GENETIC ENGINEERING

Speaker:

Ananda Kumar Saha

Department of Zoology, R.U.

GENETIC ENGINEERING may be described as the introduction of manipulated genetic material in to a cell in such a way as to replicate and be passed on to the progeny cell.

Genetic Engineering TechniqueGenetic Engineering Technique

Gene Manipulation: Gene Manipulation: Gene manipulation is Gene manipulation is defined as the formation of a new combination of defined as the formation of a new combination of a heritable material by the insertion of nucleic acid a heritable material by the insertion of nucleic acid molecules (DNA molecules) into bacterial plasmid molecules (DNA molecules) into bacterial plasmid or any other vector so as to allow their or any other vector so as to allow their incorporation in to the host organism, in which incorporation in to the host organism, in which they are capable of containing propagation.they are capable of containing propagation.

Microinjection TechniqueMicroinjection Technique

Gene CloningGene Cloning:: The insertion of a fragment The insertion of a fragment of DNA carrying a gene into a cloning vector of DNA carrying a gene into a cloning vector and subsequent propagation of recombinant and subsequent propagation of recombinant DNA molecules into many copies is known DNA molecules into many copies is known as gene cloning.as gene cloning.

Gene Cloning TechniqueGene Cloning Technique

BASIC STEPS OF GENE CLONING

Construction of recombinant DNA molecule

Transport of the recombinant DNA to the host cell

Multiplication of recombinant DNA molecule

Division of the host cell

Numerous cell division resulting in a clone

Gene cloning requires specialized tools Gene cloning requires specialized tools and techniques:and techniques:

Vehicles:Vehicles: The central component of a gene The central component of a gene cloning experiment is the vehicle, which transport cloning experiment is the vehicle, which transport the gene into the host cell and is responsible for its the gene into the host cell and is responsible for its replication. To act as a cloning vehicle a DNA replication. To act as a cloning vehicle a DNA molecule must be capable of entering a host cell molecule must be capable of entering a host cell and, once inside, replicating to produce multiple and, once inside, replicating to produce multiple copies of itself.copies of itself.

Vector:Vector: A DNA molecule, capable of replication A DNA molecule, capable of replication in a host organism, into which a gene is inserted to in a host organism, into which a gene is inserted to construct a recombinant DNA molecule.construct a recombinant DNA molecule.

Characteristic of a Vector:Characteristic of a Vector:

It must be able to replicateIt must be able to replicate There must be some way to introduce There must be some way to introduce

vector DNA into a cellvector DNA into a cell There must be some means of detecting There must be some means of detecting

its presence, preferably by plating test in its presence, preferably by plating test in petridishespetridishes

PLASMIDPLASMIDCharacteristics:Characteristics: They are small and contain 3-5 kb of DNAThey are small and contain 3-5 kb of DNA They are contain a suitable markers (antibiotic They are contain a suitable markers (antibiotic

resistant)resistant) They contain suitable restriction sites that can be They contain suitable restriction sites that can be

used for inserting DNA fragments for cloningused for inserting DNA fragments for cloning

Chromosomal DNA Plasmid DNA

Bacterium

Different types of cloningDifferent types of cloning

1.1. Recombinant DNA technology or Recombinant DNA technology or DNA cloningDNA cloning

2.2. Reproductive cloning andReproductive cloning and

3.3. Therapeutic cloningTherapeutic cloning

Recombinant DNA technology or Recombinant DNA technology or DNA cloningDNA cloning

The terms “recombinant DNA technology”, The terms “recombinant DNA technology”, “DNA cloning”, “molecular cloning” or “gene “DNA cloning”, “molecular cloning” or “gene cloning” all refer to the same process: the cloning” all refer to the same process: the transfer of a DNA fragment of interest from transfer of a DNA fragment of interest from one organism to a self-replicating genetic one organism to a self-replicating genetic element such as a bacterial plasmid. The DNA element such as a bacterial plasmid. The DNA of interest can then be propagated in a foreign of interest can then be propagated in a foreign host cell. This technology has been around host cell. This technology has been around since the 1970’s, and it has become a common since the 1970’s, and it has become a common practice in molecular biology labs today.practice in molecular biology labs today.

