N4S

Genetic Engineering concepts, specially their applications highlight the practical implications of Biotechnology. Hence, they become the most important area for UPSC. Students are expected to be thorough with it.

Genetic Engineering is an overarching term referring to modification of an organism’s genome. Here is a summary video outlining important techniques and applications which you might find interesting.

The techniques of genetic engineering include the creation of recombinant DNA, use of gene cloning and gene transfer.

A. Recombinant DNA technology

Recombinant DNA technology involves several steps in specific sequence such as

1.isolation of DNA,

2.fragmentation of DNA,

3.isolation of a desired DNA fragment,

4.ligation of the DNA fragment into a vector,

5.transferring the recombinant DNA into the host,

6.culturing the host cells in a medium at large scale and extraction of the desired product.

The DNA molecule thus produced is called recombinant DNA.

B. Gene Cloning

It is the creation of a genetic copy of a sequence of DNA or of the entire genome of an organism.

It is usually done through two approaches: Somatic Cell Nuclear Transfer(SCNT) and Artificial Embryo Twinning

1.Somatic Cell Nuclear Transfer(SCNT)

• A somatic cell is any cell of a living organism other than the reproductive cells
• The nucleus of a somatic cell is removed and kept, and the host’s egg cell is kept and nucleus removed and discarded
• The kept nucleus is then fused with the ‘deprogrammed’ egg cell. After being inserted into the egg, the lone somatic cell nucleus is reprogrammed by the host egg cell
• The egg, now containing the somatic cell’s nucleus is stimulated with a shock and will now begin to divide.
• After multiple divisions, the single cell forms a blastocyst(an early stage embryo) with almost identical DNA to the original organism.
• Dolly the sheep was the first animal ever to be cloned using SCNT
• There are two possible uses of SCNT:
  • Therapeutic Cloning: Creating cloned human embryos are not implanted into a female uterus rather used in research and therapy to obtain stem cells
  • Reproductive Cloning: Creating cloned human embryos with the intention of placing them into the uterus where it can implant and develop

2. Artificial Embryo Twinning

https://www.youtube.com/watch?v=FvjWKip6Pm8

• This technology mimics the natural process of creating identical twins
• The early stage embryo is manually separated into individual cells and allowed to divide and develop on their own.
• The resulting embryos are placed into a surrogate mother, where they are carried to term and delivered. As the embryos came from the same zygote(fertilized egg), they are genetically identical.

Summing up the similarities and differences

C. Gene Silencing

Certain genes in a cell may be defective and could cause undesirable traits. The process of regulation or suppression of such genes is called Gene Silencing.

RNA Interference(RNAi) and Antisense Technology are two common processes by which this is achieved. More details are not required.

D. CRISPR-Cas9

CRISPR-Cas9 technique has appeared extensively in news

Salient features

• CRISPR-Cas9 is a genome-editing tool. CRISPR is a naturally occurring defence mechanism in bacteria.
• The CRISPR-Cas9 consists of two molecules that are responsible for gene editing.
  • An enzyme called Cas9 that acts as a pair of molecular scissors that cuts the two strands of DNA at a specific location
  • A guide RNA(gRNA) that binds to DNA and guides the Cas9 to the right part of the genome
• The gRNA is designed to find and bind to a specific sequence in the DNA. The gRNA has bases that are complementary to those of the target DNA sequence.
• The Cas9 follows the gRNA to the same location in the DNA sequence and makes a cut across both strands of the DNA
• At this stage, the cell recognizes that the DNA is damaged and tries to repair it
• Scientists can use the DNA repair machinery to introduce changes to one or more genes in the genome of the cell of interest.

E. Gene splicing

Gene splicing is the process of chemically cutting DNA in one organism and inserting it into the DNA strand of a vector, usually a plasmid. The DNA is cut using special chemicals called restriction enzymes. Through this technique, microorganisms are genetically engineered. Gene splicing primarily involves the following steps

1.Cutting of DNA or genes from the donor cell.

Tool Used – Chemicals called restriction enzymes. Restriction enzymes are chemicals extracted from certain strains of bacteria. Different restriction enzymes recognize and cut at particular nuclear sequences.

2.Transportation of the cut or desired DNA

A.DNA Micro injection – Direct Micro-injection of the chosen gene.

B.Retrovirus mediated gene transfer. Retroviruses are used as vectors to transfer genetic material taking advantage of their natural ability to infect host cells and insert their DNA into host cell DNA.

C.Embryonic Stem Cell mediated gene transfer.

APPLICATIONS

A. Three-parent babies

Three-parent babies have repeatedly remained in news. The world’s first three-parent baby was born in 2016 in Mexico to Jordanian parents.

In 2017, UK became the first country to give licenses to use this technique to prevent incurable genetic diseases being inherited by children.
Remember we had very briefly touched upon the Mitochondria when we discussed the Cell and the Nucleus. Mitochondria contains mtDNA and mutations in it can give rise to various genetic disorders. Since mtDNA is transferred from mother to child, these disorders are also transferred. The three-parent babies technique tries to overcome this shortcoming by preventing the transfer of mitochondria from mother to child.

There are 2 versions – Pronuclear Transfer and Spindle Transfer

Pronuclear Transfer

Spindle Transfer

https://www.sciencenewsforstudents.org/article/how-make-three-parent-baby

B.Gene Therapy

It is a technique through which a new DNA is introduced into a patient to treat a genetic disease. The new DNA usually contains a functioning gene to correct the effects of a non-functioning or disease causing gene.

A four-year old girl became the first gene therapy patient. She had adenosine deaminase (ADA) deficiency, a genetic disease which left her defenseless against infections.

There are two types of gene therapy:

Somatic Gene Therapy: The genes are transferred into the somatic cells of a patient. Any modifications and effects will be restricted to the individual patient only and will not be inherited by the offspring.

Germline Gene Therapy: The genes are transferred into the germ cells(sperm or eggs). This would allow the therapy to be heritable and passed onto the offspring.

C. Genetic Modification of Plants(GM Crops)

The DNA of plants used in agriculture are modified using genetic engineering techniques

There are two primary processes through which plants are modified:

Cisgenic: Genes are artificially transferred between organisms that could be conventionally bred i.e. from a crossable or sexually compatible plant.

Transgenic: Genes from a non-plant species or from a donor plant that is sexually incompatible with the recipient plant are inserted, which is a form of horizontal gene transfer

Biotech firms make use of the following technologies to ensure sterile seeds and crops with specific properties

Terminator Technology: It is the genetic modification of plants to make them produce sterile seeds. They are also known as suicide seeds.

Traitor Technology: Traitor technology can allow traits in genetically modified seeds to be switched on and off by using special proprietary chemicals.

D. In-vitro Fertilization

https://www.youtube.com/watch?v=J1xHUA_fiQ4

It is the process of fertilization by extracting eggs, retrieving a sperm sample, and then manually combining an egg and sperm in a laboratory dish. The embryo(s) is then transferred to the uterus to induce pregnancy.