Genetic Engineering

GENETIC ENGINEERING (Recombinant DNA technology) Genetic engineering or recombinant DNA technology is defined as the manipulation of DNA of one organism (donor) and its transfer into another organism (the host) where its combines with that of the host organism.

Where it combines with that of the host organism To create as new gene combination, genetic engineers must be able to

1. Identify the Target Gene: They must first identify the specific gene that they want to transfer or modify. This gene could be responsible for a desirable trait, like disease resistance or improved growth.
2. Extract the Gene: Using special tools, genetic engineers extract the desired gene from the DNA of an organism. This is often done using restriction enzymes that cut the DNA at specific points.
3. Modify the Gene: Sometimes, the extracted gene may need to be altered. Genetic engineers can change the gene's sequence to enhance its function or remove any unwanted parts using techniques like gene editing.
4. Insert the Gene into a Vector: The gene is then inserted into a vector (a small piece of DNA, such as a plasmid or virus) that can carry the gene into a new organism. The vector helps transfer the gene into the target organism’s cells.
5. Introduce the Gene into the Target Organism: The vector carrying the new gene is introduced into the target organism (such as a plant, animal, or microorganism). This can be done through various methods, such as using a gene gun, or by electroporation (shocking the cells to allow the gene to enter).
6. Select and Grow the Organisms: After the gene has been successfully inserted, the genetic engineers select the organisms that have the new gene and allow them to grow. Only those that successfully received the new gene will be able to show the desired trait.

Techniques used to manipulate DNA Restriction Enzymes (DNA Cutting Tools)

• These enzymes cut DNA at specific sequences, like scissors.
• Scientists use them to cut out genes they want to study or transfer to another organism.

Gel Electrophoresis (DNA Sorting)

• This technique separates pieces of DNA based on their size.
• DNA is placed in a gel and an electric current is applied. Smaller pieces move faster, helping scientists see and analyze the DNA.

Polymerase Chain Reaction (PCR)

• PCR is used to make many copies of a small piece of DNA, like a photocopy machine for DNA.
• It helps scientists study DNA in detail, even if only a small amount is available.

DNA Ligase (DNA Joining)

• DNA ligase is an enzyme used to join two pieces of DNA together.
• It acts like glue, helping scientists combine different DNA pieces to create new combinations.

Gene Cloning

• In this process, a gene is copied and inserted into a host, like bacteria.
• The host reproduces, creating many copies of the gene for study or practical use.

Gene Editing (CRISPR-Cas9)

• This is a modern technique that allows scientists to cut and modify DNA precisely.
• CRISPR is very accurate and is used to fix genetic problems or create new traits.

DNA Transformation

• This involves introducing new DNA into a living organism, like bacteria or plants.
• The organism then uses the new DNA to create a desired trait.

DNA Sequencing

• Scientists use this method to read the exact order of DNA bases (A, T, C, G).
• It helps them understand genes and detect changes or mutations.

Stages

1. Adding the gene the vector DNA.
2. Formation of recombinant DNA molecule with vector.
3. Joining the DNA portions together using DNA ligase enzyme.

Illustration: Consider the diagram below showing the synthesis of insulin A clone is a group of cells of similar characteristics that are able to replicate and produce more cells. Gene cloning is a process whereby multiple copies of a given gene are produced which may then be used to manufacture larger quantities of valuable products. A transgenic organism is a genetically modified organism (GMO) ie: the organism formed as a result of genetic engineering. New genes are added into embryo of an organism. MERITS AND DEMERITS OF GENETIC ENGINEERING Merits of genetic engineering

1. Synthesis of hormones such as insulin, growth hormones etc.
2. Production of vaccine and antibiotics. Already interferon has been synthesized by genetic engineering.
3. Increases plant resistance to pests eg: cotton and potatoes.
4. Improves quality and quantity of animal products such as milk.
5. It offers endless opportunities to manipulate DNA.

Demerits

1. The materials contained in the manipulated DNA, are likely to undergo mutation.
2. The use of genetic engineering in the manufacture of biological weapons is a mis-application of genetics.
3. The use of GMO’s for human consumption is dangerous as it increases cancer chances.
4. Some practices of Genetic engineering may not be in line with ethical and moral values.

Phenylketonuria(PKU)

• PKU is a recessive autosomal condition.
• The disease is due to inability to convert the amino acid phenylalanine to another amino acid, tyrosine.

Phenylalanine Hydrolase. Phenylalanine     (PAH)           Tyrosine. The enzyme PAH is normally there in the liver. As a result of phenylalanine builds up in the body and the excess is converted into toxins which affect mental development. Affected children appear normal at birth because, while in their mother’s uterus during pregnancy, excess phenylalanine moves across the placenta and is removed by the mother’s liver. If not treated soon, harmful effects are noted. Identifying PKU in new born babiesFew days later after birth, blood test is carried on and higher level than normal of phenylalanine is detected.

