Indian Journal of Biotechnology
VOLUME 1
NUMBER 1
JANUARY 2002
CONTENTS
Editorial
7
Papers
Developments in biotechnology: an overview
Rintu Banerjee, Gargi Mukherjee, Ashok Pandey & A Sabu
9
Some aspects of biotechnology relevant in the national context
H C Chaturvedi & Savita Agnihotri
17
Biotechnology input in fish breeding
Samir Bhattacharya , Subrata Dasgupta, Malabika Datta & Dipanjan Basu
Heralding the dawn of cultured adult stem cell transplantation
Geeta Kashyap Vemuganti & D Balasubramanian
29
Therapeutic monoclonal antibodies: attractions and challenges
Navin Khanna
50
Apoptosis: an overview
Rana Anjum & Ashok Khar
58
Virus resistant transgenic plants for environmentally safe management of viral diseases
A Varma, R K Jain & A I Bhat
73
Electrostatic interactions, phase separation behaviour and partitioning of proteins in
polyelectrolyte based aqueous two-phase systems
Vandana Gupta, Sunil Nath & Subhash Chand
Development of genetic markers specific for highly endangered species, Rhinoceros
unicornis: strategies and implications in conservation
Vipra Kapur, Anu Bashamboo & Sher Ali
87
39
96
Bioinformatics: advancing biotechnology through information technology Part I:
molecular biology databases
Sudeshna Adak & Biplav Srivastava
101
Biomolecular engineering of Escherichia coli organomercurial lyase gene and its expression
Imtiyaz Murtaza, Amit Dutt & Arif Ali
117
Biotechnology—the interface between agri resources and the consumer
V Prakash
121
Instructions to Contributors
126
Indian Journal of Biotechnology
Vol 1, January 2002, pp 9-16
Developments in Biotechnology: An Overview
Rintu Banerjee, Gargi Mukherjee, Ashok Pandey and A Sabu
Human quest for improving the natural capabilities of microorganisms, making them capable of
novel processes, and discovering microorganisms with new capabilities has led to tremendous
development, today referred to as modern biotechnological advances. This has led to the development of
recombinant DNA technology, which allows modifying microbes and other organisms to create in them
highly valuable, novel and naturally non-existent capabilities. R & D in areas such as stem cell, human
genome, transgenic crops, etc. has made untreatable diseases to understand and control. However,
research in many areas such as cloning, stem cell, etc. has also posed serious concern on ethical and
societal issues, which if not taken proper measures, could be eventually very dangerous. This paper traces
the significant biotechnology developments and their impact on human life with a futuristic approach.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 17-28
Some Aspects of Biotechnology Relevant in the National Context
H C Chaturvedi and Savita Agnihotri
Biotechnology encompasses all such plant processes, which may have practical or commercial
application, albeit only temporarily a distinction may be made between fundamental and applied
biotechnological research. But in all cases, priorities must be drawn for biotechnological research in the
context of a developing nation, like, India with practicable objectives either realizable immediately or in
the near future for economic growth of the country as well as protection and improvement of human
health, putting the promise research on the back burner. Some of the aspects of Biotechnology, which may
be brought to fruition with the already existing technologies and resources having down-to-earth
objectives, are identified here. 1. Boosting citrus industry: In the face of the rich heritage of citrus
germplasm, vast orchard area and suitable agroclimates for commercially important Citrus species, the
Citrus Industry can be boosted with the application of a recent breakthrough of meristem culture if not
some other techniques also. 2. Micropropagation of forest trees for reforestation: It is estimated that the
green cover in the country is reduced to more than 55%, resulting in change of agroclimates, loss of
biodiversity and many other related evil effects besides, shortage of wood. Micropropagation of forest
trees, which is the only process to produce requisite number of plants, at a rapid rate, in the shortest
possible period of time. 3. Standardization of herbal drugs: There exists a world herbal market worth $ 50
billion, in which Indian share is hardly 0.1% despite India being the home of herbal traditional drugs of
Ayurveda. One of the main reasons for this is non -standard herbal formulations and gross
adulteration of scientifically unidentified plant materials, in which context biotechnology can
provide solutions. 4. Germplasm preservation for conservation of phytodiversity: Conservation
of biodiversity, precisely phytodiversity for the survival of the human race is imperative. And
the role of in vitro strategies for germplasm preservation and establishment of Gene Banks or
Germplasm Repositories cannot be overemphasized. 5. Bioremediation and purification of
sewage: The huge quantity of sewage is discharged everyday in the rivers, which constitute a
major source for drinking water and poses a threat to the human health. As practiced by
advanced countries, remediation of sewage by efficient biotechnological processes using
bacteria and a few algae should be done and the purified sewage can be used for irrigation
purpose, while the remaining quantity may be discharged in rivers minimising the health risk.
