Biology 5 Marks Questions

The bacterium Thermus aquaticus is employed and used for amplification of the gene of interest using PCR technique. Usually Taq (Thermus aquaticus) DNA polymerase, a thermostable enzyme is isolated from a thermophilic bacterium. The enzyme extends the two primers towards each other in order to copy the DNA segment (act as a template) lying between the two primers.
The step requires the presence of deoxynucleoside triphosphates and Mg2+ and occurs at 72°C.
If these cycles are repeated many times, the DNA segment can be amplified to approximately a billion times the DNA segment are made.
i. These are restriction endonucleases enzymes which cut the DNA molecule at the specific base sequences into fragments with sticky ends.
e.g., Eco RI, Hind II
ii. The enzyme restriction endonuclease cleaves DNA at a specific site resulting in the formation of fragments with single strand portions at the ends called sticky-ends. In practice, the digestion by the restriction enzyme keeping all other conditions at the optimum level and checked by using agarose gel electrophoresis technique.

i. They worked with bacteriophage, i.e. viruses that infect bacteria. These viruses were used because during infection they transfer their genetic material into bacteria.
ii. They used two types of culture media, containing 35S and 32P, so as to compare that which one out of DNA and proteins gets transferred from virus to bacteria and act as genetic material.
iii. A blender and centrifuge were used to open up the bacterial cells and viral particles, so, that genetic material could be visualised.
iv. They concluded that DNA is the genetic material that is passed from virus to bacteria.

a.

b. The development of an endosperm precede that of the embryo in angiosperm because the cells get packed with reserve food supplies, used for providing the nutrition to the developing embryo.
c. Cells with 16 chromosomes is called zygote and cells with 24 chromosome is called endosperm.

i. Amplification of recombinant DNA gene is done using Polymerase Chain Reaction (PCR).
It is carried out in the following steps:
a. Denaturation -The double-stranded DNA is denatured by applying high temperature of 95°C for 15 seconds. Each separated strand acts as a template.
b. Annealing -Two sets of primers are added, which anneal to the 3'end of each separated strand.
c. Extension- DNA polymerase extends the primers by adding nucleotides complementary to the template provided in the reaction. Taq polymerase is used in the reaction, which can tolerate heat. All these steps are repeated many times to get several copies of the desired DNA,
ii. The DNA polymerase used in PCR is Taq polymerase extracted from Thermus aquaticus. It is a thermostable enzyme that can withstand the high temperature used in the denaturation and separation of DNA strands. Hence, it can be used for a number of cycles of DNA amplification without being denatured.

i. Satellite DNA or repetitive DNA These sequences normally do not code for any proteins. These sequence show high degree of polymorphism.
ii. Steps carried out in the process of DNA fingerprinting technique are:
(a) isolation of DNA,
(b) digestion of DNA by restriction endonucleases
(c) separation of DNA fragments by electrophoresis
(d) transferring (blotting) of separated DNA fragments to synthetic membranes such as nitrocellulose or nylon.
(e) hybridization on using labelled VNTR probe.
(f) detection of hybridized DNA fragments by autoradiography
The applications of DNA fingerprinting technique are in Forensic science / determining population and genetic diversities / paternity test.