eBook Chapters

ABSTRACT

Pesticide degradation is the breaking down of toxic pesticides into a non-toxic compounds and, in some cases, down to the original elements from which they were derived. The most common type of degradation is carried out in the soil by microorganisms, especially the fungi and bacteria. Microbial degradation depends not only on the presence of microbes with the appropriate degradative enzymes, but also on a wide range of environmental parameters. Pesticides which are rapidly degraded are called non-persistent while those which resist degradation are termed as recalcitrant (persistent). Microorganisms are able to degrade a large variety of compounds, including pesticides under laboratory conditions. However, methods have yet to be developed to decontaminate the environment from pesticide residues. Continuous efforts are required in this direction, and at present several PGPR capable of degrading pesticides have been isolated from the natural environment. Catabolic genes responsible for the degradation of several xenobiotics, including pesticides, have been identified, isolated, and cloned into various other organisms such as Streptomyces, algae, fungi, etc. In addition, recombinant DNA studies have made it possible to develop DNA probes that are being used to identify microbes from diverse environmental communities with a unique ability to degrade pesticides. Bioremediation treatment still requires more intense research through specific microbial degradation of broad range of organic pollutants. This review describes recent advances in biodegradation of pesticides by addressing the microbes involved in pesticide degradation.

Microbial diversity is the key to human survival and economic well being. It provides a huge reservoir of resources which can be utilized for our benefit. Yeasts have been isolated from many sources for industrial purposes. The impact of yeasts on food and beverage production extends many fold beyond the popular notions of bread, beer and wine fermentations by Saccharomyces cerevisiae.

Diverse dynamic mixed populations of prokaryotic and lower eukaryotic microorganisms govern the fermentation and maturation processes and, finally, impact the quality of wine. Both of these parameters apparently contribute to regional differences in the quality of produced wines and characterization of regional wines should involve the study of microbial populations and their members active in the given region.

ABSTRACT

The current paper discusses the diversity, distribution and commercial importance of Indian Magnolias. Globally, these species are found in tropical, subtropical and temperate forests of southeastern Asia and tropical America. In India, there are 25 species of Magnolias and most of them are distributed in Northeast India, with highest number in Assam. Of all the species, two (Magnolia gustavii and M. pleiocarpa) are critically endangered, one (M. pealiana) endangered, two (M. manii and M. nilagirica) vulnerable, nine least concern and ten data deficient at global level. The highly threatened nature of most of the magnolias species calls for their immediate conservation and protection measures. Members of this genus are known to be rich in a wide variety of biologically active compounds including alkaloids, flavonoids, lignans, neolignans and terpenoids. Many of the species have been found to possess potent procognitive activity, anti-oxidative, anti-microbial, anti-inflammatory, anti-angiogenic, diuretic, anti-ulcer, analgesic, anti-helmintholytic, and anti-cancer activities. The species have huge economic potential and are used for a number of purposes including ornamental, medicinal, culinary, timber and joinery works.

INTRODUCTION

The entire Himalayan region is vast and its diverse geographical grouping is unique and has complex ecosystem and occupies about 18% of the total geographical area of India. Indian Himalaya (27O 50' – 37O 06' N and 72O 30' – 97O 25' E) includes the parts of trans, north-west, west, central and east Himalaya and covers approximately an area of 4,19,873 Km2 with 2500 Km length and 240 Km width. The north-western Himalayas generally represent the Himalayan tract from Kumaon in Uttar Pradesh to Chitral in Gilgit. The Himalayan ranges situated in this region exhibit a diverse climate, topography, vegetation, ecology and land use pattern. The annual average rainfall varies from 8 cm in Ladakh to over 200 cm in some parts of Himachal Pradesh and Uttar Pradesh. The vegetation may vary from the scrub in the lower hills to arctic in the Greater Himalayas. The unique physiography, climatic conditions and soil characteristics of the area has resulted in a variety of habitats and a significant biological and cultural diversity. Along the altitudinal gradient (300-800 m.a.s.l) the vegetation varies from sub-tropical, temperate, sub-alpine to alpine types. It supports about 8000 species (47.06% of the total flowering plants of India) of which 30% are endemics, 10.2% trees, 8.44% wild edibles and over 15% medicinal (Samant and Dhar 1997).

Abstract

Lianas are slightly lesser known angiosperms, grows typically in the form of climbing vines mostly in tropical and subtropical forests. Such plants are characterized by scandent thick stems, woody stem in nature and vary in shape and size. They depend on supports in the form of trees, fence and usually climb upward towards the sunlight as they needed for their survival. Keeping the importance of lianas in forests and household, this study was conducted to document the liana angiosperms growing in Western Himalaya along with their medicinal potential for human and animals. This study reveals a total 64 lianas species belonging to 24 families recorded from Jammu and Kashmir, Ladakh, Uttarakhand, Himachal Pradesh and Punjab. This investigation also provided with medicinal potential of these species, which is used as ethnomedicine by local people living in the interior regions of hills and mountains of Himalaya. In this communication, we highlight lianas as an example of an underestimated forest element that, on the one hand, is a key component in and around forests. This study also helps in conservation of endangered liana species in Himalaya.

