INDEX

1. Theory 03 - 45

2. Exercise - 1 46 - 54

3. Answers Key 55
 THEORY
Molecular Basis of Inheritance

Fig 6.1: Molecular basis of inheritance

● Molecular basis of inheritance is the study of genes, hereditary and genetic variations which explains how an offspring looks
similar to its maternal or paternal features.
● Deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) are the two types of nucleic acids found in living systems.
● DNA acts as the genetic material in most of the organisms. RNA though it also acts as a genetic material in some viruses,
mostly functions as a messenger.
● RNA has additional roles as well. It functions as an adapter, structural, and in some cases as a catalytic molecule.
● In this chapter we are going to discuss the structure of DNA, its replication, the process of making RNA from DNA
(transcription), the genetic code that determines the sequences of amino acids in proteins, the process of protein synthesis
(translation) and the elementary basis of their regulation.
● The determination of the complete nucleotide sequence of the human genome during the last decade has set in a new era
of genomics.
 4 MOLECULAR BASIS OF INHERITANCE

1. The DNA

Fig 6.2: The DNA

● Genetic material controls the formation and expression of traits in an organism. It can replicate and pass on from one
generation to another.
● DNA or deoxyribonucleic acid represents the genetic material of an organism which contains the genes and is responsible for
passing down characters from generation to generation.

Fig 6.3: DNA in prokaryotes and eukaryotes

MOLECULAR BASIS OF INHERITANCE 5

● The length of DNA is usually defined as the number of nucleotides (or a pair of nucleotides referred to as base pairs) present
in it.
Length of DNA in Different Organisms

Organisms Length of DNA

Bacteriophage φ ×174 5386 nucleotides
Bacteriophage lambda 48502 base pairs (bp)
Escherichia coli 4.6 × 106 bp
Haploid content of human DNA 3.3 × 109 bp

1.1 Structure of Polynucleotide Chain

● DNA is a long polymer of deoxyribonucleotides.
● Nucleotides are the building blocks of nucleic acids (DNA or RNA).
● A nucleotide has three components:
● A nitrogenous base
● A pentose sugar
● A phosphate group

Fig 6.4: Structure of a nucleotide

1.1.1 Nitrogenous Base

● Nitrogenous bases are simply a nitrogen-containing heterocyclic molecule that has the same chemical properties as a base.
There are two types of nitrogenous bases - purines and pyrimidines.

Differences between Purine and Pyrimidine

Purine
1 Purines are larger-sized nitrogen
containing biomolecules.
2 A purine is a nine-membered structure.
3 It is a double ring.
4 A purine contains four nitrogen atoms at
1, 3, 7 and 9 positions.
5 Purine bases are of two types, adenine
(A) and guanine (G).
Pyrimidine
1 Pyrimidines are smaller-sized nitrogen containing biomolecules.
2 A pyrimidine is a six-membered structure.
3 It is a single ring.
4 A pyrimidine has nitrogen atoms at two places, 1 and 3 positions.
5 Pyrimidine bases are of three types - cytosine (C), thymine (T) and
uracil (U).
 6 MOLECULAR BASIS OF INHERITANCE

Fig 6.5: Types of purines and pyrimidines

1.1.2 Pentose Sugar

● Nucleic acids contain a 5-C sugar molecule that binds to nitrogenous bases.
● In the case of RNA, the sugar is ribose.
● However, in the case of DNA, the ribose sugar has an H atom instead of OH at it’s 2’ carbon position. Hence in DNA, the
sugar is called deoxyribose (one oxygen missing).

Fig 6.6: Pentose sugar in RNA and DNA

MOLECULAR BASIS OF INHERITANCE 7

1.1.3 Bonds in a DNA

● There are three types of bonds found in a DNA molecule:
● Hydrogen bonds – In a DNA molecule, purines bond with pyrimidines through hydrogen bonds. Adenine bonds with Thymine
(T) through two H-bonds, while Guanine bonds with Cytosine (C) through three H-bonds.

Fig 6.7: Hydrogen bonds between purines and pyrimidines

● Glycosidic bonds – A bond between a nitrogenous base and a pentose sugar is called a glycosidic bond. The 1’ carbon of the
sugar molecule is bonded to a nitrogen atom of the base. Therefore, the bond is also called as N-glycosidic linkage.

