Single-stranded conformational polymorphism

Single-stranded conformational polymorphism

Single-stranded conformational polymorphism (SSCP) is a technique used in molecular biology to detect genetic variation. It is based on the principle that single-stranded DNA (ssDNA) molecules with different sequences can adopt different conformations, which can be separated by electrophoresis. This method is particularly useful for identifying mutations in genes.

Principle

SSCP relies on the fact that when a double-stranded DNA is denatured into single strands, each strand can fold into a unique three-dimensional structure. These structures, or conformations, are influenced by the nucleotide sequence of the DNA. When subjected to non-denaturing polyacrylamide gel electrophoresis, these conformations migrate at different rates, allowing for the detection of sequence variations.

Procedure

The basic steps of SSCP are as follows:

1. DNA Denaturation: The double-stranded DNA is denatured to produce single strands. This is typically achieved by heating the DNA in the presence of a denaturing agent.

2. Electrophoresis: The single-stranded DNA is then subjected to electrophoresis on a non-denaturing polyacrylamide gel. The gel conditions are carefully controlled to maintain the single-stranded nature of the DNA.

3. Detection: After electrophoresis, the DNA is visualized using a staining method such as silver staining or autoradiography if the DNA is radioactively labeled.

4. Analysis: The pattern of bands is analyzed. Differences in band patterns indicate the presence of polymorphisms or mutations.

Applications

SSCP is widely used in genetic research and clinical diagnostics. Some of its applications include:

• Mutation Detection: SSCP is used to screen for mutations in genes associated with diseases such as cancer.
• Genetic Mapping: It helps in identifying genetic markers linked to specific traits or diseases.
• Population Genetics: SSCP can be used to study genetic variation within and between populations.

Advantages and Limitations

Advantages

• Simplicity: SSCP is a relatively simple and cost-effective method for mutation detection.
• Sensitivity: It can detect single nucleotide changes in DNA sequences.

Limitations

• Resolution: SSCP may not resolve all mutations, especially if they do not significantly alter the conformation of the DNA.
• Optimization: The technique requires careful optimization of gel conditions to achieve reliable results.

Also see

• Polymerase chain reaction
• Gel electrophoresis
• DNA sequencing
• Restriction fragment length polymorphism