Question

2. (3pts) Describe the lysing process that occurs when we perform the ""slow freeze-quick thaw"" procedure for preparing a crude extract of rGFP?

Answer 1

Answer:

Slow freeze-quick thaw is lysis protocol used widely for mammalian and bacterial cells. It works by dehydrating the sample through crystal formation.

Explanation:

Slow Freeze - Quick Thaw:

• A cell lysis procedure that involves slowly freezing cells in dry ice or ethanol or even liquid nitrogen (-196°C) and then quickly thawing them at room temperature.
• Slow freezing involves cooling below freezing point to form ice crystals in the extracellular environment. As water around the cell freezes, the presence of high concentration of solutes (salt and sugar) and cryoprotective agents induces a hypertonic environment outside the cell. This forces water outside the cell, consequently dehydrating it. Crystals also form in the cell wall, disrupting the cellular transport.
• Upon quick thawing, as the extracellular environment turns hypotonic, water rushes into the cells which swell up and burst due to osmotic shock.