1. Introduction

Fungi are ubiquitously present in air, water, soil and organisms, or on organism surfaces. They are a group of eukaryotic system that are of great applied and scientific interest to common man, microbiologists, agriculturists and industrialists. They are a diverse group of organisms with respect to taxonomy, size, morphological appearance, mode of nutrition and reproduction. Fungi are eukaryotic spore bearing protists that lack chlorophyll. Fungi comprise the molds and yeasts. Yeasts are usually unicellular measuring only several microns in diameter, whereas molds are filamentous and multicellular which may grow up to several meters in diameter like the extremely large polypores. Fungi generally reproduce both sexually and asexually (Gow and Gadd 1995).

Although it is not easy to define fungi, biologists currently use the term “Fungus” to include nucleated, spore bearing, achlorophyllous organisms with absorptive nutrition which generally reproduce sexually and asexually, and whose filamentous branched somatic structures are typically surrounded by cell walls containing cellulose of chitin or both. A feature which all fungi have in common is their mode of nutrition. Like animals, fungi are heterotrophic organisms, which must consume preformed organic matter. They may live as saprophyte or parasites. They are of great economic importance to man and play an important role in the disintegration of organic matter and affect us directly by destroying food, fabric, leather and other commercial goods. They are responsible for a large number of diseases of plants, animals and man.

Horticulture is a wide field and includes a great variety and diversity of crops. In general, these crops are classified as follows:

1. Crops grown for the food and/or vitamin and mineral content, for e.g., apple and sweet potato.

2. Crops grown for their beverage properties, for e.g., tea and coffee.

3. Crops grown to beautify the environment, for e.g., the grasses of the well planned lawn.

4. Crops grown to express sentiments, for e.g., cut roses, jasmine.

5. Crops grown for perfume for e.g., rose, jasmine.

6. Crops grown for spices for e.g., pepper, nutmeg.

UNTIL THE 1940S BIOLOGISTS ARGUED ABOUT WHETHER DNA OR PROTEIN WAS THE MOLECULE OF HEREDITY

Nuclei observed to divide when cells reproduced themselves

Large amounts of both DNA & protein in the nucleus

Many though protein was most likely molecule of heredity

Proteins made of 20 different amino acids- could have more variety than DNA (made of only 4 different bases)

Abstract

Despite several controversies surrounding the genetically modified (GM) crops or their produce, the production of such crops is constantly increasing, especially in developing countries. To regulate the GM produce in the market, several countries have set very strict regulatory procedures for the approval to grow, import or utilize these GM crops as food or food ingredients. Hence, for regulatory compliance on GM labeling and identification of genetic traits, the development of reliable, sensitive and precise detection assays have become essential. The detection of GM produce is carried out by detecting either the inserted foreign DNA or the novel transgenic protein specifically expressed in GM crops. This article broadly focuses on some DNA and protein-based detection methods employed for GM produce and significant current advances in such detection technologies.

Delineating species boundaries correctly and also identifying species are crucial to the discovery of life’s diversity because it determines whether different individual organisms are members of the same entity or not (Dayrat, 2005). The identification of species depends on the knowledge held by taxonomists whose work cannot cover all taxon identification requested by non-specialists. To deal with these difficulties, the ‘DNA Barcode of Life’ project aims to develop a standardized, rapid and inexpensive species identification method accessible to non-taxonomists.

DNA barcoding is a technique in which species identification is performed by determining and comparing the sequences of several hundred base pairs of nucleotides from a small fragment of an identified gene, with the aim of contributing to a wide range of ecological and conservation studies in which traditional taxonomic identification is not practical (Krishnamurthy and Francis, 2012). The two main ambitions of DNA barcoding are to assign unknown specimens to species and to enhance the discovery of new species and facilitate identification, particularly in cryptic, microscopic and other organisms with complex or inaccessible morphology (Hebert et al., 2003a). Apart from these, identification of species previously defined by other criteria to also is an objective of barcoding. Another major objective could be the definition of operational units for ecological studies. This may also be used for forensic purposes by linking the specimens with already identified databases.

Aim: To identify culturable gut bacteria from insects.

Materials required: Micro scissors, Micro pestle, 1.5 ml centrifuge tubes, Pipettes, Petri plates, Inoculation loop, 0.5 ml PCR tube, Glass slides, non absorbent cotton etc.

Instruments required: Autoclave, Refrigerator, Laminar Air Flow, Homogenizer, Biological Oxygen Demand (BOD) incubator, Thermal cycler, Gel Doc unit, Deep freezer pH meter, Spirit lamp.

Chemicals required: 70% ethanol, Phosphate Buffer Saline (PBS solution, pH 7.4), Distilled water, 0.01 M TE buffer, Forward primer - 27F (5’-AGAGTTTGATCCTGGCTCAG-3’) and reverse primer - 1492R(5′ AAGGAGGTGATCCAGCCGCA), PCR Master mix, Nuclease free water, Agarose, Ethidium bromide, crystal violet, Grams’s iodine, 95% ethanol, Safranin, 3% hydrogen peroxide, Tryptophan broth, Kovac’s reagent, King’s Base agar.

Protocol for DNA extraction using the Qiagen DNeasy Blood and Tissue Kit (Catalogue No. 69504) presented here

Contents of the kit: DNeasy mini spin columns (2 ml), collecting tubes (2 ml), tissue lysis buffer (ATL), lysis buffer (AL), wash buffer 1 (AW1), wash buffer 2 (AW2), elution buffer (AE) and proteinase-K.

