Nucleic Acid and Protein Sample Preparation
Standing on the shoulders of giants, Fred Sanger received the Nobel Prize for Chemistry twice: in 1958 for sequencing the protein insulin and in 1980 for his contribution to the sequencing of DNA. Sample preparation - preparing nucleic acids or proteins for analysis - played a vital role in both achievements.
Without effective sample preparation, there is a risk that incorrect conclusions will be drawn. For example, important mRNA transcripts that have been degraded along the way will be missed, and damaged enzymes will give incorrect kinetics data.
Methods in sample preparation are generally based on properties such as: differential solubility in solvents, precipitation, adsorption to solid-phase, highly selective purification by affinity, or separation by size or charge. There are specific methods for isolation from individual samples of nucleic acids and proteins, and even both. Sample preparation may also be necessary between downstream steps, to remove reaction components that would inhibit or disturb the next step.
The main aim is to prepare a product that meets the requirements of the most demanding of the downstream methods you will be using. This may mean obtaining the optimal purity, retaining molecular structure, biological activity, or relative levels of all biomolecules of interest. The method must remove contaminants that degrade the biomolecule of interest, interfere with immediate downstream analysis, or threaten long-term sample storage. The method chosen will always be a compromise between various factors such as effectiveness, time, effort, and cost.
Initial extraction starts with cell lysis (chemical or physical) into a protective environment that immediately ensures minimal degradation of the target biomolecule. The sample can then be refined by degrading or removing contaminating biomolecules, and concentrated if necessary.