BIOTECH Q3M2
Genetic Engineering
The process of altering the genetic makeup of organisms to achieve desired traits. This often involves the introduction of new DNA from different species.
Examples of Genetically Engineered Organisms:
Glow-in-the-dark animals: Such as genetically modified zebrafish with fluorescent protein genes.
Pollution-fighting plants: Engineered to absorb heavy metals from soil, improving phytoremediation.
Fast-growing salmon: Modified to grow faster than non-engineered counterparts, increasing aquaculture efficiency.
Banana vaccine: Genetically engineered to produce vaccines against diseases like hepatitis B when consumed.
Medicinal eggs: Chickens modified to produce eggs containing therapeutic proteins for human use.
Tools and Techniques in Handling DNA
A. Gel Electrophoresis
Purpose: To separate DNA fragments by size for analysis or purification. Materials: Agarose gel, buffer solution, electricity, DNA samples, and a gel electrophoresis chamber.
Process:
Prepare agarose gel by melting agarose powder in a buffer solution.
Pour the gel into a mold and let it solidify.
Load DNA samples along with a DNA ladder (size marker) into wells of the gel.
Apply electrical current; DNA fragments migrate toward the positive electrode due to their negative charge.
Smaller DNA fragments move faster and cover greater distances than larger fragments, leading to separation.
Stain the gel post-run to visualize the DNA bands for analysis.
B. Blotting Techniques
Purpose: Transfer DNA, RNA, or proteins from a gel to a membrane for detection and characterization.
Types of Blotting:
Southern Blotting: Detects DNA.
Northern Blotting: Detects RNA, crucial for studying gene expression.
Western Blotting: Detects proteins, using antibodies for specific identification.(Note: Southern blotting is the primary focus of this module.)
C. Southern Blotting
Developed by: Sir Edwin Southern in the 1970s.Steps involved:
Extract DNA from cells and cut it into fragments using restriction enzymes.
Separate the fragments through gel electrophoresis.
Transfer the separated fragments to a nitrocellulose or nylon membrane.
Label fragments with a specific DNA probe for hybridization to visualize targeted sequences.
Use autoradiography or chemiluminescence to visualize the hybridization patterns.
D. Gene Gun Technique
Description: A revolutionary method allowing direct injection of foreign DNA into a cell.
Process:
Uses small particles coated with DNA (such as gold or tungsten).
Initially developed for plant transformation; has since been adapted for some animal cells.
The device operates using high-pressure gas to accelerate the particles into the target cells.
Effective for tissues where traditional methods are less successful.
E. Electroporation
Description: A technique that utilizes electric current to create transient pores in cell membranes, enabling foreign DNA to enter cells.
Purpose: Widely used for introducing plasmids and other nucleic acids into bacterial and plant cells efficiently.
Usage: Particularly beneficial for transforming cells that are difficult to transfect using conventional methods.
F. Microinjection
Description: The process of directly inserting DNA into a cell's cytoplasm or nucleus using specialized equipment.
Requires: Holding pipette, microinjection needle, and an inverted microscope to accurately guide and inject into the desired cell.
Applications: Commonly used in fertilized eggs for creating transgenic organisms or in specific cell lines for functional gene studies.
Summary of Techniques
Gel Electrophoresis: Separates DNA based on size using agarose gel and electricity.
Southern Blotting: Enables detection of specific DNA sequences after separation.
Gene Gun: Transforms cells directly with DNA-coated particles.
Electroporation: Facilitates DNA entry by creating temporary pores in membranes.
Microinjection: Directly introduces DNA into cells using fine-tipped needles.
from agriculture to medicine.