Note: Check for the validity of the product before dilution. Dilute the ATL and AL buffer with molecular gradient ethyl alcohol to a specified volume mentioned on the bottle before starting experiments. Keep a tick mark the bottle cap after dilution with ethyl alcohol for easy identification.

Equipments required: Microscope, centrifuge, vortex mixture, water bath, pestle and mortar/homogeniser, micropipettes, micro tips, eppendorf tubes, needle, micro scissor, forceps, camel hair brush, thermometer, tube stand, masks and gloves.

Previous Year Questions

1. What is DNA fingerprinting? Discuss the applied aspects in modern agriculture. (15M, CSE 2021)

• The term DNA fingerprinting in its original sense refers to the method developed in 1985 by Sir Alec Jeffreys (Jeffreys et al., 1985) and his associates for the simultaneous detection of highly variable DNA fragments by hybridisation of specific multilocus probes to electrophoretically separated restriction fragments?

• The DNA fingerprints, resembling barcodes, are unique to the individual and hence can be used in much the same way as conventional fingerprints - to identify individuals with absolute certainty?

DNA Fingerprinting Techniques

• A large number of fingerprinting techniques are in currency, which are variations and combinations of restriction fragmentation and polymerase chain reaction (PCR).

• Restriction fragment length polymorphism (RFLP) involves digesting DNA with restriction enzymes, separating the resultant DNA fragments by gel electrophoresis, blotting the fragments to a filter and hybridising probes to the separated fragments. Specific probe-enzyme combinations give highly reproducible patterns.

• With the advent of PCR technology a number of simpler techniques have become available for molecular characterisation.

• These include, random amplified polymorphic DNA (RAPD) and its several variants, intersimple sequence repeats (ISSR), amplified fragment length polymorphism (AFLP) and sequence tagged microsatellites sites (STMS).

ABSTRACT

DNA fingerprinting is increasingly being used for identification of varieties, quantification of similarity between varieties and study of genetic diversity available in the population.

This is all the more important in crops where morphological variability is less or one variety is known by different names or different varieties are known by same name.

DNA fingerprinting using RAPD method has been standardized for tapioca. Dellaporta method has been used for extraction of DNA and amplification was done following William’s method. This technique has been used for molecular characterisation of accessions and identification of duplicates in the germplasm maintained at Central Tuber Crops Research Institute, Thiruvananthapuram. This technology has been extended for DNA fingerprinting of released varieties and also some of the elite breeding lines.

Eleven varieties released from the institute were fingerprinted using a few primers, which showed high polymorphism in tapioca. One single primer gave distinct banding pattern for all the 11 varieties and it was capable of earmarking each variety.

Individual variety was finger printed for 20 different primers and the results were documented. This technique is being extended to promising land races as well. DNA fingerprinting will be useful for protecting Plant Breeders’ Right as well as Farmers’ Right in the new Patent Regime.

Apoptosis is triggered by extrinsic and intrinsic stimuli as radiation, oxidative stress and genotoxic chemicals. It is defined by characteristic changes in the nucleus morphology including chromatin condensation and fragmentation, overall cell shrinkage/rounding and formation of apoptotic cell bodies. Among the many markers of apoptosis, DNA fragmentation is considered a hallmark. The DNA ladder assay described here is a simple, sensitive, cost-effective and rapid method for estimating apoptosis in cells.

Isolation of Genomic DNA

Genomic DNA from primary cells culture of PMNC and splenocytes of rats is isolated by using standard phenol-chloroform DNA isolation protocol of Sambrook and Russel (2001) as described below:

1. Preparation of primary cells culture of different cells, namely-PBMNCs, thymocytes and splenocytes form Wistar rats and viability checked by trypan blue dye exclusion test.

2. 106 cells/ml of primary cell culture is used and seeded in 24 well culture plates containing along with treatment with xenobiotics at different µM concentration.

3. Cell culture plates are incubated in CO2 incubator at 37°C with 5% CO2 for 12 hrs.

4. The cells are collected in 1.5 ml of micro centrifuge tube and centrifuged at 3200 rpm for 10 min at 4°C; supernatant is discarded and pellet is washed with PBS (pH 7.2).

5. DNA extraction buffer (500 µl/tube) is added in tubes. Gently tap to disperse the cell pellet in extraction buffer and keep in water bath for 1.0 hr at 37 °C (it is to ensure that pellet is completely suspended so that cells are accessible to SDS and Proteinase K).

6. 10 % SDS is added (20 µl/ml) to cell suspension and the tubes are gently mixed by inverting the tube. The contents of tube become viscous which indicates lysis of cells. After that sample is handled gently from onward to avoid shearing of DNA.

7. Proteinase K (15 µl) of 20 mg Proteinase K/ml of buffer is added in tubes in two pulse i.e. half the requirement is added to tube in 1st pulse, mixed gently end to end and kept in water bath at 50°C. After 3-4 hrs, the second pulse of the remaining amount of Proteinase K is added and tubes are incubated at 50°C overnight.

8. Equal volume of equilibrated phenol (Tris saturated phenol pH> 7.8 ) is added in the tubes next day morning. Mix the contents of tube gently by inverting the tube for 15 min till a light coffee coloured uniform solution without any balls of phenol is formed and then centrifuge at 3400 rpm for 15 min.

Objective: To isolate DNA from plant sample by CTAB method and quantification by spectrophotometer.

