UPSC Biology Genetics Genetics and Biotechnology

 

GENETICS AND BIOTECHNOLOGY

 

 

GENETICS

 

Heredity is the transmission of genetic characters from parent to off springs. Individuals of same species have some differences, these are called variation.

 

MENDEL’S FINDINGS

1. Mendel’s Law of Dominance and Recessive

• Each of the \[{{f}_{1}}\] generation plant shows inheritance of Y allele from one parent and a G allele from the other. When the \[{{f}_{1}}\] plants breed, each has equal chance of passing on either Y or G allele to each offspring.

In all the seven traits that Mendel examined, one form appeared dominant over the other i.e., it marked the presence of the other allele, e.g.

 

 

BASIC TERMS USED IN INHERITANCE STUDIES ·   Allele: It is an alternative form of a gene which are located on same position (loci) on the homologous chromosome. Term allele was coined by Bateson. ·   Homozygous: A zygote is formed by fusion of two gametes having identical factors is called homozygote and organism developed from this zygote is called homozygous. Ex. TT, RR, tt. ·   Heterozygous: A zygote is formed by fusion of two different types of gamete carrying different factors is called heterozygote (Tt, Rr) and individual developed from such zygote is called heterozygous. ·   Hemizygous: If individual contains only one gene of a pair then individual is said to be hemizygous. Male individual is always hemizygous for sex linked gene. ·   Phenotype: It is the external and morphological appearances of an organism for a particular character. ·   Genotype: It is the genetic constitution or genetic make-up of an organism for a particular character. ·   Back cross: A back cross is a cross in which \[{{F}_{1}}\]individuals are crossed with any of their parents. ·   Test cross: When \[{{F}_{1}}\] progeny is crossed with recessive parent then it is called test cross. ·   Monohybrid test cross: The progeny obtained from the monohybrid test cross are in equal proportion, means 50% is dominant phenotypes and 50% is recessive phenotypes. It can be represented in symbolic forms as follows. \[{{F}_{1}}\]progeny (hybrid) \[\times \] Recessive parent   Monohybrid test cross ratio =1:1 ·   Dihybrid test cross: The progeny is obtained from dihybrid test cross are of four types and each of them is 25%.

 

2. Law of Segregation: According to this law, for any particular trait, the pair of alleles of each parent separate and only one allele passes from each parent to an off spring. Allele in a parent’s pair of allele is inherited as a matter of chance (we now know segregation of alleles occurs during the process of meiosis).
3. Law of Independent Assortment: This is also known as ‘Inheritance law’. According to this law, different pairs of alleles are passed to offspring independently of each other.

 

PARENTAL CROSS

 

 

 

 Exceptions of Conclusions of Mendel

Exception of Dominance

There are two exceptions of law of dominance-

(i)         Incomplete dominance

(ii)         Co-dominance

 

LINKAGE

• Linkage is the phenomenon of certain genes staying together during inheritance through generations without any change or separation. This is due to their location on the same chromosomes.
• Linkage was first time seen by Bateson and Punnett in Lathyrus odoratus and gave coupling and repulsion phenomenon. But they did not explain the phenomenon of linkage.
• Sex linkage was first discovered by Morgan in Drosophila and coined the term linkage. He proposed the theory of linkage.

 

Sex Linkage

When the genes of vegetative/somatic characters are present on sex-chromosome, it is termed as sex linked gene and such phenomenon is known as sex-linkage.

 

	Sex linkage
	\[\downarrow \]                                               \[\downarrow \]
X- linkage		Y- linkage
Genes    of    somatic characters are found on X-chromosome.     The inheritance of X-linked character may be through the males and females, e.g. Haemophilia, Colour blindness.		The genes of somatic characters are located on Y- chromosome. The inheritance of such type of character occur only through the males, such type of character is called holandric character. These characters are found only in male. E.g.: Hypertrichosis (excessive hair on ear pinna.)

