Essential amino acids

 Essential or indispensable amino acids :- 

                                                                                                              The amino acids which can not be synthesized by the body and, therefore, need to be supplied through the diet are called essential amino acids. They are required for proper growth and maintenance of the individual. The ten amino acids listed below are essential for humans,

1. Arginine, 
2. Valine, 
3. Histidine, 
4. Isoleucine, 
5. Leucine, 
6. Lysine, 
7. Methionine,
8. Phenylalanine, 
9. Threonine, 
10. Tryptophan,

Nonessential amino acids

Nonessential Amino Acid or dispensable amino acids:- 

 The body can synthesize about 10 amino acids to meet the biological needs, hence they need not be consumed in the diet. These are:-

1. Glycine, 
2. Alanine, 
3. Serine,
4. Cysteine, 
5. Aspartate, 
6. Asparagine, 
7. Glutamate, 
8. Glutamine, 
9. Tyrosine and
10.  Proline,

Recombinant DNA technology

 Recombinant DNA technology:-

Basic Functions of a Fermenter

 Basic Functions of a Fermenter:-

• It should provide a controlled environment of optimum biomass yields.
• It should permit aseptic fermentation for a number of days reliably and dependably, and meet the requirements of containment regulations Containment involves prevention of escape of viable cells from a fermenter or a downstream processing equipment into the environment.
• It should provide adequate mixing and aeration for optimum growth and production, Without damaging the microorganisms.
• The power consumption should be minimum,
• It should provide easy and dependable temperature control.
• Facility for sampling should be provided.
• It should have a system for monitoring and regulating pH of the fermentation broth.
• Evaporation losses should be as low as possible.
• It should require a minimum of labour in maintenance, cleaning, Operating and harvesting operations.
• It should be suitable for a range of fermentation processes. But this range may often be restricted by containment regulations.
• It should have smooth production internal surfaces, and joints should be welded wherever possible.
• The pilot scale and production stage fermenters should have similar geometry to facilitate scale-up.
• It should be contrasted using the cheapest materials that afford satisfactory results.
• There should be adequate service provisions for individual plants.

Ti-Plasmid

                                                           Introduction

 Agrobacterium tumefaciens is a rod shaped, gram negative Bacteria, soil born motile bacterium. This bacterium infects the parts of the plant which are in contact with soil. The genus Agrobacterium inclides four species of bacteria. They are A.tumefaciens, A.radiobacter, A.rubi, A.rhizogens. They  are pathogenic in nature and cause cancerous growth in the infected plant tissues. They infect the plants and cause tumberous growth in the infected portion. The induction of tumerous growth is due to the presence of a large sized plasmid named Ti Plasmid. Accourding to Zaenen et al. (1974) almost all the strain of A.tumefaciens contain Ti plasmid.

Agrobacterium is a bacterium which is used to produce transgenic plants because it contain tow kinds of vectors which are used in the transfer of genes of one plant to the genome of another plant:-

 1. Ti plasmid, 

 2. Ri plasmid,

                                                              Structure  

T-DNA carry genes for phytohormones, AND opine

Auxin:- The main function of auixn is to help plant grow. Auxin stimulates plant cell to elongate, and the apical meristem of a plant is one of the main places where auxin is produced.

Cytokinin:- Increased cell division by stimulating the process of mitosis. Increased formation of shoot and buds, as well as development of fruits and seeds.

Opin:- Nutritional source for agrobacteria and fall into different chemical grouping such as nopaline, Octopine, Mannopine, agrocinopine etc.

Vir genes:-

 Vir A:- Its a kinase portein in bacterial membrane. Act as receptor Phenolic compund relased by wounded plant.

Vir A and Vir G :- They expessed continuously respond to phenolic compund released by wounded plant.

Vir G :- Activate other vir gene (transcription gene factor)

Vir E/E2 :- Protect T-DNA against nuclease and target T-DNA to plant cell. Act as SSB protein (single standard binding protein).

Vir C :- Stimulate transfer/promotes high effciency T-stand Synthesis.

