in terms of the performance of the hybrids. This analysis elucidates the nature and magnitude of various types of gene action involved in the expression of quantitative traits (Dhillon, 1975). Thus, the knowledge of combining ability serves as an useful tool for the selection of plants for hybridization and further exploitation. Combining ability consists of general and specific combining ability.

Sprague and Tatum (1942) defined general combining ability (gca) as the “average performance of a line in hybrid combinations” and specific combining ability (sca) “as those cases in which certain combinations do relatively better of worse than would be expected on the basis of the average performance of the lines involved. The general combining ability provides an indication of the importance of genes of largely additive in nature, while specific combining ability indicates the importance of non-additive gene effects.

Several biometrical techniques are available to estimate the general combining ability of parents and specific combining ability of hybrids. Among them, line × tester analysis is extensively used both in self and cross pollinated crops. The advantage of line × tester analysis is that a large number of inbred lines can be evaluated than diallel analysis. Besides, it provides additional information on the performance of the specific hybrids on gene action in contrast to top cross or polycross. The line × tester analysis is an efficient biometrical approach for assessing the combining ability of parents and hybrids (Kempthorne, 1957).

Commercial Crop Breeding

The term commercial crop breeding is straightaway equivalent to ‘commercial plant breeding’ and remains largely plant breeding with slight twist that commercial plant breeding aims at delivering commercial products to be delivered to the end users that is the farmers as per the need of the consumers/ markets. Thus, the farmers plant those products (open-pollinated cultivars and hybrids) which are easily accepted by the consumers and which are easy to be handled by the traders as the products reach to the traders and then from traders to the retailers and ultimately to the consumers.

Any commercial plant breeding programme run by the private seed companies typically called as Research and Development (R & D) is carried out through various models operating. There is no fixed organizational set up and different companies adopt the working set up as per size of their programme, availability of manpower and resources, area of the breeding station and several other considerations. One thing is clear that any commercial plant breeding programme will have more numbers of crop breeders than the supporting scientists.

Many developing countries, for example India, China, Thailand, Brazil, Mexico, Egypt and South Africa, utilise the so-called modern biotechnology, based on genetic engineering and genomics. Agricultural biotechnology is the most widespread biotechnology in developing countries, but only a few of them are able to carry out all of the research and development activities leading to the commercialization of genetically modified seeds. These include basic research in molecular and cell biology and genetics, greenhouse and field trials according to  internationally agreed biosafety standards, risk assessment and management, respect for intellectual property rights relating to the transferred genes and to the creation of new crop varieties, production of genetically modified (GM) seeds by private corporations or working in cooperation with the public agricultural research sector, extension activities aiming at delivering the new seeds to the farmers and biovigilance in the fields of GM crops so as to detect any abnormalities or any hazards caused to the environment and to conventional crops (Sasson and Elliot, 2004).

Biotechnology is defined as a set of tools that uses living organisms or parts of living organisms to make or modify a product, improve plants, trees or animals or develop microorganism for specific uses. Agricultural biotechnology is the term used in crops and livestock improvement through biotechnology tools. Biotechnology encompasses a number of tools and elements of conventional plant breeding techniques, bioinformatics, microbiology, molecular genetics, biochemistry, plant physiology and molecular biology.

Principle

Plasmids requires hosts to replicate, hence it has to be inserted into bacteria. But efficiency of uptake of plasmid DNA by bacteria is low and will not do so without getting degraded. Therefore, to enhance the probability of uptake of the plasmid DNA by bacteria it is necessary to make the bacterial cells “competent” to absorb the plasmid DNA. One of the methods to increase efficiency of cell competence is given below. It should be noted that competent cells are more fragile than normal bacteria. Vortexing the cells, heating the cells above 42ºC or to 42°C for prolonged periods, or exposure of the cells to abusive treatments may kill them

The chapter on completely randomized design (CRD) presents an overview of its theoretical principles, structure, and randomization methods. It includes the mathematical modeling of CRD and the analysis of ANOVA tables. Additionally, the chapter facilitates an understanding of how to analyze CRD and factorial CRD data using R, offering valuable practical insights for research applications.

