What is DNA Fingerprinting?
𝗗𝗡𝗔 𝗳𝗶𝗻𝗴𝗲𝗿𝗽𝗿𝗶𝗻𝘁𝗶𝗻𝗴 is a laboratory technique used to identify the 𝘂𝗻𝗶𝗾𝘂𝗲 𝗽𝗮𝘁𝘁𝗲𝗿𝗻𝘀 𝗼𝗿 𝘀𝗲𝗾𝘂𝗲𝗻𝗰𝗲𝘀 of an individual's (person or organism) DNA. It was first developed by 𝗦𝗶𝗿 𝗔𝗹𝗲𝗰 𝗝𝗲𝗳𝗳𝗿𝗲𝘆𝘀 in 𝟭𝟵𝟴𝟰–𝟭𝟵𝟴𝟱.
- About 𝟵𝟵.𝟵% of human DNA is identical across all humans.
- The remaining 𝟬.𝟭% made up of non-coding repetitive sequences called 𝗩𝗡𝗧𝗥𝘀 (𝗩𝗮𝗿𝗶𝗮𝗯𝗹𝗲 𝗡𝘂𝗺𝗯𝗲𝗿 𝗼𝗳 𝗧𝗮𝗻𝗱𝗲𝗺 𝗥𝗲𝗽𝗲𝗮𝘁𝘀) forms each person's unique genetic fingerprint.
Principle of DNA Fingerprinting
The fundamental principle relies on the fact that specific regions of the human genome contain DNA sequences that repeat multiple times, known as 𝘁𝗮𝗻𝗱𝗲𝗺 𝗿𝗲𝗽𝗲𝗮𝘁𝘀.
- The 𝗹𝗲𝗻𝗴𝘁𝗵 and 𝗻𝘂𝗺𝗯𝗲𝗿 of these repeating units vary among individuals.
- This variation is called 𝗽𝗼𝗹𝘆𝗺𝗼𝗿𝗽𝗵𝗶𝘀𝗺.
- Polymorphism is detected using 𝗿𝗲𝘀𝘁𝗿𝗶𝗰𝘁𝗶𝗼𝗻 𝗲𝗻𝘇𝘆𝗺𝗲𝘀 and the 𝗦𝗼𝘂𝘁𝗵𝗲𝗿𝗻 𝗯𝗹𝗼𝘁𝘁𝗶𝗻𝗴 technique.
- Since VNTR regions are highly variable but stably inherited, they act as reliable genetic markers passed from parents to offspring, which is why the technique also works for paternity testing.
Step-by-Step Procedure of DNA Fingerprinting
The DNA fingerprinting process follows six major laboratory steps:
1. 𝗜𝘀𝗼𝗹𝗮𝘁𝗶𝗼𝗻 𝗼𝗳 𝗗𝗡𝗔
DNA is extracted from a biological sample such as blood, hair root/follicle attached, skin, saliva, or semen.
- The sample is treated with lysis buffer and enzymes (like proteinase K) to break open cells and remove proteins.
- Pure DNA is then precipitated out using chilled ethanol and dissolved in a buffer for further use.
- Even a very small quantity of biological material (a single hair root or a few microliters of blood) is enough, since the technique is highly sensitive.
2. 𝗗𝗶𝗴𝗲𝘀𝘁𝗶𝗼𝗻 𝗯𝘆 𝗥𝗲𝘀𝘁𝗿𝗶𝗰𝘁𝗶𝗼𝗻 𝗘𝗻𝘇𝘆𝗺𝗲𝘀
The extracted DNA is cut into smaller fragments using restriction endonuclease enzymes (e.g., HinfI, EcoRI).
- These enzymes recognize specific short DNA sequences and cleave the DNA at those exact points.
- Because the number and position of VNTR repeats differ between individuals, restriction digestion produces DNA fragments of different lengths. This variation forms the basis of DNA fingerprinting.