Vir D :- Responsible for virulence activity of bacterium.

Vir D1:- Topoisomerase helps virD2 to recognise and cleave within 25bp L.B sequence.

 Vir D2 :- Endonuclease ( vir D2 cut T-DNA right border, help to integration of T-DNA into host chromosome)

Vir D1 :- Prevent attack of exonuclease at 5' end of T-DNA, important for cutting phosphodiester bond.

Vir B/ Vir D4 :- ATP dependant linkage of  protein complex necessary for T-DNA transfer from bacterium of plant cell ( assemble into a secreation system which spns the inner and outer bacterial membrane.)

Vir D2/Vir E2 :- Have nuclear localization signal.

Vir F:- Plant cell cytosole, vib1protein.

                                                               Advantage

• Occure naturally in Bacteria.
• Have different restriction sites.
• Replication completely independent of Bacteria.
• Genes are easily inserted into plasmids.
• Easily transformed into bacteria.
  
  

what is bioinformatics

What is Bioinformatics:-

Translation of billions of characters in DNA sequences that make the genome into biologically meaningful information has given birth to a new field of science called "bioinformatics". The term bioinformatics has been derived by combining biology and information.

                          The key to biotechnology discoveries is locked in the genomes of organisms. The bioinformatics holds the key to unlock these data for the next generation of innovations.

intro germplam conservation

 

Germplasm Conservation:-

Germplasm broadly refers to the hereditary material transmitted to the offspring through germ cells.

Germplasm provides the raw material for the breeder to develop various crops. Thus, conservation of germplasm assumes significance in all breeding programmes.

As the primitive man learnt about the utility of plants for food and shelter, he cultivated the habit of saving selected seeds or vegetative propagules from one season to the next one. In other words, this may be regarded as primitive but conventional germplasm preservation and management, which is highly valuable in breeding programmes.

The very objective of germplasm conservation is to preserve the genetic diversity of a particular plant or genetic stock for its use at any time in future. In recent years, many new plant species with desired and improved characteristics have started replacing the primitive and conventionally used agricultural plants. It is important to conserve the endangered plants or else some of the valuable genetic traits present in the primitive plants may be lost.

A global body namely International Board of Plant Genetic Resources (IBPGR) has been established for germplasm conservation. Its main objective is to provide necessary support for collection, conservation and utilization of plant genetic resources throughout the world.

Cryobiology

 Cryobiology deals with the study of metabolic activates and their responses in plant materials and animal cells stored at low temperature (-196'C) by using liquid nitrogen in the presence of cryoprotectants. 

Fungal Cells

 1. Yeast:- Yeast is an unicellular eukaryotic fungus containing a small well characterized genome. Unlike plant or animal cells, it has rather fast growth rate and itself is a non-pathogenic fungus. Most of its gene contain introns which are spliced during purification of mature mRNA. It appears that intron found in yeast contain sequences for correct splicing as they are totally absent in higher eukaryotes. Moreover, yeast can carry out post-translational modification such as removal of signal sequence from a precursor polypeptide after the secretion of cell. This reveals a major advantage of yeasts over the bacteria. Success in DNA cloning in yeast depends on uptake of foreign DNA by its spheroplast in the presence of calcium ion and polyethyleneglycol (PEG). The spheroplasts develop cell wall after the incorporation of DNA.

Properties of Good Host

 A good host have the following features:-

1. Should be easy transform,                                                                                                          
2. Should support the replication of recombinant DNA,                                                                     
3. Should be free from elements that interfere with replication of recombinant DNA,                         
4. Lack active restriction enzyme, e.g. E.coli K12 sub strain HB 101,                                   
5. Should not have Methylases since these enzyme would methylate the replicated recombinant DNA which, as a result, would become resistant to useful restriction enzymes,                                                                                                                              
6. Should be deficient in normal recombination function so that the DNA insert is not altered by recombination events.                                                                                                                                                                                                                                     E.coli support several type of  vectors, some natural, constructed, which can be grouped as follows:-                                                                                                                                                                                                                                                    1. Plasmid                                                                                                                          2. Bacteriophages (both natural)                                                                                      3. Cosmids,                                                                                                                        4. Phasmids                                                                                                                      5. Shuttle vectors,                                                                                                            6. Artificial chromosomes                                                                                                  7. Phagemids,                                

Types of Restriction Endonucleases

There are the following four distinct types of restriction endonucleases:-

Type 1st restriction endonucleases:- are complex endonucleases, and have recognition sequences of about 15 bp; they cleave the DNA about 1000 bp away from the 5'-end of the sequence "TCA" located within the recognition site, e.g, EcoK, EcoB, etc.

