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Questions
Name and explain the technique used for separating DNA fragments and making them available for biotechnology experiments.
Name and describe the technique that helps in the separation and isolation of DNA fragments.
Solution
Gel electrophoresis is a technique which is used for separating DNA fragments in a gel on the basis of their electric charge and size.
The process of gel electrophoresis is as follows:
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The fragments obtained after cutting with restriction enzymes are separated by using gel electrophoresis.
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Electric field is applied to the agarose gel matrix and the negatively-charged DNA fragments move towards the anode.
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Fragments separate according to their size by the sieving properties of agarose gel. The smaller the fragment, the farther it moves.
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Staining dyes such as ethidium bromide, upon exposure to UV radiation, are used to visualise the DNA fragments.
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DNA fragments are visible as bright orange-coloured bands in the agarose matrix.
- These bands are cut from the agarose gel and extracted from the gel piece (elution).
- DNA fragments are purified and used in constructing recombinant DNAs
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Match the Column I (Restriction enzyme) with Column II (Recognition sequence) and select the correct option.
| column I (Restriction Enzyme) | column II (Recognition sequence) | ||
| i. | Alu | P. | 5'---G- ↓-G-A-T-C-C---3' 3'---C-C-T-A-G- -G---5 |
| ii. | BamHI | Q. | 5'--- A-G- ↓ -C-T---3' 3' ---T-C- G-A--- 5' |
| iii. | Eco RI | R. | 5'--G-T-C- ↓ -G-A-C--3' I 3'--C-A-G-t-C-T-G--5' |
| iv. | HindII | S. | 5'---G- ↓ -A-A-T-T-C---3' 3'---C-T-T-A-A- -G---5' |
The following identifies the major features of technology that differentiate modem biotechnology from classical biotechnology.
i. Capability of science to change the genetic material for getting new specific products through rDNA technology, polymerase chain reaction (PCR), microarrays, cell culture and fusion, and bioprocessing.
ii. Ownership of technology and its sociopolitical impact.
iii. Modem biotechnology makes use of only fermentation technology.
iv. Classical biotechnology relies only on principles of plant and animal tissue culture.
Following are the four statements regarding separation of DNA fragments using Gel electrophoresis. Identify the INCORRECT statements.
i. DNA is negatively charged molecule and so it is loaded on gel towards the Anode terminal.
ii. DNA fragments travel along the surface of the gel whose concentration does not affect movement of DNA.
iii. Smaller the size of DNA fragment, larger is the distance it travels through it.
iv. Pure DNA can be visualized directly by exposing to UV radiation.
The vector Baculovirus is used for gene transfer in ____________.
The gene transfer between unrelated plants is brought about by ____________.
Which of the following viruses has the capacity to produce lysozymes?
Name the plant disease caused by Agrobacterium tumefaciens.
To be a cloning vector, a plasmid require ______.
The structure below shows pUC18, which is similar to pBR322 in its function. However, they differ in some of their restriction sites and the number of ori. The ori number for pBR322 is approximately 20.
- How are puc18 and pBR322 used in biotechnological studies?
OR
What will be the impact if the ori in the above structure gets damaged? - The lac z gene has many recognition sites. Study the segment of DNA given below and answer the questions.
5’... ATC GTA AAG CTT CAT…3’
3’... TAG CAT TTC GAA GTA…5’
i) Applying your knowledge of palindrome sequences identify and mark the possible region where the restriction enzyme X will act.
ii) Restriction enzyme Y was used to extract a gene of interest from a plant. This gene needs to be inserted in the given DNA segment which has been treated with restriction enzyme X. Will there be a successful recombination? Explain with a reason. - Which one of the two (pUC18 and pBR322) would you prefer for biotechnological studies? Justify.
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Give a reason for the following:
DNA cannot enter directly into the host cell.
