eChapters

Primary cells are often preferred to established cell lines as they better represent the cells in vivo in terms of activity and functions. On the other hand, there is significant difference between original culture and present culture and even between cultures of the same cells in different laboratories. The rate of growth increases and there is change in chromosome numbers of lines over the time. The differences in phenotypes can cause problems in comparison and interpretation of results. The advantage of cell lines are ease handling, grow continuously, large yield and readily available. Primary cell cultures compared to established cell line are more fastidious in their growth requirements because many cell lines are derived from tumour tissue and demonstrate reduced growth factor requirements.

Abstract

Real-time PCR, also known as qPCR, is a highly sensitive technique used to determine the real-time amplification of a target DNA sequence. However, the presence of secondary structures or spurious amplicons can distort the results. When using SYBR Green chemistry, the features of qPCR primers differ from those of standard endpoint PCR primers. As a result, designing primers for SYBR Green qPCR requires extra caution and attention. This chapter outlines the essential features and conditions for designing qPCR primers, including primer length, melting temperature, GC content, specificity, and avoiding primer-dimer formation.

Abstract

PCR primer design is a crucial skill for molecular biologists, as it enables the study of gene expression, gene amplification, and molecular markers. Primer3 is an online tool widely used for designing optimal PCR primers, which are pairs of short oligonucleotides that initiate DNA synthesis in vitro using a thermocycler. This tool uses a range of parameters to generate high- quality primers, including target sequence length, melting temperature, GC content, specificity, and product size. Understanding and effectively utilizing these parameters is essential for achieving accurate and reliable PCR results. In this chapter, the practical aspect of primer designing using the online tool Primer3 has been discussed. The parameters and the purpose of their usage have been given in a tabulated manner.

The cell is the unit of structure of all plants and animals. With increasing complexity of organization, masses of different cell types tend to become localized and to form recognizable patterns. An aggregation of cells forming a definite pattern is called tissue. A still higher level of organization, tissues aggregate to form organs. The animal cell is a highly organized structure. Plant cell is different from animal cell  by having a cellulose cell wall.  There are a number of well defined structures viz., nucleus, mitochondria, lysosomes, Golgi apparatus etc in the cell. Animal cells also have centrioles whereas plant cells contain plastids such as chloroplasts. The remainder of the cell is the cytoplasm.

The prospects of Farm Animal Genetic Resources (FAnGR) are exceedingly uncertain. The FAnGR characterize the genetic makeup of the thousands of local breeds worldwide which are well-adapted for producing in harsh environments, and which for millennia have been essential in nourishing human communities. Currently, the genetic diversity in numerous livestock breeds across the world is threatened. Decades of consistent selection intended mainly for production increase has caused irreversible loss of genetic diversity (FAO, 2009).

True/False

Q.1.    The set of all possible outcomes of a variable along with the frequency (the number of occurrence) of each outcome or a group of outcome is called “Frequency Distribution”.
Q.2.    The frequency of an occurrence of each outcome of a trail divided by total frequency is known as “Relative frequency” of the outcome.
Q.3.    The distribution of total probability “1” into probabilities of occurrence of each outcome of a trial, is known as probability distribution of the outcome.

11.1 Learning Objectives
After comprehending this unit, we will be capable to:
i) Understand the conception of probability distribution;
ii) Appreciate the significance and applications of Binomial distribution;
iii) Understand the importance and uses of Poisson distribution;
iv) Know about the normal distribution and its’ applications.

11.2 Introduction

A probability distribution is a mathematical function that describes the likelihood of obtaining the possible outcomes of a random variable. In other words, it provides the probabilities of different events or values that a random variable can take. The sum of all probabilities in a probability distribution is equal to 1.

A probability distribution is a statistical function that describes all the possible values and likelihoods that a random variable can take within a given range. This range will be restricted between the minimum and maximum possible values.

The probability distribution shows how the set of all possible mutually exclusive events are distributed, and can be presented as an equation, a chart or a table.

A variable which can take different values with given probabilities is called a random variable.

There are numerous probability distributions which may be distinguished by whether the random variable is discrete, taking only a finite set of possible values, or continuous, taking an infinite set of possible values in a range of values.

A discrete random variable with only two possible values is called a binary variable, e.g. pregnant or not pregnant, diseased or healthy.

Probability distributions for discrete random variables The probability distribution for a discrete random variable y is the table, graph or formula that assigns the probability P(y) for each possible value of the variable y.