6. Production of bioethanol fuel: In view of the acute shortage of petrol, production of
bioethanol to be used as fuel in automobiles is not only unavoidable, but also feasible by
utilizing sugarcane as a raw material; sufficient quantity of sugarcane crop can be produced
through well-established procedure for its micropropagation.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 29-38
Biotechnology Input in Fish Breeding
Samir Bhattacharya, Subrata Dasgupta, Malabika Datta and Dipanjan Basu
Pisciculture, to increase the fish production, has a crucial limitation. Fish cultured in land-locked
water bodies usually do not breed without the hormonal induction. Technology of fish breeding suffered
for a long time due to the lack of suitable commercial product to induce the spawning of economically
important fish. Hypophysation technique, where pituitary extract was used to induce the breeding, faced
more failures than success besides the limitation of the source. After the discovery of a brain peptide,
gonadotropin releasing hormone (GnRH), which binds to the membrane receptor of pituitary gonadotroph
cell and releases gonadotropic hormone (GTH), the situation has changed dramatically. Final maturation
and release of germ cells, spermatozoa and oocyte, to the water for fertilization, depends on an acute surge
of GTH which is about 10 times greater than the normal circulatory level of GTH. Injection of GnRH
causes this acute GTH surge. GnRH is a decapeptide but its gene encodes 92 amino acid containing large
molecule. Extensive post-translational processing is necessary to secrete GnRH from the neural cells. For
this reason recombinant DNA technology could not be employed for the production of GnRH. Instead,
chemical synthesis of this decapeptide is easier and cheaper. Numbers of fish GnRH structure have been
elucidated and depending on salmon GnRH peptide sequence, a chemical analogue has now been
marketed under the the name of “Ovaprim”. There is a strong research background to understand GnRH
mechanism of action and signal transduction pathway involved in GnRH mediated GTH function on germ
cell maturation and release. These scientific investigations have contributed significantly in designing the
superactive GnRH analogues. The only GnRH available in India is from the brain of a freshwater murrel,
Channa punctatus. Combination of two murrel GnRH variants, GnRH I and GnRH II, produces far more
superior effects than “Ovaprim”. Biotechnology input in fish breeding is no doubt highly appreciable as it
provides the cultivators almost a riskless method. However, research in different laboratories is still in
progress to have a more potent molecule with the possible addition of some metabolic hormones.
Pisciculture is now a booming industry all over the world, which imposes a larger demand for “Ovaprim”
like products. Murrel GnRH, therefore is expected to be a highly competitive product in the global
market.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 39-49
Heralding the Dawn of Cultured Adult Stem Cell Transplantation
Geeta Kashyap Vemuganti and D Balasubramanian
Over the last few years, research on stem cells has received much public attention both for its extraordinary
potential and for the associated social, legal and public issues. One of the fundamental questions in developmental
biology is: how does a single cell-the fertilized cell give rise to a complex, multicellular organism and what are the
factors involved during this remarkable development? With that began the quest to define stem cells, identify and
isolate the embryonic, fetal and adult stem cells. Then came the question of whether these stem cells derived from
various sources can be used to determine the pharmacokinetics of drugs and toxins; and ultimately be used to replace
cells in diseased tissues. These questions were answered by series of experiments on animals followed by clinical trials
using cultured adult stem cells, since the latter evoke the least debate on the ethical and legal fronts. Apart from the
clinical use of hematopoietic stem cells, the other adult stem cells that have been tested and put to clinical use include:
myoblasts for cardiac and skeletal muscle loss or degeneration; cultured beta cells for diabetes mellitus, and cultured
limbal epithelial cells for reconstruction of ocular surface in limbal stem cell deficiency. This review briefly describes
the concept of stem cells, the regulatory mechanisms that govern them, with special emphasis on stem cells in epithelial
tissues and the technique of culturing limbal stem cells that has been developed at our institute.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 50-57
Therapeutic Monoclonal Antibodies: Attractions and Challenges
Navin Khanna
Due to the technological advances made during the past decade, therapeutic monoclonal antibodies
