DNA recombinant

technology
Definition of recombinant DNA
technology( 基因重組 )
A series of procedures used to
recombine DNA segments.. Under
certain conditions, a recombinant DNA
molecule can enter a cell and replicate.
History of recombinant DNA
technology

Recombinant DNA technology is one

of the recent advances in
biotechnology, which was developed
by two scientists named Boyer and
Cohen in 1973.
Basic principle of recombinant
DNA technology

The DNA is inserted into another

DNA molecule called ‘vector’( 載體 )

. The recombinant vector is then

introduced into a host cell where it
replicates itself, the gene is then
produced
Basic principle of recombinant
DNA technology
 Applications of
Recombinant DNA
Technology
Large-scale production of human
proteins by genetically
engineered bacteria.

Such as : insulin, Growth

hormone, Interferons and
Blood clotting factors (VIII & IX)
Production of Human
Insulin( 胰島素 )
1) Obtaining the human insulin gene
Human insulin gene can be obtained by
making a complementary DNA (cDNA) copy
of the messenger RNA (mRNA) for human
insulin.
 2)Joining the human insulin gene
into a plasmid( 質粒 ) vector

The bacterial plasmids and the cDNA are

mixed together. The human insulin gene
(cDNA) is inserted into the plasmid through
complementary base pairing at sticky ends.
3)Introducing the recombinant
DNA plasmids into bacteria
The bacteria E.coli is used as the host cell. If E.
coli and the recombinant plasmids are mixed
together in a test-tube.
 4)Selecting the bacteria which
have taken up the correct
piece of DNA
The bacteria are spread onto nutrient agar. The
agar also contains substances such as an
antibiotic which allows growth of only the
transformed bacteria.
Vaccinedevelopment ( 疫苗
生產 )
The surface antigen of Plasmodium
falciparum, one of the 4 species of malaria
has been transferred to E. coli to produce
amounts large enough to develop a vaccine
against this form of malaria( 瘧疾 ). It
works well enough for people who will
visit a malarious region for a relatively
short period of time
 Hemophilia( 血友病 ) A and
B
The genes encoding factors 8 and 9 are on the X
chromosome.
Like other X-linked disorders, hemophilia A and B are found
almost exclusively in males because they inherit just a single
X chromosome, and if the gene for factor 8 (or 9) on it is
defective, they will suffer from the disease.
There are many different mutant versions of the genes for
factors 8 and 9. Although some produce only a minor effect
on the function of their protein, others fail to produce any
functioning clotting factor.
 Treating Hemophilia A and B

Factor 8 and 9 can be extracted from donated blood, usually

pooled from several thousand donors, and purified. Injections of
this material can halt episodes of bleeding in hemophiliacs and
have allowed countless young men to live relatively normal lives.
However, blood contaminated with the human immunodeficiency
virus (HIV) was unknowingly used to manufacture preparations of
factors 8 and 9. Many have since died of AIDS.
The Future of Treating Hemophilia A
and B
•all donated blood is now tested to see if the
donor has been infected with HIV (as well as
hepatitis B and C);
•plasma-derived preparations of factors 8 and 9
are now treated with heat and/or solvents to
destroy any viruses that might be present;
•recombinant factor 8 and recombinant
factor 9 made by genetic engineering are now
available.
Now the treatment become more safety!
Gene therapy for genetic diseases
Still in the experimental stages, it may be possible to
transfer the gene for normal adult hemoglobin into
marrow stem cells of an individual with sickle-cell
anemia( 鐮刀形貧血 ). The goal is to promote the
growth of enough cells to produce enough normal
hemoglobin to alleviate the symptoms of sickle-cell
anemia. One hundred percent (100%) is NOT
required to attain the alleviation of symptoms.
 Safety Issues in relation to
Recombinant DNA Technology
As bacteria is commonly used in recombinant DNA work,
there has always been a concern among scientists and a
worry among people that there is a possibility that a clone
of highly pathogenic recombinant bacteria were made by
accident, then escaped from the laboratory and caused an
epidemic for which no drugs were available.

Recombinant DNA Advisory Committee (RAC) was

established in 1974 in the United States, which
responds to public concerns regarding the safety of
manipulation of genetic material through the use of
recombinant DNA techniques.
2 types of control : physical
containment and biological
containment ( 封鎖 )
Effective biological safety programs were
operated in a variety of laboratories, which
include a set of standard practices generally
used in microbiological laboratories, and
special procedures, equipment and laboratory
installations that provide physical barriers of
varying degrees.
Biological containment
In considering biological containment, the
vector (plasmid, organelle, or virus) for the
recombinant DNA and the host (bacterial,
plant, or animal cell) in which the vector is
propagated in the laboratory will be considered
together.

(i) survival of the vector in its host outside the

laboratory, and (ii) transmission of the vector
from the propagation host to other non-
laboratory hosts.
 Dangerous of DNA
recombinant technology

It is always possible that an

antibiotic-resistant plasmid could be
accidentally incorporated into a
dangerous pathogen with serious
medical consequences.
http://www.ied.edu.hk/biotech/eng/clas
srm/explain/gene5.htm