eChapters

Principle

Since the entire DNA sequence of the bacteriophage is known, we know exactly where a restriction enzyme will cleave the viral DNA and the size of the DNA fragments generated. This restriction enzyme digest will create a pattern of specific DNA fragments that will serves as a “DNA ladder”.

Reagents

Nuclease-Free Water: 14µl

10X Restriction Buffer: 2µl

Acetylated BSA (1mg/ml): 2µl

DNA (~1µg): 1µl

Restriction Enzyme (10u): 1µl

Final Volume: 20µl

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)

For most of the past 50 years world food production has outpaced rising demand. World population has doubled since World War II, whereas food production has tripled. In the developing world the calories available per person per day increased from an average of 1,925 calories in 1961 to 2,540 in 1992. World food production has expanded due to the Green Revolution—adoption of crop rotation, the mass production and use of petroleum-based fertilizers and chemical pesticides, expanded irrigation, and the introduction of genetically superior, disease-resistant cultivars. The trend may now be changing for the worse, however. Since about 1990 global grain production has risen only slightly and, despite slower rates of population growth, grain supplies per capita have fallen. In the worst case, Africa now produces nearly 30% less food per person than it did in 1967. As per FAO estimates, world food production would have to double to provide food security for the 8 billion people projected for 2025. By 2050, when world population is projected to be over 9 billion, the situation would be even more challenging.

Introduction
At the beginning of the 21^st century, with a population of 6.1 billion in 2000 and expected to be headed for 9.1 billion by 2050 (UN News Centre, 2005), the challenge of yet again increase food production by many folds only in 50 years has become a daunting task in itself. The situation is further worsen because now, we need to increase food production in sustainable way by 2050 on approximately the same area of arable land using less resources, particularly, fossil fuel, water and nitrogen, at a time when we must also mitigate some of the enormous challenges associated with climate change. Furthermore, there is the critical and urgent humanitarian need to alleviate poverty, hunger and malnutrition which is afflicting more than 1 billion people

Q.1.    What is an experiment ?

    Answer
    
    An experiment is a means of getting an answer to the question that an experimentor (researcher) has in mind. This may be to decide which of the several pain relieving drugs that are available in the market, is the most effective or whether they are equqlly effective.
    
    An experiment may be planned to compare the yield of different varieties of wheat. so that the varity giving the maximum yield may be recommended to the farmers to increase their production.

Preliminaries
    
Minimum Variance Unbiased Estimator and Cramer, Rao Inequality
    
Minimum Variance Unbiased Estimator (M.V.U.E.) and Black Wellisation
    
Results on UMVUE and MVUE
    
Efficiency and Consistency
    
Consistency
    
Sufficiency and Completeness
    
Distribution Possessing Sufficient Statistics
    
Sufficient Statistic for Several Parameters
    
Completeness of a Sufficient Statistic
    
Some Important Results in Theorem Forms
    
Minimal Sufficient Statistic
    
Results on Complete Sufficient Statistic

Biotechnology Products and Technologies
    
Bottlenecks in Fast Progress of Animal Biotechnology
    
Why Biotechnology should be Used in Animal Husbandry and Dairy Production
    
Contribution of Livestock Sector to Food Basket
    
Biotechnology and Environment
    
Biotechnology of Nutrient Partitioning
    
Genetic Programming for Better Animals
    
Biotechnology of Reproduction
    
Biotechnology of Animal Health

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.

Principle

The rate of joining of DNA fragments is affected by seven parameters:

DNA concentration,

Concentration of compatible ends,

Size of the fragments,

Presence or absence of complementary, single-stranded ends (some nucleases form blunt ends and in these cases special techniques are needed for joining),

G + C content and length of the complementary single strands,

Temperature of the reaction and

Ionic composition of the solution.

Steps for Cloning

Prepare DNA.

Select restriction enzymes for your insert and vector, and determine the appropriate reaction buffers.