(MAbs) now represent an important and growing class of bio-therapeutics. Sales of MAbs are increasing
rapidly from virtually nil in 1996 to potentially over $ 1 billion in 2001. With the potential new targets
resulting from genomics and with methods now in place to make fully human antibodies, the potential of
antibodies as valuable therapeutics in oncology, inflammation and cardiovascular disease is being fully
realised. There are 235 MAbs under different stages of development. More than 100 clinical trials are in
progress, 14 of them in phase III (mostly for cancer diagnosis and therapy). To meet the increased demand
of the monoclonals, worldwide manufacturing capacity for MAbs must increase dramatically, during the
next few years, if the industry segment is to avoid a manufacturing bottleneck. Many novel expression
strategies are being evaluated to increase the production capacity. Factors including molecular fidelity and
cost of products will be critical in the selection of the expression systems.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 58-72
Apoptosis: An Overview
Rana Anjum and Ashok Khar
Apoptosis, a highly ordered cascade of enzymatic events that culminates in cell death and entails the
autolytic degradation of cellular components, is characterized by blebbing of cell membrane, nuclear and
cytoplasmic condensation, protein fragmentation and DNA degradation followed by rapid engulfment of
cell corpses by neighbouring cells. It is essential for maintenance of cellular homeostasis and deregulation
of this process leads to a spectrum of pathological manifestations. Caspases, which form the proteolytic
network within the cell are among the critical components of cell death process. They bring about the
cleavage and degradation of a number of proteins that result in cell demise. Mitochondria are regarded as
the central control point in the execution of apoptosis. They release a potent cocktail of pro-apoptotic
proteins into the cytosol, which results in the amplification of the death cascade, the most prominent one
being cytochrome c. The mechanism by which cytochrome c is released into the cytosol is controversial
but seems to be regulated by the Bcl-2 family of proteins.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 73-86
Virus Resistant Transgenic Plants for Environmentally Safe
Management of Viral Diseases
A Varma, R K Jain and A I Bhat
Plant viruses are one of the major yield reducing factors for agricultural and horticultural crops. In
India, most destructive diseases are caused by gemini-, poty-, and tospoviruses. Virus resistant transgenic
plants (VRTPs), developed by the transfer of transgenes from virus, plant or other origins, have been
found resistant to a wide range of viruses. The most successful approach is the viral coat protein mediated
resistance (CPMR). Other transgenes of viral origin, which have shown promise are: replicase protein,
movement protein, proteases, and antisense sequences. ‘R’-genes from plants, plantibodies and yeast
RNase genes are also useful for developing VRTPs. In a large number of VRTPs developed using
transgenes of viral origin, resistance is conferred by post-transcriptional gene silencing (PTGS); in some
cases PTGS has been overcome when plants are infected by a heterologous virus, indicating need for
cautious approach. Overall, the bio-safety concerns in the use of VRTPs get insignificant, but these must
be addressed scientifically. In India, initiatives have been taken for developing VRTPs to manage
important plant viral diseases. The present world area under VRTPs is about 0.4 mha. Judging from the
success of various strategies, the area under VRTPs is expected to grow at a fast rate in the coming years.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 87-95
Electrostatic Interactions, Phase Separation Behaviour and Partitioning
of Proteins in Polyelectrolyte Based Aqueous Two-Phase Systems
Vandana Gupta, Sunil Nath and Subhash Chand
Electrostatic interactions play a major role in the purification of proteins by different methods and they tend to be
specific in nature in the presence of salts and environmental hydrogen ion concentration (pH). These can be readily
exploited as the basis for protein isolation and purification by aqueous two-phase system (ATPS) using a
polyelectrolyte as one of the polymeric components. Polyelectrolytes are water-soluble charged polymers and the phase
separation in the polyelectrolyte based ATPS is dependent on the ionic composition of the system and the charge
density of polymers. This review outlines the mechanism of the phase separation in non-ionic polymers based ATPS on
the basis of water structure in polymeric solution and validity of the hypothesis is further discussed for polyelectrolytebased systems. Partitioning of proteins in polyeletrolyte based ATPS is dominantly controlled by the electrostatic
