Nucleic acid extraction

Nucleic acid extraction reagents are combinations of various chemical reagents used for extracting nucleic acids (including DNA and RNA) from biological samples. The following is a detailed introduction:

I. Main Components and Their Functions

1. Lysis Buffer
   • Components: Usually contains detergents (such as sodium dodecyl sulfate, SDS), salts (such as sodium chloride, magnesium chloride, etc.), and buffer substances (such as Tris – HCl).
   • Functions: Detergents can disrupt the cell membrane and nuclear membrane structures of biological samples, releasing nucleic acids inside the cells. Salts can maintain the ionic strength of the solution, which helps with the dissolution and stability of nucleic acids. Buffer substances can adjust the pH of the solution, keeping it within the appropriate pH range for the stable existence of nucleic acids, generally around 7 – 9.
2. Proteinase K
   • Component: It is a serine protease.
   • Function: During the nucleic acid extraction process, proteinase K can break down the proteins in the samples. It can degrade the proteins bound to nucleic acids, preventing these proteins from interfering with the extraction and purification of nucleic acids in subsequent steps, ensuring the obtainment of pure nucleic acids.
3. Binding Buffer (if the Magnetic Bead or Centrifugal Column Method is Used)
   • Components: In the magnetic bead method, the binding buffer contains substances that enable nucleic acids to specifically bind to magnetic beads; in the centrifugal column method, the binding buffer helps nucleic acids to adsorb onto the filter membrane of the centrifugal column. For example, the binding buffer in the magnetic bead method may contain chemical groups that can have ionic interactions with nucleic acids.
   • Functions: Promote the binding of nucleic acids to the corresponding carriers (magnetic beads or centrifugal column filter membranes) for the subsequent separation and purification of nucleic acids.
4. Washing Buffer
   • Components: Generally, it is a solution containing ethanol or other organic solvents, and also includes buffer substances and salts.
   • Functions: After nucleic acids bind to carriers (magnetic beads or centrifugal column filter membranes), it is used to wash away impurities, such as undegraded proteins and polysaccharides. Organic solvents can reduce the solubility of nucleic acids, making them adhere more firmly to the carriers and helping to remove impurities. Buffer substances and salts maintain the stability of the solution environment.
5. Elution Buffer
   • Components: Usually, it is a buffer with low ionic strength, such as TE buffer (Tris – HCl and EDTA).
   • Functions: Used to elute the purified nucleic acids from carriers (magnetic beads or centrifugal column filter membranes), obtaining nucleic acid solutions available for subsequent experiments or tests.

II. Types

1. Reagents Based on Different Extraction Methods
   • Reagents for Magnetic Bead – based Extraction: Specially designed for nucleic acid extraction using the magnetic bead method. The magnetic beads in the reagents have special surface modifications and can specifically bind to nucleic acids under the action of the binding buffer. This reagent relies on an external magnetic field to manipulate the magnetic beads during the operation to achieve the separation and purification of nucleic acids.
   • Reagents for Centrifugal Column – based Extraction: Applicable to nucleic acid extraction using the centrifugal column method. Its components are designed based on the characteristics of the centrifugal column, enabling nucleic acids to adsorb onto the filter membrane of the centrifugal column under the action of centrifugal force and obtain pure nucleic acids through washing and elution steps.
   • Reagents for Traditional Phenol – Chloroform Method: Although this method is gradually being replaced by more advanced techniques, it is still used under certain research or laboratory conditions. The reagents for the phenol – chloroform method enable proteins to denature and be distributed into the organic phase through the action of organic solvents phenol and chloroform, while nucleic acids remain in the aqueous phase, thereby achieving nucleic acid extraction.
2. Reagents for Different Sample Types
   • Reagents for Nucleic Acid Extraction from Blood: Blood contains various cellular components such as red blood cells, white blood cells, and platelets, as well as complex matrices such as plasma. The lysis buffer formulations of the reagents for nucleic acid extraction from blood are optimized to quickly and effectively lyse white blood cells (which are the main source of nucleic acids in blood) while avoiding interference from substances such as hemoglobin in red blood cells in the nucleic acid extraction process.
   • Reagents for Nucleic Acid Extraction from Tissues: The cellular structures and components of different tissues vary greatly, such as muscle tissue, brain tissue, and liver tissue. The reagents for nucleic acid extraction from tissues take into account the characteristics of tissue samples and generally contain stronger lysis components to ensure the full disruption of tissue – cell structures and the release of nucleic acids.
   • Reagents for Nucleic Acid Extraction from Swab Samples (such as Throat Swabs and Nasal Swabs): The nucleic acid content in swab samples is relatively low, and there may be more exogenous contaminations. These reagents focus on efficiently extracting nucleic acids from small – quantity samples and removing substances that may inhibit subsequent nucleic acid tests, such as fiber impurities from the sampling swabs introduced during the swab sampling process.