Combine the following in a microfuge tube (30 µL total volume):
2 µg DNA
1 µL Each Restriction Enzyme
3 µL 10x Buffer
3 µL 10x BSA
x µL H₂O (to bring total volume to 30 µL)

Reproductive Biotechnology
    
Artificial Insemination (AI)
    
History and Future Perspectives of Artificial Insemination
    
Semen Cryopreservation
    
Buffalo
    
Goat
    
Cryoprotection and Anti Oxidants
    
Cryopreservation and Genetic Stability of Livestock Spermatozoa
    
Insemination Technique
    
Cattle and Buffalo
    
Sheep and Goat
    
Conception Rate with Artificial Insemination
    
Multiple Ovulation and Embryo Transfer (MOET)
    
Advantages of Embryo Transfer
    
MOET Includes the Following Steps
    
Superovulatory Treatments
    
Insemination
    
Recovery of Embryos
    
Synchronization of Recipients
    
Cattle
    
Buffaloes
    
Goats
    
Horses
    
In vitro Maturation, fertilization and culture of Oocytes
    
Historical Development
    
In Vitro Maturation of Oocytes
    
In vitro Sperm Capacitation
    
Scope
    
Limitations
    
Embryo Cryopreservation
    
Ultrasonography in Reproductive Management
    
Ultrasound-guided Transvaginal Oocyte Pick-up (OPU)
    
Embryo Cloning or Nuclear Transfer
    
Blastomere Separation
    
Embryo Splitting
    
Nuclear Transfer Cloning (NT)
    
Embryonic Stem Cell Complementations
    
Limitations and Future Prospects of Cloning
    
Sexing of Embryos and Separation of X and Y Chromosomes Bearing Sperms
    
PCR  in Sex Determination
    
Transgenic Animal Production
    
Gene Transfer Through Direct DNA Microinjection
    
Gene Transfer with Retroviral-Vector Assistance
    
Gene Transfer through Production of Germ-line Chimeras
    
Parthenogenetic Embryos: An Emerging Tool in Reproductive Biotechnology for Multiplication of Superior Germ Plasm
    
Applications of Parthenogenesis
    
Livestock Genomics and Marker Assisted Selection (Use of DNA Level Markers)
    
Stem Cells
    
Vaccine for Fertility Control and Increase Prolificacy
    
Auto-immunization to Increase Prolificacy
    
Biotechnology of Conservation of Livestock Breed Biodiversity
    
Gene for Prolificity: Booroola Merino
    
Role of Biotechnology in Enhancing Production
    
Milk
    
Biotechnology of Rumen Manipulation
    
Genetically Modified Ruminal Microorganisms

Biotechnology of Animal Feeding
    
Probiotics
    
Biotechnology Crops as Animal Feed
    
Biotechnology of Animal Products
    
Some New Products made Possible by Biotechnology Include

Vaccines
    
Veterinary v/s Human Vaccines
    
Modern Veterinary Vaccines
    
rDNA Technology to Prepare Vaccines
    
Veterinary Viral Vaccines
    
Live Vaccines
    
Killed Vaccines
    
Genetically Engineered Viral Vaccines
    
Veterinary Bacterial Diseases Vaccines
    
Veterinary Parasitic Vaccines
    
Vaccines against cancer
    
Swine Flu
    
Foot and Mouth Disease
    
Adenovirus Vector Vaccine
    
Johne’s Disease
    
Diseases of Poultry
    
Newcastle Disease in Poultry
    
Avian Influenza (Bird Flu)
    
Rabies
    
Disease Diagnostics
    
The Nucleic Acid-hybridization Technique
    
Diagnostics Based on pcr
    
Diagnosis by Restriction Fragment Length Polymorphisms and Related DNA-based Approaches
    
Diagnosis by DNA Probes and DNA Microarray Technology
    
Antigen-Capture Enzyme-Linked Immunosorbent Assay (ELISA) Proteomics
    
Disease Diagnosis Kits Developed
    
Brucelosis
    
Rabies
    
Canine Distemper and Canine Parvo Virus
    
Canine Hepatitis Virus
    
Foot and Mouth Disease
    
New Castle Disease
    
Egg Drop Syndrome Virus-76 (EDS-76)
    
Proteomics for Disease Diagnostics
    
Nanotechnologies in Diagnosis and Vaccine Development
    
Monoclonal Antibodies
    
Immunotherapy
    
Problems
    
Animal Genomics for Disease Resistance
    
Gene Knockout Technology
    
Gene Pharming
    
Gene Therapy
    
Types of Gene Therapy
    
Germ Line Gene Therapy
    
Somatic Gene Therapy

Principle

To retrieve and purify any specific DNA fragment from an agarose gel slice; the expected yield is from 50-75% of the amount in the gel slice.

