Biotechnology: Principles and processes
Biotechnology Principles and Processes Class 12 notes:
What
is biotechnology?
● The European Federation of
Biotechnology (EFB) characterizes biotechnology as "the joining of
inherent science and creatures, cells, parts thereof and atomic analogs for
items and administrations".
Basis
of Modern Biotechnology
● Genetic engineering − Introduction of unfamiliar hereditary
material (DNA/RNA) into the host's genome and modifying its aggregate
● Aseptic techniques − Involves support of contamination-free feeling
in substance designing cycles for assembling of items like anti-infection
agents, antibodies, etc. This is done as such as to empower the development of
just wanted organisms answerable for a bioprocess.
Biotechnology Principles and processes Class 12 notes
for NEET
Genetic
Engineering
● Asexual reproduction saves
the hereditary data while sexual multiplication jam varieties.
● Plant and creature
hybridization methodology regularly bring about presentation of unfortunate
qualities alongside advantageous ones.
● Genetic engineering conquers
this constraint.
● Genetic engineering incorporates:
1.
Creation of recombinant DNA
2.
Gene cloning
3.
Gene transfer into host creature
● The presented piece of DNA doesn't repeat in
the host except if it is coordinated with the chromosome of host.
● For getting replicated, the
unfamiliar DNA should coordinate into the host DNA arrangement having
'beginning of replication'. At the point when this joining happens, unfamiliar
DNA is repeated and many duplicates are shaped. This cycle is called cloning (the course of arrangement of
different indistinguishable duplicates of DNA).
Biotechnology Principles and Processes Class 12 notes:
Development
of a Recombinant DNA
● Plasmid (independently duplicating,
roundabout, extra chromosomal DNA) is segregated.
● Plasmid DNA acts as a vector since it is utilized to move the
piece of DNA appended to it to the host.
● Plasmid DNA additionally contains qualities
answerable for giving anti-infection protection from the microscopic organisms.
● Plasmid DNA was cut with a
particular restriction catalyst ('atomic scissors' − that cut a DNA at specific
locations).
● The DNA of interest (to be embedded) was
likewise cut with a similar restriction compound.
● The DNA of interest is
hybridized with the plasmid with the assistance of DNA ligase to frame a Recombinant DNA.
● Recombinant DNA is then moved to a host like
E.coli, where it imitates by utilizing the host's duplicating hardware.
● When E.coli is refined in a
medium containing anti-toxin, just cells containing recombinant DNA will
actually want to make due because of anti-toxin obstruction qualities and one
will actually want to disconnect the recombinants.
Biotechnology: Principles and processes NCERT
Restriction
Enzymes as Tools of RDT
● Restriction proteins are
specific chemicals that perceive and cut a specific succession of DNA.
● Nucleases are of two kinds:
o
Endonucleases − Cut the DNA at explicit
situations inside the DNA
o
Exonucleases − Cut the DNA at the closures
(Remove the nucleotides at the finishes of the DNA)
● Every restriction enzyme recognizes
various arrangements (Recognition successions). More than 900 restriction proteins
have been segregated, all of which perceive various successions.
● Recognition groupings are palindromic - Palindromes are the
succession of base combines that read same both backward and forwards (i.e., same and bearing).
● Restriction compounds remove
a little from the focal point of palindrome site, however between similar two
bases on the contrary strands.
● Thus, overhangs (called sticky
ends) are produced on each strand.
● Sticky closures structure
hydrogen bonds with their integral partners with assistance of DNA ligases.
● All these cycles structure
the premise of RDT.
● Naming restriction enzyme
o
I st letter − Genus of the life form from which
the enzyme is derived
o
II nd and III rd letters − Species of the organism
o
IV th letter − Name of the strain
o
Roman number − Order of seclusion E.g., In
EcoRI − Derived from E.coli, strain R. It is the I st to be found.
Gel
Electrophoresis
● The sections acquired in the
wake of cutting with restriction compounds are isolated by utilizing gel
electrophoresis.