3. 𝗚𝗲𝗹 𝗘𝗹𝗲𝗰𝘁𝗿𝗼𝗽𝗵𝗼𝗿𝗲𝘀𝗶𝘀
The DNA fragments are separated based on size using agarose gel electrophoresis.
- Fragments are loaded into wells of an agarose gel and an electric current is applied.
- Since DNA is negatively charged, fragments migrate toward the positive electrode (anode).
- Smaller fragments move faster and travel farther, while larger fragments remain closer to the well creating a size-based separation pattern.
4. 𝗕𝗹𝗼𝘁𝘁𝗶𝗻𝗴 (𝗦𝗼𝘂𝘁𝗵𝗲𝗿𝗻 𝗕𝗹𝗼𝘁𝘁𝗶𝗻𝗴)
The separated DNA bands are transferred from the fragile gel onto a solid nitrocellulose or nylon membrane.
- Before transfer, the DNA is denatured (converted to single strands) using an alkaline solution.
- The gel is placed over the membrane, and capillary action or vacuum blotting moves the DNA fragments onto the membrane, preserving their exact band positions.
- This step, named after its inventor Edwin Southern, makes the DNA pattern stable enough for the next stage.
5. 𝗛𝘆𝗯𝗿𝗶𝗱𝗶𝘇𝗮𝘁𝗶𝗼𝗻
A labeled VNTR probe, traditionally radioactive and in some modern applications fluorescent, is introduced to bind to complementary DNA sequences.
- The membrane is incubated with the probe under controlled temperature and salt conditions to ensure specific base-pairing.
- Excess, unbound probe is washed away, leaving the probe attached only at the VNTR sites.
6. 𝗔𝘂𝘁𝗼𝗿𝗮𝗱𝗶𝗼𝗴𝗿𝗮𝗽𝗵𝘆
The membrane is exposed to an X-ray film, producing dark bands known as the DNA fingerprint.
- Wherever the radioactive/fluorescent probe has bound, it exposes the film, forming a unique banding pattern.
- This pattern is photographed and compared against a suspect, parent, or reference sample matching or mismatching bands confirm or rule out identity.
Applications of DNA Fingerprinting in Zoology
- 𝗙𝗼𝗿𝗲𝗻𝘀𝗶𝗰 𝗦𝗰𝗶𝗲𝗻𝗰𝗲 (𝗖𝗿𝗶𝗺𝗲 𝗜𝗻𝘃𝗲𝘀𝘁𝗶𝗴𝗮𝘁𝗶𝗼𝗻): Identifying criminals, solving rape cases, and matching biological evidence in murder investigations.
- 𝗣𝗮𝘁𝗲𝗿𝗻𝗶𝘁𝘆 𝗗𝗶𝘀𝗽𝘂𝘁𝗲𝘀: Accurately determining the biological parents of a child.
- 𝗖𝗼𝗻𝘀𝗲𝗿𝘃𝗮𝘁𝗶𝗼𝗻 𝗕𝗶𝗼𝗹𝗼𝗴𝘆 (𝗪𝗶𝗹𝗱𝗹𝗶𝗳𝗲 𝗖𝗼𝗻𝘀𝗲𝗿𝘃𝗮𝘁𝗶𝗼𝗻): Tracking endangered animal populations and combating illegal poaching.
- Population Genetics & Individual Identification: DNA profiling can be used to study genetic variation, identify individuals, and assess genetic relationships within animal populations.
Conclusion
References
- Biotechnology By P.K Gupta
Suggested Reading
- Centrifuge and Centrifugation: Principle, Procedure, Applications, Examples & Uses
- Polymerase Chain Reaction (PCR): Principle, Steps, Primer Design, Applications & Diagram
- Gel Electrophoresis: Principle, Types (AGE & PAGE), SDS-PAGE, Buffer Chemistry & Applications |
- CO₂ Incubator: Principle, Working, Parts, Uses & pH Regulation