Fig 6.8: Glycosidic bond

 8 MOLECULAR BASIS OF INHERITANCE

● Phosphodiester bonds – Adjacent nucleoside pairs are connected together to form long polynucleotide chains via phosphate
groups. These groups attach to the hydroxyl groups at the 5’C of one sugar molecule and at 3’C of the next sugar molecule.
The bond between a phosphate group and the hydroxyl group is termed as an ester bond (O-R). Hence, these bonds are called
3’-5’ phosphodiester bonds.

Fig 6.9: Phosphodiester bond

MOLECULAR BASIS OF INHERITANCE 9

1.1.4 DNA Double Helix Model

● In 1869, Friedrich Miescher discovered the DNA as an acidic substance present in nucleus and called it ‘Nuclein’.
● Rosalind Franklin and Maurice Wilkins gave the X-ray diffraction pattern of DNA.
● In 1953, Watson and Crick proposed the double helical model of DNA. This was based on the findings of Erwin Chargaff
(known as Chargaff’s rules) and X-ray diffraction studies.
● Erwin Chargaff pointed out that in a particular DNA molecule, the total number of purines is always equal to the total number
of pyrimidines.
A+G=C+T
● The number of A-T bonds differ from the number of G-C bonds, but the base ratio (A+T/G+C) is constant for a species but
may vary from one species to another. This ratio can therefore be used to identify a species.

Fig 6.10: Chargaff’s rule

NOTE:

The phosphodiester bonds in the backbone and hydrogen bonds between the bases make the DNA a double helical structure.

Fig 6.11: DNA double helix model

10 MOLECULAR BASIS OF INHERITANCE

Salient features of the Double-helix structure of DNA are as follows:

● It is made of two polynucleotide chains, where the backbone is constituted by sugar-phosphate, and the bases project inside.

Fig 6.12: Sugar phosphate backbone of DNA

● The two chains have antiparallel polarity. One chain has the polarity 5'→3', the other has 3'→5'.
MOLECULAR BASIS OF INHERITANCE 11

Fig 6.13: Antiparallel polarity of DNA

● The bases in two strands are paired through hydrogen bonds forming base pairs (bp). A purine always comes opposite to a
pyrimidine. This generates approximately uniform distance between the two strands of the helix.

Fig 6.14: Hydrogen bonding in DNA

● The two chains are coiled in a right-handed fashion. The pitch of the helix is 3.4 nm and there are roughly 10 bp in each turn.
Consequently, the distance between a bp in a helix is approximately 0.34 nm.
 12 MOLECULAR BASIS OF INHERITANCE

Fig 6.15: Pitch of DNA

● The plane of one base pair stacks over the other in a double helix. This, in addition to H-bonds, confers stability of the
helical structure.

1.2 Central Dogma of Molecular Biology

● Francis Crick proposed the Central dogma in molecular biology. It represents the scheme which shows how information in the
genes is used to form proteins, which ultimately forms all the structural and functional features of an organism.

Fig 6.16: Central dogma of molecular biology

MOLECULAR BASIS OF INHERITANCE 13

1.2 Packaging of DNA Helix

Fig 6.17: Packaging of DNA double helix in a cell

● Taken the distance between two consecutive base pairs as 0.34 nm (0.34×10–9 m), if the length of DNA double helix in a
typical mammalian cell is calculated (6.6 × 109 bp × 0.34 × 10-9 m/bp), it comes out to be approximately 2.2 meters.
● In order to accomodate a DNA molecule inside a cell, it needs to be compacted and packaged.
● Histones are positively charged proteins which are rich in basic amino acids like Lysine and Arginine, that help in binding to
the negatively charged DNA backbone.
● There are five types of histone proteins: H1, H2A, H2B, H3, H4

Fig 6.18: Nucleosome showing different histones

● DNA molecules coil around the histone octamer, forming the nucleosome core.
● Multiple nucleosome cores are joined via a linker DNA.
● This DNA-histone complex appears like “Beads on a String” under electron microscope.
14 MOLECULAR BASIS OF INHERITANCE

Fig 6.19: Beads on string - EM picture

● The beaded string is coiled to form a cylindrical coil or solenoid having 6 nucleosomes per turn, which forms a solenoid of 30
nm diameter. The solenoid structure undergoes further supercoiling to produce a chromatin fibre.
● The packaging of chromatin at a higher level requires an additional set of proteins, that are referred to as Non-histone
chromosomal (NHC) proteins.