● Electric field is applied to
the electrophoresis grid (normally agarose gel) and adversely charged DNA parts
move towards the anode.
● Fragments separate as per their size by the
sieving properties of agarose gel. More modest the section, farther it moves.
● Staining colors, for
example, ethidium bromide followed
by openness to UV radiations are utilized to picture the DNA parts.
● DNA sections are noticeable
as dazzling orange shaded groups in the agarose grid.
● These bands are cut from the agarose gel and
extricated from the gel piece (elution).
● DNA sections are refined and
these sanitized DNA pieces are utilized in building recombinant DNAs.
NCERT Biotechnology Book Class 12
Cloning vectors and host as apparatuses of RDT
Cloning
Vectors
● Plasmids and bacteriophages
are usually utilized as cloning vectors.
● Both of these can recreate
inside the bacterial cells free of the chromosomal DNA.
● Bacteriophages − Have high duplicate number (of genome) inside the
bacterial cell
● Plasmids − May have 1 − 2 duplicate number to 15 − 100 duplicate
number for every cell
● If unfamiliar DNA is
connected to these vectors, then, at that point, it is increased to the number
equivalent to the duplicate number of vector.
● Features present in the
actual vector help in the simple separation of recombinants from the
non-recombinants.
Parts
of a plasmid cloning vector
●
Origin of replication (ori)
○ Replication begins from ori.
Any part of DNA when connected to ori can be made to recreate.
○ With the assistance of this,
the genetic engineer might control duplicate number of the recombinant DNA. To
recuperate a big number, appropriate beginning of replication should be picked.
● Selectable marker
○ These qualities help to
choose recombinants over non-recombinants.
○ Antibiotic opposition
qualities like amp R (ampicillin resistant), tet R (tetracycline resistant)
fill in as selectable markers normally.
Biotechnology Principles and Processes Class 12 notes:
● Cloning sites
○ These locales allude to the
acknowledgment locales for restriction compounds (like EcoRI, Hind III, PvuI ,
BamHI, and so on)
○ These are the sites where restriction
catalysts cut the DNA.
○ Cloning process becomes
finished when more than one acknowledgment sites are available.
○ Therefore, ligation is done
distinctly at the restriction sites present on the antibiotic resistance genes.
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How
anti-infection opposition qualities help in choosing recombinants?
● Suppose tet R quality has
Bam HI acknowledgment site.
● When BamHI is utilized for restriction,
foreign DNA section is embedded inside the tet R gene.
● Hence, tetracycline. obstruction is absent in the
recombinants.
● Recombinants will develop on
the media containing ampicillin, yet will kick the bucket on media containing tetracycline..
● On the other hand, non-recombinants
will develop on medium containing ampicillin just as on medium containing tetracycline.
● In this way, antibiotic resistance gene
helps in choosing transformants.
Substitute
selectable marker
● Other than anti-infection
obstruction qualities, elective markers can be utilized.
● One of them is quality
coding for a^- galactosidase.
● When unfamiliar quality is
embedded inside a^galactosidase quality, the catalyst a^¬galactosidase gets
inactivated (insertional inactivation)
Then the microscopic organisms
are become on a chromogenic substrate.
● Non-recombinants will deliver blue-colored
provinces.
● Recombinants will deliver
dry settlements.
Biotechnology Principles and Processes Notes PDF
Cloning
vectors for plants and creatures
● Ti plasmid (tumor-inducing
plasmid) alludes to the plasmid of Agrobacterium tumefaciens.
o
A. tumefaciens is a plant microorganism. It
produces growths in the plants it taints.
o
Ti plasmid can be adjusted into a cloning
vector by eliminating the qualities liable for pathogenicity.
●
Retrovirus − These are the infections that contaminate creatures. They
produce malignant growths in creatures.
o
Retroviruses can be incapacitated to be
utilized as a cloning vector.
competent
host
● Competent host alludes to
the bacterial cells that can take up the vector (containing Recombinant DNA).