Background information:

DNA isolation from different plant tissues viz., leaf, stem, root, flower and seed is a challenging task due to hindrance of variety of variety of biochemical compounds present and further problem increases with diverge plant species. In case of animal tissues the diversity in cellular bio-molecules in different species is much lower compared to plants. Two major phyto bio-molecules that have wide variation between species and pose serious problems in DNA isolation are: polyphenols and polysaccharides. These phyto bio-molecules if contaminated in DNA then may hinder further down applications like PCR, restriction digestion, sequencing, etc.

Several methods have been developed and reported that can remove polyphenols and polysaccharides efficiently during DNA isolation from plants. For this purpose two different chemicals are widely used,the first one is CTAB (cetyl trimethyl ammonium bromide) that is a cationic detergent and helps in removal of polysaccharides and the second one is polyvinylpyrrolidone that helps in removal of polyphenols. Due to above properties the CTAB based DNA extraction buffer is very common for plant DNA extraction.

Significance of different types of chemicals found in CTAB based plant DNA extraction method is explained as under:

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.

ABSTRACT

The process of developing new crop varieties involves many steps and takes several years. In recent times, biotechnological advances have helped in reducing these considerably. Apart from the techniques of haploid culture and genetic engineering, one of the tools which is making it easier for scientists to select plant traits and develop new varieties is the use of molecular markers. Molecular markers can be used for (a) characterization of germplasm (b) varietal identification and clonal fidelity testing (c) assessment of genetic diversity (d) validation of genetic relationships and (e) marker- assisted selection. These have widespread applications in management of genetic resources as well as in breeding programmes. Marker-assisted selection has practically revolutionized breeding programmes. Traditionally, plant breeders have selected plants based on their visible or measurable traits called the phenotype. But this process can be difficult especially in the case of multigenic or quantitative traits. Molecular markers can be used to trace linkages with traits of importance. In the area of horticulture, this is a very useful tool in tree breeding, where the breeder has to wait for several years for phenotypic expression of the desired trait. A number of DNA marker technologies are now available and rapid advances are being made in the development of more sensitive marker technologies. The techniques involved, the various applications of DNA marker technologies in horticultural crops and future perspectives in the national context have been discussed.

Introduction

DNA denaturation, also called DNA melting, is the process by which double-stranded deoxyribonucleic acid unwinds and separates into single-stranded strands during the breaking of hydrogen bonding between the bases. Both terms are used to refer to the process as it occurs when a mixture is heated, although “denaturation” can also refer to the separation of DNA strands induced by chemicals like urea. For multiple copies of DNA molecules, the melting temperature (T_m) is defined as the temperature at which half of the DNA strands are in the double-helical state and half are in the “random-coil” states. The melting temperature depends on both the length of the molecule, and the specific nucleotide sequence composition of that molecule.

Applications of DNA Denaturation

The process of DNA denaturation can be used to analyze some aspects of DNA. Because cytosine / guanine base-pairing is generally stronger than adenosine / thymine base-pairing, the amount of cytosine and guanine in a genome (called the “GC content”) can be estimated by measuring the temperature at which the genomic DNA melts. Elevated temperatures are associated with high GC content. GC ratios within a genome are found to be markedly variable. These variations in GC ratio within a genome of higher organisms results in a mosaic like formation with islet regions called isochores. This results in the variations in staining intensity in the chromosomes. The isochores include in them essential protein coding genes, termed housekeeping genes and thus determination of ratio of these specific regions contributes in mapping these essential genesbacterial systematics has recommended use of GC ratios in higher level hierarchical classification. For example, the Actinobacteria are characterised as “high GC-content bacteria”. In Streptomyces coelicolor A3(2), GC content is 72%. The GC-content of Yeast (Saccharomyces cerevisiae) is 38%, and that of another common model organism Thale Cress is 36%. Because of the nature of the genetic code, it is virtually impossible for an organism to have a genome with a GC-content approaching either 0% or 100%. A species with an extremely low GC-content is Plasmodium falciparum (GC% = ~20%), and it is usually common to refer to such examples as being AT-rich instead of GC-poor.

Preparation of Solutions

GTE buffer

50 mM Glucose,

25 mM Tris-Cl,

10 mM EDTA,

pH 8

NaOH/SDS lysis solution

0.2 M NaOH,

1% SDS

Reagents

EDTA

Ethanol, absolute

Isopropanol

Phosphate Buffered Saline (PBS), 1X

Proteinase K

Sodium Acetate, 3 M pH 5.2 (Molecular Biology Grade)

Sodium chloride (NaCl)

Sodium Dodecyl Sulfate (SDS ), 10%

Tris EDTA (TE), pH 7.4

Tris EDTA (TE), pH 8.0

Tris HCl, pH 8.0

Reagents

Chloroform

EDTA, 0.5 M

Ethanol, absolute

Isoamyl alcohol

Phenol

Phosphate Buffered Saline (PBS), 1X

Proteinase K

RNAse A

Sodium dodecyl sulfate (SDS) solution, 10%

Reagents

Chloroform

EDTA, 0.5 M

Ethanol, absolute

Is amyl alcohol

Phenol

Proteinase K

RNase A

Sodium acetate, pH 5.2

Sodium chloride, 5 M
Sodium thiocyanate (NaSCN), 1 M

TE Buffer (Tris-EDTA), pH 7.4

Tween 20

Xylene

DNA replication. The double helix is unwound and each strand acts as a template. Bases are matched to synthesize the new partner strands.

