Topic: RFLP, PCR
Question: Explain RFLP.Describe clinical uses
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RFLP stands for Restriction Fragment Length Polymorphism. It is a technique used to detect variation in DNA sequences. It involves:
1. Isolating DNA from cells or tissues. The DNA contains specific sequences or patterns that are unique to each individual.
2. Cutting the DNA into fragments using restriction enzymes. These enzymes cut DNA at specific nucleotide sequences known as restriction sites. The cut sites are different for each individual due to small differences or variations in their DNA.
3. Separating the DNA fragments according to size using gel electrophoresis. The fragments form a unique “barcode” or “fingerprint” pattern on the gel that can identify the individual.
4. Analyzing the DNA fragment sizes and patterns to detect genetic differences between individuals. Variations even in a single nucleotide can result in the gain or loss of a cut site, producing different sized DNA fragments.
The clinical uses of RFLP include:
1. Diagnosing genetic diseases: RFLP can detect mutations that create or abolish restriction sites. This can diagnose disorders caused by defects in a single gene like sickle cell disease, cystic fibrosis, etc.
2. Determining disease risk: RFLP may identify people who carry one copy of a disease-causing gene mutation (carriers) by detecting loss of a cut site. Carriers can then be counseled about the risk of passing the mutation to offspring.
3. Identifying chromosomal abnormalities: RFLP can detect variations in the sizes and numbers of fragments from X and Y chromosomes to diagnose conditions like XO Turner’s syndrome.
4. DNA fingerprinting for forensic analysis and paternity testing: The unique RFLP pattern of each individual’s DNA can establish identity or biological relationships between individuals.
5. Detecting loss of heterozygosity: This can detect the loss of one copy of a gene in certain cancers like retinoblastoma. One DNA fragment may disappear or change in size, indicating loss of one allele.
6. Linkage analysis: RFLP can track the inheritance of alleles between generations based on their location on the same chromosome. This is used to locate disease-causing genes.
So in summary, RFLP is a DNA-based technique that relies on detecting changes in the sizes and patterns of restriction fragments to diagnose genetic diseases, determine disease risks, identify chromosome abnormalities, establish identity/paternity and track inheritance of disease genes within families. When combined with gene mapping, it is a useful tool for locating and isolating specific genes.
