UNIT 4: Transfection, Screening and Selection Technique
Exam Focus: Transfection methods are commonly asked as short notes (Microinjection, Lipofection, Electroporation). For selection, understand the principle and utility of the **HAT selection** medium, especially in Hybridoma technology.
Table of Contents
1. Transfection
Transfection is the introduction of **foreign nucleic acids (DNA or RNA)** into eukaryotic cells. The various methods aim to bypass the cell membrane barrier.
Microinjection
A physical method where a fine glass needle is used to **manually inject** the DNA or RNA directly into the cell nucleus or cytoplasm.
- **Pros:** Highly efficient and precise.
- **Cons:** Low throughput (one cell at a time).
Lipofection
A chemical method using **liposomes** (lipid vesicles) or **lipofection** reagents (cationic lipids) that form complexes with the negatively charged nucleic acids. These complexes fuse with the cell membrane, mediating DNA entry.
Electroporation
A physical method using a brief, high-voltage electric pulse to create **transient pores** in the cell membrane, allowing the nucleic acid in the surrounding medium to enter the cell.
Sonication
A physical method using **ultrasound waves** to temporarily disrupt the cell membrane structure, facilitating the uptake of nucleic acids from the surrounding medium.
| Method | Mechanism | Type |
|---|---|---|
| Microinjection | Direct physical delivery via needle. | Physical |
| Lipofection | Cationic lipid vesicles fuse with cell membrane. | Chemical |
| Electroporation | Electric pulse creates transient membrane pores. | Physical |