● Methods to bring recombinant DNA into
capable host:
o
Cells are treated with divalent cations (for
example Ca 2+ ). Then, at that point, these cells are hatched with recombinant
DNA on ice, trailed by heat shock (at 42º), and afterward setting them back
aside momentarily. By this, microbes can take up recombinant DNA.
○ Microinjection−
Recombinant DNA is straightforwardly infused into the core of creature cell.
○ Biolistics (Gene Gun)− Cells are assaulted with high speed miniature
particles of gold or tungsten.
○ Disarmed vector as if there should be an
occurrence of A. tumefaciens and retrovirus.
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Biotechnology Principles and Processes Class 12 notes:
Processes
of RDT
Isolation
of Genetic Material (DNA)
● For the cycles of RDT, DNA
should be accessible in its unadulterated structure.
● First of all, cells are
treated with explicit synthetic compounds to tear open the cell to deliver cell
parts like DNA, RNA, proteins, and so on This is finished by chemicals like
lysozymes (bacterial cell), cellulase (plant cell), and chitinase (parasitic
cell).
● Contaminants, for example,
RNA and proteins are processed with the assistance of ribonucleases and
proteases separately.
● Addition of chilled ethanol
eventually hastens out the refined DNA, which can be viewed as assortment of
fine strings in the suspension.
Cutting
of DNA at Specific Locations
● DNA is cut into pieces with
the assistance of restriction proteins.
● Fragments produced after restriction
are detached with the assistance of gel electrophoresis.
● Recombinant DNA is acquired by hybridizing
'quality of interest' with vector, with the assistance of enzyme DNA ligase.
Polymerase
Chain Reaction (PCR)
● Recombinant DNA can be
enhanced by PCR. A few indistinguishable duplicates of it tends to be combined
in vitro.
● Two arrangements of primers (chemically combined
oligonucleotide extends that are integral to a locale of DNA), enzyme DNA polymerase,
and deoxynucleotides are added.
● PCR comprises of 3 stages:
Denaturation−
Double helical DNA is denatured by giving high temperature. DNA polymerase
doesn't get corrupted in such high temperatures since the DNA polymerase
utilized in this response is thermostable as it is disconnected from
thermophilic microscopic organisms, Thermus aquatics.
Extension −
Replication of DNA happens in vitro.
This cycle is rehashed a few
times to produce up to 1 billion indistinguishable duplicates of the DNA.
Addition
of Recombinant DNA into Competent Cells
● Insertion of recombinant DNA
into host is done by a few techniques:
○ Transformation if there should be an
occurrence of microbes
○ Disarmed vectors,
biolistics, and micro¬injections in the event of plant and creature cells
○ The cells bearing recombinant DNA are chosen
on the grounds that the recombinants only have selectable marker present in
them (like anti-toxin obstruction).
Acquiring
the Foreign Gene Product
● This is the stage for which
the recombinant DNA was created.
● The cell containing recombinant DNA will
deliver a clever protein item (positive item/Recombinant protein).
● For enormous scope creation
of the advantageous item (anti-toxins, immunizations, chemicals), ideal
conditions are to be given.
● Continuous culture − Used
culture media is depleted from one side and new culture media is added from the
opposite side.
o
Cells are kept all through in their
log/outstanding stage.
o
Larger biomass is delivered by this strategy
prompting better return.
● Bioreactors − Large vessels in which
enormous volumes (100 − 1000 liters) of culture can be created
○ Optimal development conditions for
microorganisms are available (temperature, pH, substrate, salts, vitamins
nutrients, and so on)
○ A bioreactor has the accompanying parts ¬
fomenter framework, oxygen conveyance framework, froth control framework,
temperature and pH control framework, testing ports.
Biotechnology Principles and Processes Class 12 notes:
● Biosynthesis of many
mixtures like proteins, alcohols, and anti-microbials occur inside the
bioreactor.
● The items so acquired are unrefined and
require partition, cleaning, and completing, which is done under downstream
handling (DSP).
● DSP makes an unrefined bio item attractive.
● After legitimate division and purging,
additives are added and the completed item is made to go through clinical
preliminaries and quality checks prior to being shipped off market.