The theory of probability has its origin in the games of chance related to gambling such as throwing a die, tossing a coin, drawing cards from a pack of cards etc. Jerame Cardon (1501-1576), an Italian mathematician was the first man to write a book on the subject entitled “Book on Games and Chance”, which was published after his death in 1663. The systematic and scientific foundation of the mathematical theory of probability was laid in mid-seventeenth century by two French mathematicians B. Pascal (1623-1662) and Pierre de Fermat (1601 1665).

Important terms and concepts

To understand probability, let us first get acquainted with some important terms and concepts associated with probability.

• Trial: An experiment is any natural or planned activity for which it is not possible to predict the particular result called a trial.

• Events: The possible outcomes of experiments of a trial are called events. For example: Tossing of a coin in an experiment and one toss of the coin is called trial. In tossing of a coin there are only two outcomes or events, i.e. head or tail.

Types of events

Events can be of following different types:

• Exhaustive events

• Equally likely events

• Mutually exclusive events

• Dependent events

• Independent events

• Null or impossible events

• Exhaustive events: It is the total number of all the possible outcomes of an experiment.

True/False

Q.1.    A set containing all possible outcome of a trial in a study is known as “Sample space”.
Q.2.    If we are interested in observing the blood type of a child born in a hospital then sample space may be a set S is of the form S = {A, B, AB, O}.
Q.3.    If outcomes are continuous in nature, the sample space is specified as an interval that contains all possible outcome values.
Q.4.    When the occurrence of one event excludes the occurrence of other events, the events are said to be “Mutually exclusive events”.
Q.5.    In case of testing the type of blood of a newly born baby in a hospital, the blood types A, B, AB, O or gender (male, female) are exclusive events.

A. Definitions of probability

1. A probability relies on having an understanding of the theoretical model defining the set of all possible outcomes of a trial; we evaluate the probability solely on the basis of this model, without recourse to performing the experiment at all. It is often called an a priori probability. So, for example, we know that there are two equally likely outcomes when an unbiased coin is tossed: either a head or a tail. This is the model from which we can deduce that the probability of a defined event, obtaining a head, say, is 1/2 = 0.5.

2. The next approach to defining a probability, and the one commonly used in statistical inference, is to regard a probability as the proportion of times a particular outcome (the event) will occur in a very large number of ‘trials’ or ‘experiments’ performed under similar conditions.

The result of any one trial should be independent of the result of any other trial, so whether or not the event occurs in any one trial should not affect whether or not the event occurs in any other trial.

7.1 Introduction

The theory of probability has its origin in the games of chance related to gambling such as throwing a die, tossing a coin, drawing cards from a pack of cards etc. Jerame Cardon (1501-1576), an Italian mathematician was the first man to write a book on the subject entitled “Book on Games and Chance”, which was published after his death in 1663. The systematic and scientific foundation of the mathematical theory of probability was laid in mid-seventeenth century by two French mathematicians B. Pascal (1623-1662) and Pierre de Fermat (1601-1665).

While developing the toolkit for Participatory Curriculum Development participatory approaches like Focus Group Discussions and Field Level Workshops were adopted. The goal is to provide demand-driven services and to do so, extension professionals need to respond for farmers’ requests rather than deliver predetermined packaged solutions. It is also time to enable the farmers to empower and make their voices heard. Equally important is to make the service providers accountable to the farmers, which is possible when the farmers pay for the services they receive.

Environmental agents (xenobiotics) that are injurious to cells trigger a spectrum of responses that can range from cell death to adaptation, repair, and proliferation. Cell death, in turn, can take the form of apoptosis (a programmed form of cell death). Xenobiotics have been implicated in toxic cell injury, among them free radicals (nitrogen, oxygen, and carbon derivatives) are known to interact with cellular constituents including membrane lipids and chromagen materials.

Cytotoxicity

Cytotoxic: Toxic to cells, cell-toxic, cell-killing. Any agent or process that kills cells. The prefix cyto- denotes a cell. It comes from the Greek “kytos” meaning hollow, as a cell or container. Toxic is from the Greek “toxikon” = arrow poison.

In nutshell, cell cytotoxicity refers to the ability of certain chemicals or mediator cells to destroy living cells. By using a cytotoxic compound, healthy living cells can either be induced to undergo necrosis (accidental cell death) or apoptosis (programmed cell death).