GENES

Genes are responsible for the characteristics features (or traits) of organism-plant or animals. The characteristics or traits of parents are transmitted to their progeny (off springs) through geneses present on their chromosomes during the process of sexual reproduction.

• Genes are arranged linearly along long chains of DNA sequence, called chromosomes. The DNA of the chromosome is associated with proteins that organise, compact and control the access to the DNA, forming a material called chromatin. In eukaryotes, chromatin is composed of nucleosomes - segments of DNA wound around histone protein. The full set of hereditary material in an organism i.e., the combined DNA sequences of all chromosomes is called genome.

 

Lethal Gene

• Gene which causes death of individual in early stage when it comes in homozygous condition is called lethal gene.
• It may be dominant or recessive both, but mostly recessive for lethality. Lethal gene was discovered by L. Cuenot in coat colour of mice.

 

MULTIPLE ALLELE

• More than 2 alternative forms of same gene is called as multiple allele. Multiple allele is formed due to mutation and located on same locus of homologous chromosome.

Example of multiple allele

• ABO blood group : ABO blood groups are determined by allele IA, allele IB, allele ID

IA = dominant

IB = dominant

ID = recessive

Possible phenotypes - A, B, AB, O

 

Blood group	Genotype	Antigen or agglutinogen	Antibody or agglutinin
A	\[{{I}^{A}}{{I}^{A}},{{I}^{A}}{{I}^{O}}\]	A	b
B	\[{{I}^{B}}{{I}^{B}},{{I}^{B}}{{I}^{O}}\]	B	a
AB	\[{{I}^{A}}{{I}^{B}}\]	A & B	None
0	\[{{I}^{O}}{{I}^{O}}\]	None	A & b

 

 

Possible genotype number =\[\underline{3\,\,(3\,\,\,\,\,1)}=6\,\,genotype\]

 

SEX DETERMINATION

• Sex determination is a biological system that determines the development of sexual characters in an organism. Most sexual organisms have two sexes: Males and females.
• In a human, the sexual chromosomes complement is 46, 44 of which are autosomes while 2 distinct chromosomes are the sex chromosomes, which determine the sex of an organism and various sex-linked characteristics.
• In humans, sex is predetermined in the sperm gamete. The egg gamete mother cell is said to be homogametic because all its cells possess the XX sex chromosomes, sperm gametes are said to be heterogametic because around half of them contain the X-chromosome and others possess the Y-chromosome to compliment the first X-chromosome.

 

Deoxyribose Nucleic Acid (DNA)

• Deoxyribose nucleic acid (DNA) carries the genetic information. It is a constitute of chromosome.
• Structure of DNA was worked out by X-ray diffraction studies. A double helix model by DNA was proposed by Watson and Crick in 1953. They suggested that:

(i) Each DNA molecule consists of two polynucleotide chains.

(ii) The chains are helically coiled around a common axis.

(iii) DNA molecule has a diameter of 20 A are complete turn of helix is 3.4 A. So there are 10 bases per turn of helix.

(v) Each DNA chain is complementary chain to second chain.

• Deoxyribose and a nitrogeneous base together form a nucleoside. A nucleoside and a phosphate together form a nucleotide.

Nucleoside = Deoxyribose + Nitrogenous base

Nucleotide = Deoxyribose + Nitrogenous base + Phosphate

• Deoxyribose is pentose sugar with five carbon atoms, four of the given carbon atoms plus a single atom of oxygen for a five numbered ring.

 

GENETIC DISORDERS

• A genetic disorders is a disease that is caused by an abnormality in an individual’s DNA.
• Genetic disorder may be grouped into two categories- Mendelian disorders and chromosomal disorders.
• Mendelian disorders are chiefly determined by alteration or mutation in the single gene. Eg. haemophilia, cystic fibrosis, sickle cell anaemia, thalassemia, colour blindness, phenyl ketonuria, etc.

(i)         Haemophilia is an inherited disorder of blood in which essential clotting factor are either partly or completely missing.