Completely Randomized Design

Completely Randomized Design is a type of experimental design commonly used in statistical analysis and research. It is applied under specific conditions:

• Homogeneous units: CRD is suitable when the experimental units are consistent and uniform across the study area.

• Adherence to experimental design principles: This design conforms to two principal aspects of experimental design i.e. Randomization and Replication.

• Flexibility in replication: It allows for both equal and unequal replication of treatments within the experimental setup.

• Suitable for controlled environments: CRD is particularly effective for experiments conducted in greenhouses or laboratories, where conditions can be tightly regulated

Introduction
Gene Regulation before delving into gene regulation, it’s essential to understand the concept of gene action. Gene action encompasses the expression of a particular trait or phenotype by a gene. This expression hinges on the synthesis of a specific protein encoded by the gene, followed by the protein’s involvement in a metabolic or biosynthetic pathway leading to the manifestation of the phenotype. The sequence of events in phenotype production may, therefore, be written as follows: DNA → RNA → Protein → Phenotype (Griffiths et al., 2000).
In molecular terms, gene action refers to the production of an RNA transcript from the gene (Lewin, 2004).
Gene Regulation
Gene regulation is the process by which cells control the amount of protein or RNA produced by a gene, either increasing or decreasing its production as needed (Alberts et al., 2014). Different tissues in eukaryotes produce varying sets of proteins and enzymes tailored to their specific requirements.

Composites defined as advanced generation populations maintained by random mating in the material constituted through mixing of equal quantity of seeds of crosses showing less inbreeding depression in F2 over F1. The components of composites inbreds, open-pollinated populations, locally available open-pollinated cultivars, hybrids, advanced generation lines, improved populations. Mixing the seeds of several phenotypically outstanding lines produces a composite variety and encouraging open pollination to produce crosses in all combinations among, the mixed lines. The lines used to produce a composite variety are rarely or not tested for combining ability with each other like as done in case of synthetics where the combining ability of the component lines is invariably tested for GCA. Composite are commercial varieties and are maintained by open - pollination in isolation. While mixing the seeds of various genotypes, care should be taken that the component lines are similar in maturity height, seed size, colour, etc. so that the composite maintained through random mating in isolation manifests a reasonable degree of phenotypic uniformity. The variety is maintained by open pollination. Farmers can use their own seed for 3 to 4 years.

Abstract

Advances in genome sequencing technologies have made it possible to uncover the whole genome sequences of numerous organisms from microbes to higher eukaryotes. Haemophilus influenza, a free living bacterium, was the first complete genome to be sequenced in 1995 by a team headed by J. Craig Venter and Hamilton Smith at The Institute for Genomic Research (TIGR). Immediately after this, Mycoplasma genitalium, a bacterium responsible for reproductive-tract infections and well-known for having the shortest genome of all free-living organisms was sequenced. Subsequently, TIGR unearthed the genome sequences of many microbes like Methanococcus jannaschii, Archaeoglobus fulgidus, Helicobacter pylori, Borrelia burgdorferi etc. During the years 1996-2000, the complete genome sequences of several model organisms such as bacterium Escherichia coli K-12, baker’s yeast Saccharomyces cerevisiae, nematode Caenorhabditis elegans, fruit fly Drosophila melanogaster etc were published. Since then the complete genome sequences of many other eukaryotes have been released by the genome sequencing community.

Computer vision provides an important foundation for automated plant analysis, monitoring and precision agriculture. This chapter explores computer vision approaches for plant phenotyping, disease detection, growth monitoring and yield prediction. Traditional image-processing methods are considered alongside machine learning and modern deep learning techniques, including convolutional neural networks and vision transformers. Image segmentation, object detection and classification are used to extract meaningful information from plant images and video data. The chapter also examines multispectral and hyperspectral imaging, IoT integration and real-time monitoring. Environmental variability, dataset quality, computational complexity and model generalization are discussed, together with transfer learning, data augmentation, edge computing and future directions

Introduction

Agriculture has played an important role in the global economy in recent years. The further growth of the population is leading to a gradual decrease in the area of the agricultural land and further growing stresses on the farm system. Agricultural food production methods are rising in demand for successful and healthy. Innovative sensoring and motive technologies and advanced information and communications systems and Artificial Intelligence must supplement conventional agricultural management methods to speed up farm productivity growth more accurately, thereby promoting the production of high-quality and high-performance agriculture. Computer vision monitoring systems have become valuable tools in farm operations over the past decades, and their use has increased significantly.