Time required

Restriction digest - minimum of 2 hours

Electrophoresis - approx. 4 hours

Elution - 2-5 hours depending on fragment size

DNA Purification -2 hours

Environmental Biotechnology
    
Global Warming and Animal Health and Production
    
Genome Mapping
    
Human Genome
    
Livestock Genomics
    
Cattle Genome
    
Pig Genome
    
Bioinformatics
    
Major Branches of Bioinformatics Include
    
List of Bioinformatics Software Commonly Used in Animal Genomics Research
    
Sequence Analysis Software
    
Genetics Analysis Software
    
Functional Genomics Software
    
Biofuels
    
Advantages of Bioethanol
    
Ecological
    
Economical
    
Looking to the Future: The Potential of Second-Generation Biofuels
    
Future Role of Biofuels Depends on Profitability and New Technologies

Information Sources for Biotechnology
    
Books
    
General Biotechnology Books
    
Animal Biotechnology Books
    
Review Articles
    
Journals
    
Animal Biotechnology Journals
    
Biotechnology Job Listings
    
The Scientist
    
Patents
    
Requirements for Setting up a Biotechnology Laboratory
    
Essential
    
Beneficial
    
Useful
    
PCR
    
Specifications of a Good PCR Macine
    
Gel Documentation System Electrophoresis
    
Laboratory Safety and Hazards

Checks and Balances in Biotechnology Applications
    
Biosafety Issues in Animal Biotechnology Application
    
World Organization for Animal Health (OIE)
    
Recombinant DNA Safety Guidelines (1990)
    
Foreign Trade (Development and Regulation) Act, 1992 (2006)(Draft Amendment)
    
Revised Guidelines for Research in Transgenic Plants and Guidelines for Toxicity and Allergenicity Evaluation of Transgenic Seeds, Plants and Plant Parts (1998)
    
Guidelines for Generating Preclinical and Clinical Data for rDNA Vaccines, Diagnostics and other Biologicals (1999)
    
The Food Safety and Standards Act (2006)
    
Transgenic Animals Bioreactors
    
First Patent in Biotechnology
    
Biotechnology - Ethical Concerns
    
Some Priority Areas in the Field of Biotechnology
    
Suggestions for Future Development of Biotechnology Activities in the Field of Animal Health and Production in India
    
Epilogue

Animal Science Biotechnology Research Centres
    
Current Status and Some of the Achievements in Animal Heatlh
    
National Bioresource Development Board
    
Animal Biotechnology
    
Biotechnology Parks and Incubators
    
National Institute of Immunology, New Delhi
    
Bose Institute, Kolkata
    
Jawahar Lal Nehru University, New Delhi
    
National Centre For Cell Science, Pune
    
Centre for Dna Fingerprinting and Diagnostics (Cdfd) Hyderabad
    
National Institute for Plant Genome Research (Nipgr) New Delhi
    
Institute of Bioresources and Sustainable Development,Imphal
    
Institute of Life Sciences, Bhubaneshwar
    
Bharat Immunologicals and Biologicals Corporation Limited
    
International Centre for Genetic Engineering and Biotechnolgoy (Icgeb), New Delhi
    
Animal Biotechnology Research Abroad

Principle

RNA is negatively charged at neutral pH and when an electric field is applied, it migrates towards the anode. RNA retains much of its secondary structure during electrophoresis unless it is first denatured. The addition of formaldehyde to the agarose gel maintains the RNA in its linear (denatured) form. The following protocol is based on Formaldehyde Agarose Gel Method.

Denaturing polyacrylamide/urea gels in TBE buffer

Prepare 20ml of a 5% polyacrylamide gel containing 7M urea by adding:

47.5% acrylamide:

2.5% bis-acrylamide solution 2ml

10M urea 14ml

10X TBE buffer 2ml

10% freshly prepared ammonium persulfate 0.2ml

deionized water 1.8ml