Fig 6.20: Non-histone chromosomal (NHC) proteins

● Higher order chromatin structures can be of two types:

Euchromatin
It is a relatively loosely packaged area of DNA,
which RNA or protein molecules can access, and
copy the information stored in there to form
proteins.
The euchromatin area contains actively expressing
genes.
Heterochromatin
It is a part of DNA which has a highly condensed chromatin
structure. RNA or protein molecules cannot usually access
the heterochromatin area, and therefore the information in
such parts of the DNA are not passed on to form proteins.
Heterochromatin usually contains silenced or inactive genes.
MOLECULAR BASIS OF INHERITANCE 15

2. The Search for Genetic Material

● Previous discoveries by Gregor Mendel, Walter Sutton, Thomas Hunt Morgan and numerous other scientists had narrowed the
search to the chromosomes located in the nucleus of most cells.
● There were a number of experiments that provided conclusive evidence that the chief genetic material in nearly all life forms
are the nucleic acids, and mainly the DNA. These experiments include:
● Griffith’s transformation experiment
● Hershey-Chase experiment

2.1 Griffith’s Transforming Principle

Principle:
● Griffith's experiment, reported in 1928 by Frederick Griffith, was the first experiment suggesting that bacteria are capable of
transferring genetic information through a process known as transformation.
● In Griffith’s experiment, he used two strains of pneumococcus (Streptococcus pneumoniae or Diplococcus pneumoniae)
bacteria which infect mice – a type III-S (smooth) which was virulent, and a type II-R (rough) strain which was nonvirulent.
The III-S strain synthesized a polysaccharide capsule that protected itself from the host's immune system, resulting in the death
of the host, while the II-R strain did not have that protective capsule and was defeated by the host's immune system.
● Non-virulent R-strain (R-II) does not cause pneumonia
● Virulent S-strain (S-III) - causes pneumonia
Methods
16 MOLECULAR BASIS OF INHERITANCE

Observations
Griffith observed that the live pathogen could kill the mice but the dead (heat-killed) pathogen couldn’t. However, when the dead
pathogen was injected along with a live R-strain, it’s virulent genes got transferred to the R-strain, turning the avirulent strain
virulent. Thus the donor’s genetic material could transform the recipient.

Fig 6.21: Observations of Griffith’s experiment

2.2 Biochemical Characterisation of Transforming Principle
● Avery, MacLeod and McCarty purified proteins, DNA and RNA from heat-killed S-cells and inserted them one by one into
live R-cells.
● They observed that whenever the S-strain DNA was not inserted into R, the mice lived, signifying lack of transformation.
● They also discovered that protein-digesting enzymes (proteases) and RNA-digesting enzymes (RNases) did not affect
transformation, so the transforming substance was not a protein or RNA.
● Thus, they concluded that DNA is the hereditary material.
MOLECULAR BASIS OF INHERITANCE 17

