Question 10 of 17advanced💡 UnderstandLong Answer5 marks

How are recombinants screened? Describe the methods in detail.

Correct Answer

Recombinants are screened based on the principle of difference in biological traits present in hosts with recombinant DNA from those without recombinant DNA. The selection procedure can be divided into two main types:

Direct selection involves distinguishing transformed cells from non-transformed cells based on expression of certain traits. For example, bacterial cells that take up plasmids containing antibiotic resistance genes become resistant to that specific antibiotic. These cells survive and grow in media containing the antibiotic, whereas host cells without plasmids are killed when exposed to antibiotics.

Insertional inactivation is more efficient than direct selection. In this method, a vector having two markers (either two antibiotic resistance genes, or one antibiotic resistance gene and one lacZ gene) is used. When the gene of interest is inserted into one of the selection marker genes in the vector, its expression is disrupted, hence called insertional inactivation.

Blue-white selection is another example of insertional inactivation where the expression of lacZ gene can be directly observed in bacterial colonies to select recombinant transformed cells.

Blotting techniques are also used to separate and identify DNA, RNA and proteins. Southern blotting technique detects specific sequences of DNA in DNA samples.

Exercise: EXERCISES | Q: 12 | (Page 39)
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Explanation

The answer covers all screening methods mentioned in the textbook context: (1) Direct selection using antibiotic resistance, (2) Insertional inactivation using vectors with two markers, (3) Blue-white selection using lacZ gene, and (4) Blotting techniques including Southern blotting. The context explicitly states these methods under section 3.5 and the bullet points. The principle of selection based on difference in biological traits (antibiotic resistance, β-galactosidase expression, GFP, or nutritional requirements) is also included from the context.

Solution Steps

  1. Step 1: State the principle of recombinant selection based on biological trait differences

  2. Step 2: Describe direct selection method using antibiotic resistance

  3. Step 3: Explain insertional inactivation method with two markers

  4. Step 4: Describe blue-white selection as an example of insertional inactivation

  5. Step 5: Mention blotting techniques for DNA, RNA and protein identification

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