Genetic Engineering and Genomics Notes - MH-CET 2015 by ednexa.com

1. Genetic Engineering and Genomics
Introduction -
∙ Genes are the blueprint of life of all organisms and viruses because they have
conserved
the genetic traits since million years and will do so in future too.
∙ The last two decades of 20th
Century witnessed much advancement in the areas
of biology
particularly molecular biology.
∙ Desired genes of one organism can be cut and joined to other genes and
introduced into
the cells of organisms.
∙ Moreover, techniques have been developed to screen and detect the new
inserted DNA.
∙ This is a branch of biology called as genetic engineering.
∙ Using this technology several novel products can be produced.
DNA Fingerprinting -
∙ It is curious fact that each human individual has a separate identity which is due
to the
presence of his specific genetic make up.
∙ Though we inherit parental genes, we differ a lot from them mainly due to the
recombinations of maternal and paternal genes
∙ Differences also arise due to infrequent mutations which occur during cell
division.
∙ If we trace the differences between two individuals at molecular level, the
differences lie in
the nucleotide sequences of their DNA.
∙ DNA fingerprinting – is a technique employed to assist in the identification of
individuals on
the basis of their respective DNA profiles.
∙ DNA fingerprinting is also known as DNA profiling or DNA typing.
∙ The technique was invented in 1977 by a British geneticist Alec Jeffreys
∙ Genome of a person contains about three billion base pairs arranged in a definite
sequence.

2. ∙ Technique of DNA fingerprinting is mainly based on the fact that DNA of One
individual is
about 90% identical to the DNA of another individual.
∙ Remaining 10% DNA is different and is important for DNA fingerprinting.
∙ This 10% DNA is generally the non-coding region with simple tandem repeats of
nucleotide
sequences (eg: AT TCGAT TCGAT TCG…) called as VNTRs (Variable Number
Tandem
repeats)
∙ These repeats are inherited from the parents, and are used as genetic markers in
a
personal identity test.
Sources of DNA for DNA fingerprinting:-
∙ As the DNA content of a person is the same for every cell, tissue and Organ, the
DNA
sample for fingerprinting may be obtained from traces of blood, semen, urine,
saliva,
vaginal swab, tissue bits, or a single hair.
Procedure for DNA fingerprinting:-
(1) DNA from above mentioned sources is obtained and isolated by techniques such
as High-
speed refrigerated centrifugation.
(2) If the quantity of DNA obtained is small, it is subjected to in Vitro (Outside the
body)
replication by a technique called PCR (Polymerase Chain Reaction) to obtain
more copies of DNA
(3) DNA molecules are cleaved i.e. cut into fragments at specific sites with the help
of
Restriction endonucleases (RE). This makes many pieces of DNA having
variable length. This phenomenon is called as “Restriction Fragment Length
Polymorphism” (RFLP),
(4) The restricted DNA fragments (Sample DNA) are loaded on agarose gel slab for
electrophoresis. Current is applied due to which the fragments which are
negatively charged will move towards positive pole. The movement depends
upon the length of these fragments resulting in the formation of bands.
(5) Double stranded DNA are now split into single strands by using alkaline
chemicals.

3. (6) The isolated DNA fragments in the gel are copied (blotted) onto nitrocellulose
sheet placed
on the gel. This technique is called as Southern blotting after its inventor E.M.
Southern.
(7) Special DNA probes are prepared in the laboratory, these contain repeated
sequences of
nucleotides complementary to those on sample DNA. These probes are made
radioactive.
(8) The radioactive probes bind to the Sample DNA fragments on the nylon sheet.
This is
called hybridization.
(9) An X-ray film is exposed to the nylon sheet to mark the places where the
radioactive DNA
probes have hybridized with Sample DNA. These places are marked as Dark
bands when
X-ray film is developed. This process is called as autoradiography.
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