DNA replication, the basis for biological inheritance, is a fundamental process occurring in all living organisms to copy their DNA. This process is “replication” in that each strand of the original double-stranded DNA molecule serves as template for the reproduction of the complementary strand. Therefore, following DNA replication, two identical DNA molecules have been produced from a single double-stranded DNA molecule. Cellular proofreading and error toe-checking mechanisms ensure near perfect fidelity for DNA replication.

In a cell, DNA replication begins at specific locations in the genome, called “origins”.

Unwinding of DNA at the origin, and synthesis of new strands, forms a replication fork. In addition to DNA polymerase, the enzyme that synthesizes the new DNA by adding nucleotides matched to the template strand, a number of other proteins are associated with the fork and assist in the initiation and continuation of DNA synthesis.

Watson and Crick based on their double helical DNA model, offered to explain DNA replication by a method where each single strand of DNA is template or mold for its complement and a new helix has one old strand and one newly synthesized strand. This type of DNA replication is called as semi-conservative as in this method one old strand is retained and one new complementary strand is synthesized. In contrast to semi-conservative mode of DNA replication, two more modes of DNA replication were proposed. One is conservative type where two new strands are synthesized in form of a double helix while the old double helix remains unchanged. The third type of DNA replication proposed has been dispersive mode of DNA replication which amounts to breaking down of double helical strands into small pieces along the length, their replication and random assemblage with newly synthesized pieces to form a patchwork single string of dispersed old and new pieces. These three modes of replication of DNA molecule are shown in Fig. 40.1.

7.1 Basic concept

What is DNA sequencing?

The genome of any biological organism is a sequence of nucleotides arranged in a specific manner. The nucleotides are chemically a base which usually exists in 4 different forms; Purine – Adenine (A) and Guanine (G) and Pyramidine – Cytosine (C) and Thymine (T). These four nucleotides make up an entire sequence of Deoxyribose Nucleic Acid (DNA) which is encodes the information of any cell in the form of a gene. The genome, therefore, is the complete set of all genes that is required to build and maintain that organism. DNA sequencing is the process wherein the precise order of these nucleotides along the gene is detected and analysed which includes any method or technology that assists in determining the sequence.

DNA Sequencing

DNA sequencing is the determination of the precise sequence of nucleotides in a sample of DNA. Sequencing can be done on a short fragment of DNA to cover the entire genome. The maps thus constructed using the nucleotide sequences in the correct order is called as sequence map.

In 1977, twenty-four years after the discovery of the structure of DNA, two separate methods for sequencing DNA were developed: the chain termination method (Sanger et al., 1977) and the chemical degradation method (Maxam and Gilbert, 1977). Both methods were equally popular to begin with, but, for many reasons, the chain termination method is the method more commonly used today. In chemical degradation method, chemicals specific to each base remove the last base, which has been labelled. The method uses toxic chemicals which required high care while handling.

In chain termination method, DNA polymerase extends a DNA strand in which dideoxynucleotide becomes incorporated terminating the chain extension (phosphodiester formation) at the 3’OH site making an array of fragments differing in length. This method is based on the principle that single-stranded DNA molecules that differ in length by just a single nucleotide can be separated from one another using polyacrylamide gel electrophoresis.

DNA can be sequenced in both double stranded and single stranded templates by cloning in plasmid or phage vectors. There are two sequencing methods.

Principle

Determination of the precise sequence of nucleotides in a sample of DNA is called “DNA Sequencing”. The most popular method for doing this is called the “Dideoxy method or Sanger Method” (named after Frederick Sanger, who was awarded the 1980 Nobel Prize in chemistry for inventing it). DNA is synthesized from four deoxynucleotide triphosphates. The top most figure is a normal nucleotide: deoxythymidine triphosphate (dTTP). During polymerization each new nucleotide is added to the 3' -OH group of the last nucleotide added.

The dideoxy method has been coined after the synthetic nucleotides that lack the –OH at the 3’carbon atom of the sugar and it plays an important role in this process atom (red arrow). In the figure below dideoxythymidine triphosphate — ddTTP, a dideoxynucleotide has been shown that can be added to the growing DNA strand. But the chain elongation ceases when the next nucleotide tries to get attached to the former as there is no 3' -OH for the next nucleotide to be attached to. Hence, the dideoxy method is also called the chain termination method. 

Identification of DNA as the Genetic Material

Experiments with pneumococcus suggested that DNA is the genetic material

Hershey and Chase provided evidence that the genetic material injected into the bacterial cytoplasm is T2 phage DNA

RNA functions as the genetic material in some viruses Work Before Watson and Crick

Nucleotides are the building blocks of nucleic acids

Nucleotides are linked together to form a strand

In 1869, Miescher first isolated DNA and named it “nuclein.”

In 1902, Archibald Garrod linked heredity to enzyme (protein) defects. In 1909 he published a book entitled “Inborn Errors of Metabolism.”

In 1928, Griffith identified a “transforming factor” that transmitted infectiousness in bacteria.

In 1944, Avery, MacLeod and McCarty showed that the transforming factor was DNA)

In 1950, Hershey and Chase, using their famous “blender” experiment, confirmed that DNA, not protein, is the genetic material.