Reproductive cloningReproductive cloning

Dolly was created by reproductive cloning technology, in Dolly was created by reproductive cloning technology, in a process called “somatic cell nuclear transfer” (SCNT).a process called “somatic cell nuclear transfer” (SCNT).

Scientists transfer genetic material from the nucleus of a Scientists transfer genetic material from the nucleus of a donor adult cell to an egg whose nucleus, and thus its donor adult cell to an egg whose nucleus, and thus its genetic material, has been removed. The reconstructed genetic material, has been removed. The reconstructed egg containing the DNA from of a donor cell must be egg containing the DNA from of a donor cell must be treated with chemicals or electric current in order to treated with chemicals or electric current in order to stimulate cell division.stimulate cell division.

Once the clone embryo reaches a suitable stage, it is Once the clone embryo reaches a suitable stage, it is transferred to the uterus of a female host where it transferred to the uterus of a female host where it continues to develop until birth.continues to develop until birth.

Reproductive CloningReproductive Cloning

Dolly Dolly

Therapeutic cloningTherapeutic cloning

Therapeutic cloning, also called “embryo Therapeutic cloning, also called “embryo cloning”, is the production of human embryos cloning”, is the production of human embryos for use in research. for use in research.

The goal of this process is not to create clone The goal of this process is not to create clone human beings, but rather to harvest stem cells human beings, but rather to harvest stem cells that can be used to study human development that can be used to study human development and to treat disease. and to treat disease.

Stem cells are important to biomedical Stem cells are important to biomedical researchers because they can be used to researchers because they can be used to generate virtually any type of specialized cell in generate virtually any type of specialized cell in the human body.the human body.

How can cloning technologies be used?How can cloning technologies be used? Gene therapy can be used to treat certain genetic Gene therapy can be used to treat certain genetic conditions by introducing virus vectors that carry conditions by introducing virus vectors that carry corrected copies of faulty genes into the cells of a host corrected copies of faulty genes into the cells of a host organism.organism. Genes from different organisms that improve taste Genes from different organisms that improve taste and nutritional value or provide resistance to particular and nutritional value or provide resistance to particular types of disease can be used to genetically engineered types of disease can be used to genetically engineered food crops.food crops.Reproductive cloning also could be used to repopulate Reproductive cloning also could be used to repopulate endangered animals or animals that are difficult to endangered animals or animals that are difficult to breed.breed.

What animals have been cloned?What animals have been cloned?

Scientists have been cloning animals for Scientists have been cloning animals for many years. In 1952, the first animal, a many years. In 1952, the first animal, a tadpole, was cloned. tadpole, was cloned.

Since Dolly, researchers have cloned a Since Dolly, researchers have cloned a number of large and small animals including number of large and small animals including sheep, goats, cows, mice, pigs, cats, rabbits sheep, goats, cows, mice, pigs, cats, rabbits and a gaur.and a gaur.

Can organs be cloned for use in transplants?Can organs be cloned for use in transplants?

Scientists hope that one day therapeutic cloning Scientists hope that one day therapeutic cloning can be used to generate tissues and organs for can be used to generate tissues and organs for transplants. transplants.

The stem cells would be used to generate an organ The stem cells would be used to generate an organ or tissue that is a genetic match to the recipient. or tissue that is a genetic match to the recipient.

In theory, the cloned organ could then be In theory, the cloned organ could then be transplanted into the patient without the risk of tissue transplanted into the patient without the risk of tissue rejection.rejection.

What are the risks of cloning?What are the risks of cloning?Reproductive cloning is expensive and highly Reproductive cloning is expensive and highly inefficient.inefficient. More than 90% of cloning attempts fail to More than 90% of cloning attempts fail to produce viable offspring.produce viable offspring.More than 100 nuclear transfer procedures More than 100 nuclear transfer procedures could be required to produce one viable clone.could be required to produce one viable clone. In addition to low success rates, cloned animals In addition to low success rates, cloned animals tend to have more compromised immune tend to have more compromised immune function and higher rates of infection, tumor function and higher rates of infection, tumor growth and other disorders.growth and other disorders.

Thank you