Difficulty: Easy
Correct Answer: Low temperature and Ca++
Explanation:
Introduction / Context:
Conventional chemical transformation uses divalent cations to render E. coli competent to take up plasmid DNA. Understanding the temperature and ion conditions during the binding stage explains why a subsequent heat shock is required.
Given Data / Assumptions:
Concept / Approach:
At low temperature (ice), Ca2+ neutralizes negative charges on DNA and lipopolysaccharides, allowing plasmid DNA to adhere to the cell envelope. The system is then briefly heat-shocked to create a thermal imbalance that drives DNA uptake through transient pores.
Step-by-Step Solution:
1) Competency induction: cells are incubated in cold CaCl2 to stabilize membranes and screen charges.2) DNA addition: plasmid is mixed with cells while kept on ice, maximizing surface binding.3) Heat shock: a short high-temperature pulse promotes DNA entry; this comes after, not during, the binding step.4) Therefore, the interaction environment is low temperature with Ca++ present.
Verification / Alternative check:
Standard protocols (Mandel–Higa, Hanahan) specify cold CaCl2 (often with additional salts) for binding, followed by heat shock; Mg2+ alone is not the defining ion in the classic version.
Why Other Options Are Wrong:
High temperature and Ca++: heat is used later for shock, not during initial interaction.High temperature and Mg++ / Low temperature and Mg++: Mg2+ may be present in some recipes, but Ca2+ at low temperature is characteristic of the binding step.
Common Pitfalls:
Confusing the binding step with heat shock, or assuming Mg2+ is the key ion for the conventional method.
Final Answer:
Low temperature and Ca++
Discussion & Comments