Type 2nd restriction endonucleases:- are remarkably stable and induce cleavage either, in most cases, than 350 different type 2nd endonucleases with over 100 different recognition sequences are known. They require Mg⁺² ions for cleavage. The first type 2nd enzyme to be isolated was HindII in 1970. Only type 2nd restriction endonucleases are used for restriction mapping and gene cloning in view of their cleavage only at specific sites.

Type 3rd restriction endonucleases:- are intermediate between the type 1st and 2nd enzymes; they cleave DNA in the immediate vicinity of their recognition sites, e.g, EcoP1, EcoP15, HindIII, ect. Type 1st and 3rd restriction enzyme are not used in gene cloning. The type 3rd enzymes recognize asymmetric target sites, and cleave the DNA duplex on one side of the recognition sequence up to 20 bp away.

Steps in GENE CLONING

Steps in Gene Cloning:-

The entire procedure of gene cloning or recombinant DNA technology may be classified into the following five steps the convenience in description and on the basis of the chief activity performed.

1. Production and isolation of the DNA fragments to cloned.                     
2. Insertion of the isolated gene in a suitable vector to obtain recombinant DNA.                                                                                  
3. Introduction of the recombinant DNA into a suitable organism/cell usually we used E. coli  called host(transformation).                                                                                 
4. Selection of the transformed host cells, and identification of the clone containing the desired gene/DNA fragment.                                   
5. Multiplication/expression of the introduced gene in the host.                    
6. Where needed, transfer and expression of the gene in the another organism.                                                                                                                                   

Autoimmune Disease

 Autoimmune Disease:-                                                                                       The immune system normally produces antibodies against g foreign proteins but not against the native proteins of body, that is, the immune system can distinguish between "self" and "non-self", However, in rare cases, individuals begin to produce antibodies against their own antigens. These antibodies are called autoantibodies and the diseases resulting from their presence are the autoimmune diseases. Among these diseases are paroxysmal cold haemoglobinuria, myasthenia gravis and systemic lupus erythematous. 

structure of antibody

 Structure of antibody:-

What is sterilization and Types of sterilization

 Sterilization:-

All the materials, e.g., vessels, instruments, medium, plant material, etc., used in culture work must be freed from microbes. This is achieved by one of the following approaches:

1. Dry heat,

2. Flame sterilization,

3. Autoclaving,

4. Filter sterilization,

5. Wiping with 70% ethanol,

6. Surface sterilization,

                                                            1.Dry Heat

Glassware and Teflon plastic ware, and instruments may be sterilized by dry heat an oven at 160-180°C for 3 hours. But most workers prefer to autoclave glassware and plastic ware etc. And flame sterilize instruments like forceps, etc. More recently, glass bead sterilizers (300°C) are being employed for the sterilization of forceps, scalpels, etc. these devices use dry heat.

                                     

                                                2.Flame Sterilization

Instrumental like forceps, scalpels, needles, etc. Are ordinary flame sterilized by dipping them in 95% alcohol followed by flaming. These instruments are repeatedly sterilized during the operation to avoid contamination. It is customary to flame the mouth of culture vessels prior to inoculation.

                                                  3.Autoclaving

Culture vessels, etc. Are generally sterilized by heating in an autoclave or a pressure cooker to 121°C at 15p.s.i. (pounds per square inch 1.06kg/cm²) for 15 to 40 minutes, The time being longer for larger medium volumes. Sterilization during autoclaving depends mainly on temperature. Certain types of plastic ware and some instruments, e.g., micropipettes, etc. Are also autoclave. Care should be taken to properly stopper all the vessels and to open the autoclave only its pressure gauge indicates zero pressure. 