interactions. Several factors that influence the partitioning of proteins include the properties of the polymers, pH, salt
type and concentration and the charge on protein molecules. Low polymer concentration requirement for the phase
separation and high specificity of protein purification in polyelectrolyte based ATPS holds a great promise for their
large scale applications in isolation and purification of proteins.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 96-100
Development of Genetic Markers Specific for Highly Endangered Species
Rhinoceros unicornis: Strategies and Implications in Conservation
Vipra Kapur, Anu Bashamboo and Sher Ali
Indian rhino, Rhinoceros unicornis, is a highly endangered species and unless collective efforts are
made for its protection, the future of this species will continue to remain bleak. Application of restriction
survey and minisatellite associated sequence amplification (MASA) for identification, cloning and
sequencing of the potential genomic fragments from this species, led to the development of species
specific DNA marker. These markers are envisaged to be useful for ascertaining the origin of rhino's
biological samples in the event of poaching and illegal trafficking. This approach may be used for
conservation and management of other endangered species.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 101-116
Bioinformatics: Advancing Biotechnology through Information Technology Part I:
Molecular Biology Databases
Sudeshna Adak and Biplav Srivastava
This paper is intended as a review of molecular biology databases and other Bioinformatics resources
available for biotechnologists aiming to use the wealth of genomic data available today. The genomic data
along with associated proteomic and functional data are often distributed across multiple databases,
requiring a time-consuming search by the user. The explosion of information seen in molecular biology
has created a veritable maze, through which careful navigation is required for research and innovation in
biotechnology. The paper, one of the series, introduces the readers to the major molecular biology
databases and bioinformatics tools such as BLAST for similarity searching and RasMol for protein
structure visualization. Subsequent papers will take the readers into a journey across bioinformatics and
the biotechnological discoveries that are happening with bioinformatics. Advances in computer
technologies and the birth of the internet are also part of this revolution in biology. Online databases have
given scientists and researchers across the world access to unimaginable volumes of biologically relevant
data. Bioinformatics, a truly multidisciplinary science, aims to use the benefits of computer technologies
in understanding the biology of life itself.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 117-120
Biomolecular Engineering of Escherichia coli Organo-mercurial
Lyase Gene and its Expression
Imtiyaz Murtaza, Amit Dutt and Arif Ali
Studies were carried out to characterize merB gene from five wild type strains of broad-spectrum
Escherichia coli, collected from five geographically distinct regions of India. Each strain produced 23kb
plasmid from which functional merB gene (0.64kb) was PCR amplified. The merB gene from isolate G18,
which tolerated highest concentration of organic form (PMA) of mercury was cloned in high expression
vector pQE30 and pGEMT-Easy vector. The transformants obtained demonstrated varied results in their
appropriate hosts. The transformants (IAxpress) carrying merB gene cloned in pQE30 and negative
control having pQE30 without merB insert did not grow on agar plates amended with 1g/ml PMA. Due
to the hyperexpression of merB in pQE30 most of the protein was found in nonfunctional inclusion bodies
and did not show any resistance as sensitive strain (Devoid of merB gene) against PMA. On the other
hand transformants of merB cloned pGEMT vector tolerated up to 5g/ml of PMA, which indicates that
low expression of merB in this vector produces a functional product and thus tolerates five times more
PMA than sensitive strain. The results demonstrate that this gene can be better exploited for
bioremediation of toxic form of mercury in polluted water bodies.
Indian Journal of Biotechnology
Vol 1, January 2002, pp 121-125
Biotechnology—The Interface between Agri-Resources and the
Consumer
V Prakash
April
July
October
Indian Journal of Biotechnology, started as a quarterly journal in 2002, publishes full
papers, short communications and reviews in agricultural-, animal-, environmental-,
industrial-, medical-,and microbial biotechnology, bioinformatics, and socio-legal and
ethical aspects in biotechnology.The latest developments in biotech-industry are covered
under notes and news.