Question

The cloning site present in the ampicillin resistance gene of \( \textit{E. coli}\) cloning vector pBR322 is :

• A. \( BamH I \)
• B. \( EcoR I \)
• C. \( Pst I \)
• D. \( Sal I \)

Hint:

Remember the locations of key restriction sites in pBR322 relative to the antibiotic resistance genes. Pst I, Pvu I, and Sca I are within \(amp^R\), while BamH I, Sal I, Xho I, and Cla I are within \(tet^R\). EcoR I is outside these resistance genes.

Answer 1

The Correct Option is C

The plasmid pBR322 is a widely used \textit{E. coli} cloning vector. It contains two antibiotic resistance genes: the ampicillin resistance gene (\(amp^R\)) and the tetracycline resistance gene (\(tet^R\)). It also has several unique restriction enzyme recognition sites where foreign DNA can be inserted for cloning. The cloning sites within the ampicillin resistance gene of pBR322 include Pst I, Pvu I, and Sca I. Insertion of foreign DNA into any of these sites will disrupt the ampicillin resistance gene, a phenomenon known as insertional inactivation. This allows for the selection of recombinant plasmids (plasmids containing the inserted DNA) because bacteria carrying these plasmids will be resistant to tetracycline but sensitive to ampicillin. The cloning sites within the tetracycline resistance gene of pBR322 include BamH I, Sal I, Xho I, and Cla I. Insertion of foreign DNA into these sites will disrupt tetracycline resistance. EcoR I is a unique restriction site in pBR322 but it is located outside both antibiotic resistance genes, in the region of the origin of replication (\(ori\)). Therefore, the cloning site present in the ampicillin resistance gene of \textit{E. coli} cloning vector pBR322 among the given options is Pst I.