Genetic Screening

Genetic screening is the process of detecting mutant genes to identify or predict genetic disorders. It is useful in the following situations:

1. Prenatal Diagnosis
   • Used during pregnancy to detect health problems or genetic diseases in the unborn baby.
   • Parents can receive counseling about the quality of life and other potential problems.
   • In some cases, they may opt for termination of pregnancy.
2. Carrier Diagnosis
   • Identifies people who carry a gene for a disease without showing symptoms.
   • Useful for couples planning to have children.
3. Predictive Diagnosis
   • Predicts genetic disorders that might occur later in life, even if there are no current symptoms.
   • Helps in early preparation and medical intervention.

Techniques for Prenatal Genetic Screening

1. Chorionic Villus Sampling (CVS)
   • A small sample of chorion (placental tissue) is collected for examination.
   • Chromosomes are analyzed to detect genetic disorders and determine the baby’s sex.
   • Risk of miscarriage is slightly higher than other techniques.
2. Amniocentesis
   • Involves analyzing the amniotic fluid, which contains cells genetically identical to the fetus.
   • Detects genetic abnormalities with less risk than CVS.

Modern Mendelian Laws

1. 1st Law (Law of Segregation)
   • Pairs of chromosomes separate during meiosis, and only one chromosome from each pair is passed to gametes.
2. 2nd Law (Law of Independent Assortment)
   • During fertilization, alleles (gene pairs) combine randomly to form zygotes.

Applications of Genetics

1. Plant and Animal BreedingProducing hybrids with beneficial traits by crossing genetically different organisms. Example:
   1. Hereford: High beef production and quick maturation.
   2. Boran: Disease resistance and survival on dry pasture.
   3. Hybrid combines these traits for better productivity.
2. Blood Transfusion
   1. Ensures compatibility by matching blood groups and Rhesus factors.
   2. Prevents agglutination (clumping of blood cells).
   3. Used in parentage disputes with the ABO and Rhesus systems.
3. Genetic Counseling
   1. Advises couples with hereditary disorders on the chances of passing these disorders to their children.
   2. Helps in selecting marriage partners or planning pregnancies.
4. Medicine Production
   1. Human genes for insulin and growth hormones are transferred to bacteria to produce these medicines.
   2. Vaccines and blood clotting factors are also produced using genetic engineering.
5. Agriculture
   1. Developing crops that can fix nitrogen without relying on bacteria.
   2. Propagating plants like pyrethrum through tissue culture.
   3. Using genetically modified organisms to break down industrial waste.
6. Genetic Disorders
   1. Detecting fetal deformities to prepare or terminate pregnancy.
   2. Introducing normal genes into embryos to cure genetic disorders like sickle-cell anemia.

Symptoms of Down’s Syndrome

• Mental retardation.
• Straight hair.
• Increased risk of infections.
• Short stature.
• Heart defects.

PLEIOTROPYPleiotropic genes are genes that code for a specific metabolic process and at the same time affecting other metabolic processes. Eg. In cystic fibrosis, A gene that codes for secretion of Cl^–^also induce secretion of viscous (thick) mucus in the lungs, pancreases and gut. Perform below crosses and see if ratios can provide solutions to question (2) (a)

1. Agouti                        x          Albino.     Where; A-Agouti.

AaCc                          Aacc                     C-Black.

1. Agouti                        x          Agouti.

AaCc                          AaCc

1. Agouti                        x          Albino.

AaCa                          aacc Heterogametic sex (XY). In some insects, while the female is XX, the Y chromosome is absent in the male, which is therefore XO. In the fruit fly Drosophila, the female is XX and the male is XY. SEX-LINKAGE

• Sex linkage refers to the carrying of genes on the sex chromosomes.
• The X chromosome carries many such genes, the Y chromosome has very few.
• The non – homologous part of the X – chromosome with Y – chromosome is the one which bears extra genes.

• Those genes that are inherited together with sex chromosomes are called sex – linked characters (traits).
• Two well known sex-linked genes in humans are those causing haemophilia and pale green colour blindness. Both are linked to the X-chromosome and both occur almost exclusively in males.

For the condition to arise in females, requires the double recessive state and as the recessive allele is relatively rare in the population, this is unlikely to occur. In females the recessive allele is normally masked by the appropriate dominant allele which occurs on the other X – chromosome. These heterozygous females are not themselves affected but are capable of passing the recessive allele to their offspring. For this reason such female are termed Carriers. When the recessive allele occurs in males it expresses itself because the Y – chromosome cannot carry any corresponding dominant allele. HAEMOPHILIAHemophilia is the inability of blood to clot, leading to slow and persistent bleeding especially in the joints. Unlike colour blindness it is potentially lethal. Hemophilia is a sex linked character caused by a recessive allele which is carried by the X – chromosome. Consider the following genetic attributes:- X^H^X^H^ – Normal female. X^H^X^h^ – Normal but carrier female. X^h^X^h^ – Hemophilia female. X^H^Y – Normal male. X^h^Y – hemophilia male. Hemophilia females are rare in nature because:-

1. Mostly, they do not grow beyond the first menstrual flow.
2. For a female to have hemophilia, both X – chromosomes must bear the allele h, in the heterozygous state, one is normal.

ExampleConsider a cross between a carrier female and a normal male. From the cross,

1. All the females are phenotypically normal.
2. 50% of the males, are hemophiliac.

Considering a reciprocal cross we have