                                                4. Filter Sterilization

Some growth regulators, e.g., GA3, zeatin, ABA (abscisic acid), urea, certain vitamins, and enzymes are heat labile. Such compounds are filter sterilized by passing their solution through a membrane filter of 0.45μ or lower pore size. The membrane filter is held in a suitable assembly, the  assembly together with the filter is sterilized by autoclaving before use. Filter sterilized heat labile compounds are added to autoclaved and cooled media, in case of agar medium, they are added when the medium has cooled to about 40°C and is still liquefied.

Laminar flow cabinets are used to create an aseptic working space by blowing filter-sterilized air through an enclosed space. The air is first filtered through a coarse profiteer to remove large particles, it is then passed through a HEPA (high efficiency particulate air) filter, which filters out all particles larger than 0.3μm. This sterilized air blows through the cabinet at 1.8 km/hr. which is sufficient to keep the enclosed working area aseptic. 

        

                               5. Wiping with 70% Ethanol 

The surfaces that can not be sterilized by other techniques, e.g., platform of the laminar flow cabinet, hands of the operator, are sterilized by wiping them thoroughly with 70% ethyl alcohol is allowed to dry.

                                    6. Surface Sterilization

 All the plant materials to be used for culture are treated with an appropriate sterilizing agent to inactive the microbes present on their surface, This is called surface sterilization. Surface sterilization protocol will depend mainly on the source and the type of tissue of the explant is the excised piece of tissue or organ used for culture.

The sterilizing agent used for surface disinfection are calcium hypochlorite (9-10%), sodium hypochlorite (2%), mercuric chloride (0.1-1%), silver nitrate (1%), Bromine water (1-2%), H2O2 (10-12%) and antibiotics (4-50 mg/l). Of these, calcium or sodium hypochlorite and HgCl2 (satisfactory results) are the most commonly used. The duration of treatment varies form 15-30min. Since these agents are also toxic to plant tissues, the duration and the concentration used should be such as to cause minimum tissue death, and the rinsing after treatment should remove them as completely as possible.

Thanks For reading............................,

Definition of Biotechnology

             Definition of Biotechnology,

Biotechnology  consists of "the controlled use of biological agents, Such as, micro-organisms or cellular components, for beneficial use."                                                                                                                                                        -U.S National Science Foundation,

Biotechnology is "the integrated use of of biochemistry, microbiology And engineering science in order to achieve technological application of the capabilities of micro-organisms, cultured tissue/cells and parts thereof."                                                                                                                                                                        -European Federation of Biotechnology, 

Biotechnology comprises the "controlled and deliberate application of simple biological agents living or dead, cells or cell components-in technically useful operations, either of productive manufacture or as service operation."                                                                                                                                                          -J.D. Bu'lock,1987,

"The application of biological organisms, systems or processes" constitutes biotechnology.                                                                                     -British Biotechnologist,

Biotechnology may be defined as "the use of living organisms in system or processes for the manufacture of useful products: it may involve algae, bacteria, fungi, yeast, cells, of higher plants and animals or subsystems of any of these or isolated components from living matter."                                                                                                                                                                                                                           -Gibbs and Greenhalgh, 1983,

Amis Of Plant Tissue Culture

1. Development of disease free plant from disease plants.                                                          
2. Reducing the time it takes for the reproductive cycle to complete.                                          
3. Fixed enrichment of haploid plants.                                                                                             
4. Creating a new plant from body hybridization and production cells.                                    
5. To develop strain resistance plants.                                                                                 
6. Cultivation of transgenic plants.                                                                                             
7. To produce economically important plants in large numbers in a short time.                           
8. It does not allow the process of sexual reproduction to be necessary for crop development ensures the growth of new plants from any plant.                                        
9. Creating unusual hybrids, Such as those with protoplast conjugation with new trails develop.