(ii)         In sickle-cell anaemia glutamic acid (glutamine) is replaced by valine at the sixth position in b chain of haemoglobin. It is a blood disease. Where the red blood blood cells become sickle shaped as compared to normal one.

• The chromosomal disorders are caused due to absence or excess or abnormal arrangement of one or more chromosomes. Failure of segregation of chromatids during cell division cycle results in the gain or loss of a chromosome(s), called aneuploidy.
• Types of chromosomal disorders are – Down’s Syndrome, Klinefelter’s Syndrome and Turner’s Syndrome.
• Down’s Syndrome is caused by the presence of an additional copy of the chromosome number 21 (trisomy of 21). The affected individual is short statured with small round head, furrowed tongue and partially open mouth. Palm is broad with characteristic palm crease. Physical, psychomotor and mental development is retarded.
• Klinefelter’s Syndrome is caused due to the presence of an additional copy of X-chromosome resulting into a karyotype of 47, XXY. Such an individual has overall masculine development, however, the feminine development (development of breast, i.e., Gynaecomastia) is also expressed. Such individuals are sterile.
• Turner ‘s Syndrome is caused due to the absence of one of the X chromosome, i.e., 45 with XO, Such females are sterile as ovaries are rudimentary besides other features including lack of other secondary sexual characters.

 

BIOTECHNOLOGY

 

The Convention on Biological Diversity (CBD) defines biotechnology as any technological application that uses biological systems, living organisms, or derivatives thereof, to make or modify products or processes for specific use.

 

	Principles of Biotechnology
	\[\downarrow \]                                                 \[\downarrow \]
Genetic engineering		Biochemical engineering
It is the manipulation of genes of an organism. Genetic engineering refers to artificial synthesis, isolation, modification, combination, addition and repair of the genetic material (DNA) to alter the phenotype of the host organism to suit human needs.		These are the processes that help the growth of desired microbe/eukaryotic cell in large quantities in a sterile medium (Tissue culture technique) for the manufacture and multiplication of biotechnological product (antibiotics, vaccines, enzymes, medicines, hormones, etc.)

 

RECOMBINANT DNA TECHNOLOGY

Recombinant DNA technology, popularly known as ‘genetic engineering’ is a stream of biotechnology which deals with the manipulation of genetic material by man in vitro.

There are two distinct techniques/or introducing foreign genetic material into plant cell genome — indirect and direct transfer.

The first is indirect transfer through a vector which requires-

• selection and isolation of the desirable fragment(s) of DNA which contains gene sequence(s) that needs to be cloned known as insert;
• Generation of recombinant DNA (r DNA) molecule by insertion of these inserts (DNA fragments) into a carrier DNA molecule, termed as vector (i.e., the bacterial Agrobacterium tumefaciens, a virus, a plasmid or any other vector) that can replicate within a host cell; Recombinant DNA (r DNA) = Vector + insert
• Introduction of the r DNA molecules into host cells.

 

The second, through direct introduction of DNA, involves-

• Co-cultivation, i.e., culturing the recipient protoplast with purified DNA
• Electroporation, i.e., application of electric impulses to change the porosity of protoplasts so that the directly imbibe the purified DNA.
• Micro-injection, i.e., direct injection of DNA fragments with the help of a micropipette. Other methods of gene transfer are - liposome mediated gene transfer, calcium phosphate precipitation method, transformation by ultrasonifation and transformation using pollen or pollen tube.

Thus, genetic engineering can be defined as the generation of new combination of heritable material by the insertion of desired genes or DNA of the cell, into any carrier system so as to allow their incorporation into a host organism in which they do not normally of cur but in which they are able to perform normal behaviour and propagation.

 

Basic Tools of Recombinant DNA Technology

Enzyme

A number of specific kind of enzymes are employed in genetic engineering. These include lysing enzymes, cleaving enzymes, synthesizing enzyme and joining enzymes.

• Lysing enzymes: These are used to open up the cells to get DNA for genetic experiments. It is commonly used to dissolve the bacterial cell wall.
• Cleaving enzyme: These are used to break DNA molecule.

