كلية التقنية الطبية صرمان
قسم التقنيات الحيوية والهندسة الوراثية الطبية
DNA
MICROINJECTION
By:Nehal omar Elhamessi
Introduction
DNA microinjection:
Is a transgenic technique also known as pronuclear microinjection,
a very fine glass pipette is used to manually inject DNA from one
organism into the eggs of another.
The generation of transgenic mice by DNA microinjection is a
powerful tool to investigate the molecular regulation of gene
expression, development, and disease.
The power of this technology is that foreign DNA can be introduced
into every cell of a developing organism and the phenotypic impact
of this genetic modification can be investigated in a system under
the constraints of normal development and physiology.
The generation of transgenic mice requires the preparation of the
transgene DNA construction, collection of one-cell fertilized mouse
embryos, injection of the transgene into mouse embryos, and
transfer of the surviving embryos. Mice born from such
manipulations are then screened for the presence of the transgene .
The Process of DNA Microinjection In mice:
1. Isolation the piece of DNA as per requirement and clone it into a
vector ( plasmid ).
2. Harvest newly fertilized egg before pronuclei fuse. The best time for
injection is early after fertilization when the ova have two pronuclei
( When the two pronuclei fuse to form a single nucleus)
3. Cells to be microinjected are placed in a container .
4. Inject of isolated DNA into the pronucleus of the sperm cell is done
under a powerful microscope using a glass micropipette tip of 0.5 mm
diameter.
5. The tip of micropipette is injected through the membrane of the cell to
deliver the contents of the needle into the cytoplasm and then the
empty needle is taken out.
6. The DNA is then microinjected into the eggs, and the eggs are
re-implanted into a pseudopregnant female mouse (the ovum is
transferred into the oviduct of a recipient female, or foster mother,
that has been induced by mating with a vasectomized male)
7. If the implantion successfully, test her offspring by remove a small
piece of tissue from the tail and examine its DNA for the desired
gene.
8. About 10_30%of offspring contain injected foreign DNA .
9. Mice expressing foreign DNA are bred to continue DNA in germ line.
ADVANTAGE of DNA Microinjection
1. No requirement of a marker gene.
2. Introduction of the target gene directly into a single cell.
3. Easy identification of transformed cells upon injection of dye
along with the DNA.
4. No requirement of selection of the transformed cells using
antibiotic resistance or herbicide resistance markers.
5. It can be used for creating transgenic organisms,
particularly mammals.
6. the production of these animals can serve as an invaluable
research resource.
DISADVANTAGE
1. The procedure is time consuming .
2. The execution of these procedures requires a highly efficient
system.
3. the cost of the generation of these mice can be cost prohibitive .
Application of Transgenic mice produced by DNA
Microinjection
1. Transgenic animals are used in medical research in as disease
model.
2. Transgenic animals are also used in the field called toxicology as
a source of detection of toxicants.
3. They are also used in Biotechnology as they produce specific
proteins.
4. They are also used in Xenografting.
5. In molecular biology, they are used for analyzing regulation of
gene expression.
6. Transgenic animals have a great role in pharmaceutical
industry, They are used for drug testing and drug production.