The work of chemist Phoebus Levene showed that DNA includes deoxyribose, nitrogenous bases, and phosphates, in equal proportions.

Chargaff s data showed that the number of purines equals the number of pyrimidines.

Using Rosalind Franklin’s X-ray diffraction patterns, Watson and Crick deduced that DNA is a double helix

In 1868, Johann Friedrich Miescher, a young Swish medical student got excited with an acidic substance that he isolated from pus cells obtained from bandages used to dress human wounds. To obtain large quantities of pus for his experiment, Miescher began purchasing used bandages from surgical clinics in Tuebingen, Germany. He separated pus cells from bandages and treated the cells with pepsin, a proteolytic enzyme that he isolated from stomach of pigs. After pepsin treatment, he recovered an acidic substance that was named as “nuclein” by him. This nuclein was unusual in the sense that it was rich in nitrogen and phosphorous, two elements known at that time to coexist only in certain types of fat. Miescher’s paper on this investigation was published in 1871. At that time, importance of Miescher’s nuclein was not fully appreciated. Even the existence of polynucleotide chains, the key component of the acidic material in Miescher’s nuclein was not documented until 1940s. The role of nucleic acid in storing and transmitting genetic information remained un-established until 1944 and the double-helix structure of DNA was not discovered until 1953 by James Watson and Francis Crick. Even in 1953, many geneticists remained skeptical to accept the role of nucleic acid as the genetic information carrier rather than proteins as nucleic acids exhibited less structural variability than the proteins (Snustad et al., 1997).

A laboratory is essential for research work in science and development. Most of the research institutes have dedicated laboratories for researchers and students to conduct experiments, perform research work, as well as to provide hands-on learning/training to students. There are certain rules and regulations laid down by the laboratory in-charge which need to be followed strictly for safety purposes. Some of the laboratories keep ‘safety manual’ for researchers to read before entering into the laboratory to conduct experiments. Whenever safety is compromised in a laboratory, there is increase in risk of unwanted incidents in the laboratory. Storing food/beverages in the same refrigerator where laboratory chemicals are kept may lead to disastrous effects on health. This chapter aims to provide instructions in brief on the things to be performed (Do’s) before and after entering onto a laboratory as well as the things never to be done (Don’ts) in a laboratory for the safety reasons.

Good Laboratory Practices (Do’s)

• P Wear apron/labcoat while working in the laboratory.
• P Use gloves/respiratory mask/fumehood while handling toxic/volatile/inflammable chemicals/reagents.
• P Use pippet or pippetor for safety in transfer of solutions.
• P Protect eyes, face and naked parts of the body from UV radiations. Use safety goggles or face mask/apron.
• P Keep your eyes away from Liquid Nitrogen and use cryogloves while handling it.
• P Arrange all the necessary materials/solutions before starting the experiment.
• P Periodically fumigate/ozonize culture and inoculation rooms.
• P Autoclave all contaminated materials before discarding.
• P Pre-treat gel or solution containing Ethidium Bromide (EtBr) before discarding into laboratory waste-disposal bin, and use gloves while handling them.
• P Burn all the transgenic plant materials in an incinerator after taking necessary observations.
• P Make entry in the designated log-book and clean the instrument after using it properly.
• P Switch-off the instrument after use or when not in use.
• P Wash hands with soap and remove the labcoat before leaving/going out of the laboratory7.
• P Ensure that switches for fans/lights, and water taps are off while leaving the laboratory.
• P Follow the biosafety guidelines suggested for handling and disposal of the hazardous chemical.

INTRODUCTION

Ethno-entomophagy is the term used to describe the practice of consumption of insects as food by native human populations. Insects are an important source of protein for humans, either through direct consumption or as food supplements for livestock (DeFoliart, 1989). They are nutritious in terms of protein (40–75 g/100g dry weight) and minerals. Their protein is of high quality and has high digestibility (77–98%) with good concentration of essential amino acids (46–96% of the nutritional profile) (Verkerk et al., 2007). The biochemical composition and chemical analysis have revealed that they are comparable with other foods enriched with high proteins, fats, lipids, vitamins and mineral salts (Prasad, 2007). Insects generally have higher food conversion efficiency than other animals. They show a food conversion efficiency of about twice as efficient as pig and boiler chick and more than five times as efficient as beef cattle (Durst and Shono, 2010). Some of the important edible insects include grasshoppers, caterpillars, beetle grubs, adult winged termites, bees, wasps, winged ants, cicadas and a variety of aquatic insects.

1. Introduction

Phytochemists and pharmacologists from all over the world obtain several patents for herbal drugs developed on the basis of the indigenous knowledge systems of different ethnic groups. Many countries rich in traditional medicinal knowledge have been conducting ethnobotanical survey of medicinal plants (Patil and Bhaskar, 2006). In India the native people have exploited a variety of herbal medicines for effective curing of various diseases. The plants used, preparation and administration of drug varies from area to area (Rajaram, 2006). The ethnobotanical research still plays its evident scientific role in stimulating further phytochemical and pharmacological studies (Leporatti and Corradi, 2001).