They are further of 3 kinds—

(i) Exonucleases - which cut off nucleotides from 5’ or ‘3 ends of DNA molecule;

(ii) Restriction endonucleases — which cleave DNA duplex at specific points called restriction sites in such a way that single-stranded free ends project from each fragment of DNA duplex. These single-stranded free ends are called ‘sticky ends’ because they can join similar complementary ends of DNA fragment from some other source. Restriction endonuclease is called molecular scissors or a chemical scalpel.

 3’- CTTAAG-5’           5’-GAATTC – 3’

 

• Joining enzyme: These enzymes help in joining the DNA fragment. For example DNA ligase from Escherichia coil is used to join DNA fragments by forming a phosphodiester bond.

 

 

 

Vectors

• Vectors are cloning vehicle required to transfer DNA of interest from one organism to another.
• Desirable properties of cloning vector are -

High copy number, Presence of origin of replication (Ori), Presence of selectable marker, Presence of unique recognition site or cloning site and Ability to sustain in bacterial cell

 

• Ori is sequence where replication starts and any piece of DNA linked here will be replicated. Ori also controls copy number of vector.
• Selectable marker allow to select those host cells that contain the vector amongst those which do not. Selectable markers helps in identifying eliminating nontransformants & selectively permitting the growth of transforments.
• In cloning site, vector should have single recognition site for restriction enzyme. Presence of more than one recognition site will cut the vector into many fragments.
• Transformation efficiency of vector is percentage of competent bacterial host cell receiving desired DNA at specific recognition sequences which are palindromic.

 

 

Process of Recombinant Technology

Recombinant DNA (rDNA) is DNA created artificially by combining the DNA from two or more organisms into a single “recombinant”   molecule. The term "recombinant DNA technology" also commonly known as 'DNA cloning,’ ‘molecular cloning' and 'gene cloning’, refers to the transfer of a segment of DNA from one organism to another organism (the “host cell”) where it reproduces.

 

Following are the steps in Recombinant DNA Technology

Step 1: Isolation of genetic material

At first required DNA sequence from the donor cell is identified. Later the required DNA is cut by the help of restriction enzymes at specific site, which result is single stranded sequence with “stick ends”.

Step 2: Cutting of DNA of specific locations

Second, cloning vectors from the host cell are identified and removed with the same restrictive enzyme applied to the donor DNA. Cloning vectors are DNA molecules in which another DNA fragment (i.e., foreign DNA) can be integrated and which are capable of independently replicating themselves and the foreign DNA once inserted into the host cell.

Step 3: Amplification of Gene of interest using PCR

PCR stands for Polymerase Chain Reaction. It is the process of amplifying a desired gene of interest into a number of copies. In this reaction, multiple copies of the gene (or DNA) of interest is synthesised in vitro using two sets of primers (small chemically synthesised oligonucleotides that are complementary to the regions of DNA) and the enzyme DNA polymerase.

Step 4: Insertion of Recombinant DNA into the Host Cell/Organism

There are several methods of introducing the ligated DNA into recipient cells. Recipient cells after making them ‘competent’ to receive, take up DNA present in its surrounding. So, if a recombinant DNA bearing gene for resistance to an antibiotic (e.g. ampicillin) is transferred into E. coli cells, the host cells become transformed into ampicillin-resistant cells.

Step 5: Obtaining the Foreign Gene Product

After having cloned the gene of interest and having optimised the conditions to induce the expression of the target protein, one has to consider producing it on a large scale. If any protein encoding gene is expressed in a heterologous host, is called a recombinant protein.