What is Computer Vision?

Computer vision can be characterized as a field of AI for extracting information from digital images. The type of data obtained from an image may be different from recognition, navigational space measurements or applications of increased reality.

The applications of computer vision can also be described. Computer vision builds algorithms that recognize and use the quality of pictures for other purposes.

Cross-pollinated species, or outbreeders, are highly heterozygous, heterogeneous and carry a higher mutation load than the inbreeders. Their mode of reproduction bestows upon them considerable heterozygous balance under panmitic population structure. Inbreeding such species disturbs the natural rhythm and leads to inbreeding depression. It is, therefore, essential that if selfing has created any disturbance in outbreeders, the heterozygous structure be restored. Since artificial selfing and outcrossing can be carried out in such crops at will, all breeding methodologies, even those meant for self-pollinated crops, can be used for their improvement. In fact, allogamous crops have provided opportunities for innovating breeding procedures to achieve the maximum efficiency by combining selfing and outcrossing cycles.

Thus, the classical separation of breeding procedures for self- and cross-pollinated crops has become unnecessary, particularly, for outbreeders which are amenable to selfing. Exploitation of heterosis has been the major objective in cross-pollinated crops and most breeding procedures developed earlier aimed at maximising heterosis. In later years, with better understanding of genetic structure of populations and availability of efficient biometrical methods of estimating genetic parameters, newer procedures of accumulating desirable genes have been developed. The presently available breeding procedures for cross-pollinated crops can be broadly classified under two heads: (1) Hybrid development, and (2) Population improvement. Both these methodologies, though independent, are not mutually exclusive. The steps for operating the two procedures, however, have contrasting requirements. For hybrid development, fixation of genes through cycles of selfing is necessary whereas such fixation is avoided while developing populations. As can be seen from the scheme given below, in both cases the homozygosity attained through initial selfing is converted to heterozygosity at the end of breeding programme. Irrespective of whether hybrids, synthetics or composites are being developed, great emphasis is placed on the attainment of homogeneity in the end products.

Man-directed evolution of distinct types and cultivars (cultivated populations) of self-fertilised crops had their beginning about 10,000 to 12,000 years ago. As a breeding system, self-fertilisation favoured constancy and fitness (Stebbins, 1950) of the genotypes to the immediate environment. This provided an alternative to the slash and burn migrant agriculture of the Neolithic farmers (7,000-6,000 B.C.) and to stabilisation of production.

Manu Smriti (700–500 B.C.) proclaimed Subeejam Sukshetre Jayate Sampadyate meaning that good seed in a good field yields abundant produce. The knowledge that good seed is essential for a good harvest, led to conscious efforts for selecting types which provided a dependable source of food. Besides their utility as food, choice of plants and their seeds as progenitors of the next generation depended on personal fancy, observation and experience. As farmers learnt their crop plants better, they sorted them out into similar types (species). While migrating from the centres of origin they carried seeds of mixed populations.

Self-pollinated cereal crops of barley, wheat, oats and important grain legumes such as lentil, peas and chickpea spread from the Near East Centre, first to far and remote areas of Asia, Europe and Africa; and subsequently in comparatively recent times, to the Americas and Australia. In new environments, the introduced mixed populations diversified into well adapted identities of land races. These land races continued to be the main basis of cereal production till early 1900. Improvement in crop plants made by some early civilisations was mainly through introduction and mass selection, based on the natural tendency of preserving the seeds of preferred colour and size from vigorous, healthy and more productive plants.

It was only after the rediscovery of Mendel’s laws in 1900 that breeding procedures based on the knowledge of the genetic principles and the inheritance pattern of different characteristics were developed. This permitted the improvement of crops with objectivity and predictability.