The objective of this study is to interact with Kani traditional healers and document their traditional knowledge. The Kani-tribals are one of the most primitive hill tribes of Western Ghats. They occupy the slopes of the Agasthiarmalai in Tirunelveli, Tamil Nadu, India. Some ethnobotanical studies have been done in this area (Anonymous, 1948-76, Chopara et al., 1956, Satyavati, 1976, Jain, 1981, Nagendra Prasad et al., 1996). The impact of deforestation, urbanization and modernization makes the tribals shifting from their natural habitats to urban areas. Documenting the traditional knowledge through ethnobotanical studies is important for the conservation of medicinal plants and traditional medicines.

A. Answer the following questions

1.    Why protein requirement for maintenance in cat is high?

Ans: It is due to the inability of the nitrogen catabolic enzymes in the cat’s liver to adapt to changes in dietary protein intake.

1.    Why arginine is dietary essential in cat?

Ans: Because, cat is unable to synthesize ornithine which is an arginine precursorwithin theurea cycle. Aminoacid glutamic acid and proline is precursor of ornithine in the intestinal mucosa of all mammals.However, an enzyme thatis required in the pathway, proline-5-carboxylate synthatase is absent in cat’s intestinal mucosa. In the absence of dietary arginine, a high catabolism of protein generates a load of ammonia that cannot be detoxified by its incorporation to urea and hyper ammonia develops.

2.    Why taurine is dietary essential in cat?

The full name of an organism technically has eight terms. For the dog it is: Eukarya, Animalia, Chordata, Mammalia, Carnivora, Canidae, Canis, and lupus. Notice that each name is capitalized except for species, and the genus and species names are italicized. Scientists generally refer to an organism only by its genus and species, which is its two-word scientific name, or binomial nomenclature. Therefore, the scientific name of the dog is Canis lupus. The name at each level is also a taxonomy Carnivora is the name of the taxon at the order level; Canidae is the taxon at the family level, and so forth. Organisms also have a common name that people typically use, in this case, dog. Note that the dog is additionally a subspecies: the “familiaris” in Canis lupus familiaris. Subspecies are members of the same species that are capable of mating and reproducing viable offspring, but they are separate subspecies due to geographic or behavioural isolation or other factors. Dogs, which are under the species Canis lupus familiaris, are known to have a total of 78 chromosomes (2n). The diploid genes can be classified into 38 pairs with two sex chromosomes.

Dolichos bean is known as Lablab bean, hyacinth bean or sem. Grown as field crop in MP, Maharashtra, AP and TN. Genus Dolichus is renamed as Lablab (Syn. D. lablab, D. purpureus, Lablab niger, L. bvulgaris) and Lablab purpureus name is widely accepted. Dolichos lablab L. (synonyms Lablab purpureus (L.) Sweet.) is leguminous pod vegetable grown in tropics and few temperate countries. Lablab niger var. lablab is synonym to Dolichos lablab var. typicus where long axis of seed is parallel to suture and is vegetable type. Lablab niger var. lignosus synonym to Dolichos lablab var. lignosus where long axis of seed is perpendicular to suture and is pulse type.

Dolichos bean, [Lablab purpureus (L.) Sweet] var. typicus (2n=22) also known as Indian bean or sem is one of the vital leguminous vegetables grown in India for its tender green pods. Both pole types and bush types are commercially grown. It is predominantly cultivated in Maharashtra, Tamil Nadu, Andhra Pradesh, Uttar Pradesh and North-East India. Protein, minerals, vitamins and f ibers are abundant in Dolichos bean. Bush-type vegetable dolichos unlike the pole-type dolichos (which are generally grown during the winter season as they require short day) are thermosensitive and can be grown year-round.

Dolichos is a versatile and valuable leguminous crop, offering multiple benefits. It boasts a high protein content of 3.6% and fiber content of 1.8%. The dry seed is composed of a protein content ranging from 23.0% to 28.0%. The pods are abundant in phenols, with levels ranging from 1.7 to 9.67 mg/100 g, making them a valuable source of antioxidants. Furthermore, dolichos serves as an excellent source of amino acids, particularly lysine, along with vitamins such as A, C, and riboflavin, and essential minerals (Ca, Fe, Mg, S, Na and P). The seeds of dolichos contain a flavonoid, kievitone, recognized for its potential in combating breast cancer. Additionally, the presence of tyrosinase in the seeds indicates promising possibilities for treating hypertension in humans. Dolichos beans are utilized for various medicinal purposes, acting as stomachic, anthelmintic, diuretic, aphrodisiac, antispasmodic, digestive, febrifuge, and carminative agents. Furthermore, they are rich in bioactive compounds that offer significant benefits in treating specific ailments such as liver problems, diabetes and tumors.

Physical ergonomics is concerned with human anatomical, anthropometric, physiological and biomechanical characteristics as they relate to physical activity. This is the ergonomics domain we are most concerned with in the workplace, and most of the content on this site is very much focused on workplace ergonomics.Physical ergonomic principles have been widely used in the design of both consumer and industrial products for optimizing performance and to preventing / treating work-related disorders by reducing the mechanisms behind mechanically induced acute and chronic musculoskeletal injuries / disorders.

There are about 6.5 million coconut farmers in the country. Coconut production, processing, marketing and other connected activities support the livelihood of over 12 million people. Coconut crop provides natural health drink, nutritious edible products and oil for edible and non edible uses, fibre of commercial value, shell and wood for fuel and industrial uses, and a variety of miscellaneous products of utility. Coconut is a symbol of national integration and prosperity. The crop is considered reverential cutting across linguistic and religious barriers promoting national integration. Contribution of coconut to the GDP of the country is around Rs. 100,000 million.