 

 APPLICATION OF BIOTECHNOLOGY

 

BT COTTON Bt means Bacillus thuringiensis. It is a gram positive dwelling bacterium. It produces crystal [Cry] protein. This Cry protein is toxic to larvae of certain insects. Each Cry protein is toxic to a different group of insects. The gene encoding cry protein is called cry gene. This Cry protein is isolated and transferred into several crops. A crop expressing a cry gene is usually resistant to the group of insects for which the concerned Cry protein is toxic. There are a number of them, for example, the proteins encoded by the genes cry IAc and cry lAb control the cotton bollworms, that of cry IAb controls corn borer. Some strains of Bacillus thuringiensis produce proteins that kill certain insects such as lepidopterans (tobacco budworm, armyworm), coleopterans (beetles) and dipterans (flies, mosquitoes). B. thuringiensis forms protein crystals during a particular phase of their growth. These crystals contain a toxic insecticidal protein. The Bt toxin protein exist as inactive protoxins but once an insect ingest the inactive toxin, it is converted into an active form of toxin due to the alkaline pH of the gut which solubilise the crystals. The activated toxin binds to the surface of midgut epithelial cells and create pores that cause cell swelling and lysis and eventually cause death of the insect.

 

Problems related to GM foods are

• The transgene product may toxicity and or produce allergies.
• The enzyme produced by the antibiotic resistance gene could cause allergies, since it is a foreign protein.
• The bacteria present in the alimentary canal of the humans could take up the antibiotic resistance gene that is present in the GM food.

 

Application of Biotechnology in Medicine

• The rDNA technology has been used in the production of safe and more effective therapeutic drugs.
• The recombinant therapeutics do not induce unwanted immunological responses that are commonly observed with similar products isolated from non-human sources.

 

Genetically engineered insulin (humulin)

• Human insulin consists of two short polypeptide chains: chain A and chain B, linked by disulphide bridges.
• Insulin is secreted as prohormone which has to be processed before it becomes a mature and functional hormone.

 

Transgenic Animals

• Transgenic animals are those animals that have had their DNA manipulated to possess and express a foreign gene.
  • Transgenic animals are used in the following ways :

(i) Transgenic animals can be specifically designed to allow the study of how genes are regulated and how they affect the normal functions of the body and its development, e.g., Information is obtained about the biological role of insulin like growth factor.

(ii) Transgenic animals are designed to increase our understanding of how genes contribute to the development of diseases; they are made to serve as models for human diseases.

(iii) Transgenic mice are being developed for use in testing the safety of vaccines, (e.g. polio vaccine). (iv) Transgenic animals with more sensitivity to toxic substances are being developed to test the toxicity of drugs.

 

Transgenic Plants

Transgenic Plants (of some species,) are easier to produce because plants have a lot oftotipotent tissue (meristem) that can be grown in culture and then induced to develop into a whole plant. One of the most common methods utilizes a natural gene transfer system called the Ti-plasmid of Agrobacterium tumefaciens which transfers a piece of DNA called T-DNA into the genome of infected plants.

 

Table: Transgenic plants

 

Transgenic plants	Useful application
Bt Cotton	Pest resistance, herbicide tolerance and high yield. It is resistant to boll worm infestation.
Flavr Savr Tomato	Increased shelf-life (delayed ripening) and better nutrient quality.
Golden rice	Vitamin A-rich
Potato	Higher protein content
Corn, Brinjal	Insect resistance
Soyabeen, Maize	Herbicide resistance

 

 

Ethical Issues

• Genetic modification of organisms can have unpredictable/undesirable effects when such organisms are introduced into the ecosystem.
• The modification and use of such organisms for public services has also resulted in problems with the granting of patents.
• One such organisation is the Genetic Engineering Approval Committee (GEAC).

 

Bio piracy

• Basmati rice grown in India is distinct for its unique flavour and aroma, but an American company got patent rights on Basmati through the US patent and trademark office; the new variety of Basmati has been developed by this company by crossing an Indian variety with the semi-dwarf varieties.
• Now some nations are developing laws to prevent such unauthorised exploitation of their bio resources and traditional knowledge.
  • Some such developed countries use the bio resources and traditional knowledge of the other countries without proper authorisation and/or compensation to the countries concerned (Bio piracy)