Measuring cytotoxicity can prove to be very valuable tool in identifying compounds that might pose certain health risks in humans and animals. Cytotoxicity assays are widely used by pharmaceutical industry to screen for cytotoxicity in compound libraries from initial high-throughput drug screens for unwanted cytotoxicity. This can be of vital importance and prove to be quite indispensable during research phase of developing therapeutic new pharmaceutical products to ensure the safety of end-users. Additionally, understanding the mechanisms involved in cytotoxicity can likewise give researchers a more in-depth knowledge on the biological processes (both normal and abnormal) governing cell growth, cell proliferation and death. Compounds that have cytotoxic effects often compromise cell membrane integrity. Therefore, assessing cell membrane integrity is one of the most common ways to measure cell viability and cytotoxic effects. There are three commonly used methods which can be employed to assess in vitro cytotoxicity (disruption of cell integrity) in cell cultures:

Processing

On arrival in the laboratory, the specimens are processed immediately or refrigerated until processed. For inoculation into animal/cell culture/embryonated eggs (EE), swabs are shaken in fluid medium, faeces are dispersed in fluid medium and tissue specimens are homogenized in a high speed blender/pestle and mortar with the help of an abrasive like powdered glass or sand. Good quality hard sand should be properly washed in distilled water, treated with HCl, washed thoroughly, dried and sterilized before use. Generally the infective tissue is homogenized in a few ml of buffer usually having a pH of 7.2 to 7.3 to make about 10% tissue suspension. The suspension is centrifuged at about 1000g for 10-15 minutes to sediment the tissue debris and coarse materials.

Introduction

Currently, the agricultural industry is expanding exponentially. Improved economic growth and rapid population growth have spurred investor interest in the agricultural industry. The World Bank estimates it to be worth $3 trillion. With the urbanization and industrialization of the agricultural sector, the amount of waste generated has also increased, causing many environmental problems. About 5 million metric tons of biomass are produced annually from agriculture. These agricultural wastes become breeding grounds for pathogenic microorganisms if not properly treated and recycled, it is noteworthy that this agricultural waste can be used as substrates for the synthesis of valuable product or as raw material for renewable energy production.

Principle

Unlike bacteria, which can be cultivated on an artificial nutrient medium, viruses rely on a living host cell for replication. Eukaryotic or prokaryotic host cells that have been infected with viruses can be used to produce viruses. Subsequently, the medium and cells can be collected and used as a virus source. The production of viruses usually consists of a two-step procedure: f irst, the growth of host cells, and then the production phase. Understanding the complex relationship between the host cell and virus is essential. Various crucial factors have been recognised, such as the concentration of host cells during infection and the multiplicity of infection (MOI).

Materials required

Equipment

a) Biosafety cabinet level 2 (B2)

b) Carbon Dioxide (CO2 ) Incubator (set to 37°C with a 5% CO2 in air atmosphere)

c) Refrigerated centrifuge

d) Refrigerator

e) Inverted Microscope

f) Micro pipettes(1-10 μl, 10-200 μl, 100-1000 μl)

g) -86°C/-80°C deep freezer

h) Autoclave

i) Hot air oven

j) Filtration Assembly

Abstract

The data on production performance of Banni buffaloes during the 2008-09 and 2009-10 were collected from the animals selected under project “In-Situ Conservation of Banni buffalo”. The data on 208 animals were analyzed and overall means ± SE for total lactation milk yield, lactation length, peak yield, days to attain peak yield, milk yield per day of lactation length, service period and calving interval were observed as: 2917.76 ±  54.77 lit., 292.19 ± 2.88 days, 14.86 ± 0.18 lit., 47.50 ± 2.92 days, 9.99 ± 0.14 lit., 87.13 ± 4.39 days and 380.87 ± 2.36 days, respectively. Present findings on production performance indicate Banni buffalo an valuable germplasm maintained under extensive production system and arid climatic conditions of Kachchh and holds the promise of raising milk production of non-descript and low producing buffaloes through up-grading and crossbreeding. Therefore, Banni buffalo can be use as improver breed for genetic improvement of non-descript buffalo population in semi-arid and arid regions of the state / country.

Components of Project Finance
1. Capital Structure Determination: Deciding on the mix of debt, equity, and other sources of financing that will be used to fund the project. The goal is to structure the capital in a way that minimizes costs, maximizes returns, and balances risk. Here’s an elaborate breakdown of the steps involved in capital structure determination:
Understanding Financing Options
• Debt Financing: Borrowing funds that must be repaid with interest. Common sources include bank loans, bonds, and credit lines. Debt financing is often preferred for its tax benefits (interest payments are tax-deductible) and lower cost compared to equity. • Equity Financing: Raising capital by selling shares of the project to investors. This does not require repayment but dilutes ownership. Equity financing is useful for sharing risk and avoiding the burden of debt repayments.
• Hybrid Instruments: Combining features of both debt and equity, such as convertible bonds or preferred shares. These can provide flexibility and attract a broader range of investors.
• Other Sources: Grants, subsidies, and public-private partnerships (PPPs) can also contribute to project financing without adding to debt or diluting equity.