a proper description about the process microinjection and also about gene transfer. and different types of DNA delivery methods. with advantages, disadvantages, limitations and applications.

1. MICROINJECTION
NIKITA NARWAL
B.SC. BIOTECHNOLOGY (FINAL YEAR)
ROLL NO:194713
PH. NO.:9896989063

2. WHAT IS GENE TRANSFER?
• It is defined simply as a technique to efficiency and stably
introduce foreign genes into the genome of target cells.
• The insertion of unrelated, therapeutic genetic information
in the form of DNA into target cells.

3. GENE TRANSFER TECHNIQUE
• Based on the vectors used the gene transfer techniques can
be divided as,
1. Non-Viral methods.
2. Viral method.

4. NON-VIRAL DELIVERY SYSTEM
• Non-viral vectors using mechanical or chemical approaches
can efficiently transfect cells in vitro.
• Mechanical methods involve direct injection or the use of
“gene gun technology” to introduce the plasmid DNA.
LIMITATIONS
 Low levels of gene expression.
 Inability to use for systemic administration due to the
presence of serum nucleases.

5. GENERALLY THERE ARE TWO
APPROACHES OF DNA TRANSFER
1. Natural method of DNA transfer
2. Artificial method of DNA transfer

6. NATURAL METHOS OF DNA TRANSFER
AGROBACTERIUM MEDIATED TRANSFER
RETROVIRAL TRANSDUCTION
CONJUGATION TRANSPOSITION
BACTERIAL
TRANSFORMATION
PHAGE
TRANSDUCTION

7. ARTIFICIAL METHOD OF DNA TRANSFER
PHYSICAL METHOD
• MACROINJECTION
• MICROINJECTION
• PROTOPLAST FUSION
• BIOLISTIC TRANSFORMATION
CHEMICAL METHOD
• DNA TRANSFER BY CALCIUM PHOSPHATE METHOD
• TRANSFER OF DNA BY USE OF POLYETHENE GLYCOL
• USE OF DEAE-DEXTRAN FOR DNA TRANSFER
• LIPOSOME MEDIATED TRANSFER
ELECTRICAL METHOD
• ELECTROPORATION
• ELECTROFUSION

8. MICROINJECTION
• Microinjection where the DNA is directly injected into plant protoplast or
cells (specifically into the nucleus or cytoplasm) using fine tipped(0.5-1.0
micrometer diameter) glass needle or micropipette.
• This method of gene transfer is used to introduce DNA into large cells,
normally performed under a specialized optical microscope setup called a
micromanipulator.
• The process is frequently used as a vector in genetic engineering and
transgenetics to insert material into a single cell.
• Computerized control of holding pipette, needle, microscope stage and
video technology has improved the efficiency of this technique.

11. ADVANTAGES OF MICROINJECTION
• Frequency of stable integration of DNA is far better as
compared to other method.
• Method is effective in transforming primary cells as well as
cells in established culture.
• The DNA injected in this process is subjected to less
extensive modification.
• More precise integration of recombinant gene in limited
copy number can be obtained.

12. LIMITATIONS OF MICROINJECTION
1. Costly.
2. Skilled personal required.
3. More useful for animal cells.
4. Embryonic cells are preferred for manipulation.
5. Knowledge of mating timing, oocyte recovery is essential.
6. Methos is useful for protopplast and not for the walled cells.

13. APPLICATIONS OF MICROINJECTION
• Process is applicable for plant cells as well as animal cells but
more common for animal cells.
• Technique is ideally useful for producing transgenic animal
quickly.
• Procedure is important for gene transfer to embryonic cells.
• Applied to inject DNA into plant nuclei.
• Method has been successfully used with cells and protoplast of
tobacco, alfalfa etc.

14. • Microinjection is potentially a useful method
for simultaneous introduction of multiple
bioactive compounds such as antibodies,
peptides, RNAs, plasmid, diffusion markers,
elicitors, Ca2+ as well as nucleus and artificial
micro or nano particles containing those
chemicals into the same target-cells.