Domestication of Livestock

Domestication (from Latin domesticus) is the process whereby a  population  of  animals  or plants is changed at the genetic level through a process of selection, in order to accentuate traits that benefit humans. It differs from taming in that a change in the phenotypical expression and genotype of the animal occurs, whereas taming is simply the process by which animals become accustomed to human presence. In the Convention on Biological Diversity, a domesticated species is defined as a “species in which the evolutionary process has been influenced by humans to meet their needs.” Therefore, a defining characteristic of domestication is artificial selection by humans. Humans have brought these populations under their control and care for a wide range of reasons: to produce food or valuable commodities (such as wool, cotton, or silk), for types of work (such as transportation, protection, and warfare), scientific research, or simply to enjoy as companions or ornaments. Plants domesticated primarily for aesthetic enjoyment in and around the home are usually called house plants or ornamentals, while those domesticated for large-scale food production are generally called crops. A distinction can be made between those domesticated plants that have been deliberately altered or selected for special desirable characteristics and those plants that are used for human benefit, but are essentially no different from the wild populations of the species. Animals domesticated for home companionship are usually called pets while those domesticated for food or work are called livestock or farm animals.

In literal sense a species is called underutilized whose potential has not been properly realized. These underutilized crops (referred to also by other terms such as minor, orphan, neglected, underutilized, underexploited, underdeveloped, lost, new, novel, promising, alternative, local, traditional, niche crops) have been included in world-wide plans of action after having successfully raised the interest of decision makers. Wealth of plant genetic diversity currently available holds vast potential. We use only small number of crop plants for our basic requirements. Many thousands of wild plants have great economic and cultural importance and tremendous market potential for vast numbers of people throughout the world. But the majority are still unknown to science. Plant genetic resources have gained global attention in  recent past by agricultural and environmental institutions. Conservation and use of genetic resources attracted  considerable attention. Value of biological diversity in economic terms is very important for human society.

Abstract

Pomegranate, Punica granatum L., is an ancient fruit tree native to the Middle East. All plant parts are rich source of the various secondary metabolites that accounts for its antioxidant and health-promoting activity. A strong genetic control on anthocyanin production is responsible for variability in the color of peel and arils of pomegranate. Characterization, cloning and expression of the genes related with anthocyanin and tannin production provides routes for engineering of these complex pathways. Recent studies on pomegranate genome sequencing have elucidated several biochemical pathways and candidate genes that could assist to increase production of potent bioactive compounds in near future. This will further facilitate our understanding to some exclusive biological processes that plays a significant role in pomegranate breeding.

Donkeys can also be termed as ass (Equus africanus asinus) and is a domesticated member of the horse family, Equidae with chromosome number 62. The Wild ass of Africa, E. africanus is the wild ancestor of the donkey. For approximately 5000-7000 years, the donkey has been used as a working animal.

An adult male donkey is a jack or jackass, an adult female is a jenny or jennet, and an immature donkey of either sex is a foal. Jacks are often mated with female horses (mares) to produce mules, the less common hybrid of a male horse (stallion) and jenny is called hinny. NBAGR-ICAR has registered three breeds of donkey till date.

1. Kachchhi

• These donkeys are found in Kachchh district of Gujarat.

• Coat colour is mainly grey (dorsal surface grey and ventral surface white) followed by white, brown and black.

• Forehead is convex. Nasal bone is straight.

• Height at wither ranges from 77 to 115 cm.

• Docile in temperament.

• Only donkeys are used for agricultural purposes like inter cultivation for weed removing.

• Also utilized for transportation as donkey cart, as pack animal during pastoralist migration, etc.

Abstract

The milk from non-bovine species is gaining considerable attention mainly due to the fact that it is consumed by sensitive consumers (infants and elderly). Donkey milk possesses various health benefits, namely, anti-microbial activity, immunomodulation, hypoallergenicity, prevention of cardiovascular diseases which are elaborated in details here. Similarity in nutrition profile among donkey and human milk is discussed in detail. The donkey milk’s yield, its milking technique, effect of lactation on chemical composition and the microbial aspect is also described. As, availability of donkey milk is limited, therefore its processing becomes essential for its usage over a long period of time. Preparation of various donkey milk products like yoghurt, ice cream and cheese are also described here.

Doppler ultrasound is a non-invasive diagnostic tool being employed in evaluating blood flow in major vessels, veins and arteries. Information regarding perfusion in an organ or an area can also be obtained through Doppler ultrasonography. Doppler ultrasound is an important part of echocardiographic examination in dogs and cats. Its more recent application is in the investigation of the tissue wall motion while evaluating the heart. B-mode ultrasound is used to examine cardiac structures.

7.1 Measurement of Radiation Dose

Radiation dosimetry involves the measurement of the energy actually absorbed in a substrate exposed to ionizing radiation. The radiation may be either electromagnetic or corpuscular in nature. The passage of electromagnetic radiation through a substrate causes ionization by virtue of the secondary electrons released. Particle radiations on the other hand, ionize directly because they are charged, or indirectly through the charged particle resulting from a collision with a substrate. Alpha, beta particles, and protons yield primary ionization. Gamma rays and neutrons produce ionizing radiation by their interaction with the substrate in passage through it.