Fig 6.22: Avery, MacLeod and McCarty Experiment

2.3 Hershey Chase Experiment

Fig 6.23: Martha Chase and Alfred Hershey

● The unequivocal proof that DNA is the genetic material came from the experiments of Alfred Hershey and Martha
Chase (1952).
● They worked with viruses that infect bacteria called bacteriophages. The bacteriophage attaches to the bacteria and its genetic
material then enters the bacterial cell. The bacterial cell treats the viral genetic material as if it was its own and subsequently
manufactures more virus particles.
● Hershey and Chase worked to discover whether it was protein or DNA from the viruses that entered the bacteria.
● They grew some viruses on a medium that contained radioactive phosphorus and some others on a medium that contained
radioactive sulfur.
● Viruses grown in the presence of radioactive phosphorus contained radioactive DNA but not radioactive protein because DNA
contains phosphorus but protein does not.
● Similarly, viruses grown on radioactive sulfur contained radioactive protein but not radioactive DNA because DNA does not
contain sulfur.
● Radioactive phages were allowed to attach to E. coli bacteria.
● Then, as the infection proceeded, the viral coats were removed from the bacteria by agitating them in a blender.
● The virus particles were separated from the bacteria by spinning them in a centrifuge.
● Bacteria which were infected with viruses that had radioactive DNA were radioactive, indicating that DNA was the material
that passed from the virus to the bacteria.
● Bacteria that were infected with viruses that had radioactive proteins were not radioactive. This indicates that proteins did not
enter the bacteria from the viruses.
● DNA is therefore the genetic material that is passed from virus to bacteria.
18 MOLECULAR BASIS OF INHERITANCE

Fig 6.24: Hershey and Chase experiment

3. Properties of Genetic Material (DNA Versus RNA)

● A genetic material is defined as a hereditary material that can store the information regarding the functioning of a living
organism and pass down that information to its subsequent generations.
● Requirements of a genetic material:
● It should be able to generate its replica (Replication).
● It should be stable chemically and structurally.
● It should provide the scope for slow changes (mutation) that are required for evolution.
● It should be able to express itself in the form of 'Mendelian Characters’.
3.1 Difference Between DNA and RNA

Fig 6.25: Differences between DNA and RNA

MOLECULAR BASIS OF INHERITANCE 19

DNA RNA

1 It usually occurs inside the nucleus and some cell 1 Very little RNA occurs inside the nucleus.
organelles.

2 DNA is the genetic material. 2 RNA is not the genetic material except in certain viruses,
e.g., Reovirus.

3 It is double stranded with the exception of some 3 RNA is single stranded with the exception of some viruses
viruses (e.g., ϕ x 174). (e.g., double stranded in Reovirus).

4 DNA contains over a million nucleotides. 4 Depending upon the type, RNA contains 70 - 12000
nucleotides.

5 DNA is of only two types; intra-nuclear and 5 There are at least three types of RNAs - mRNA, rRNA and
extranuclear. tRNA.

6 It contains deoxyribose sugar. 6 It contains ribose sugar.

7 Nitrogen base thymine occurs in DNA along with 7 Thymine is replaced by uracil in RNA. The other three are
three others - adenine, cytosine and guanine. similar - adenine, cytosine and guanine.

8 Renaturation after melting is slow. 8 It is quite fast.

9 Hydrogen bonds are formed between 9 Base pairing through hydrogen bonds occurs only in the
complementary nitrogen bases of the opposite coiled parts.
strands of DNA (A-T, C-G).

10 DNA is spirally twisted to produce a regular helix. 10 The strand may get folded at places to produce a secondary
helix or pseudo helix.

11 It replicates to form new DNA molecules. 11 It cannot normally replicate itself.

12 DNA replication requires a primer. 12 No primer is needed during the formation of RNA.

13 DNA transcribes genetic information to RNA. 13 RNA translates the transcribed message for forming
polypeptides.

14 DNA controls metabolism and genetics including 14 It only controls metabolism under instructions from DNA.
variations.

15 Purine and pyrimidine bases are in equal numbers. 15 There is no proportionality between the number of purines
and pyrimidine bases.

16 It is long lived. 16 Some RNAs are very short lived while others have
somewhat longer life.

17 It can be hydrolysed by DNA-ase. 17 It is hydrolysed by RNA-ase.

4. RNA World
20 MOLECULAR BASIS OF INHERITANCE

Fig 6.26: RNA

● Though DNA is the most prevalent genetic material, the first genetic material to evolve on earth was Ribonucleic acid
or RNA.
● The RNA world theory states that in the earliest life forms, an RNA could carry out all the functions responsible for the
functioning of a living organ.
● Experimental evidence proves that essential life processes like translation, splicing etc. evolved around RNA.
● It can also function as a catalyst in the form of ribozymes.
● But, RNA being a catalyst was reactive and hence unstable.
● Therefore, DNA has evolved from RNA with chemical modifications that make it more stable.

5. DNA Replication
● Replication or Duplication of DNA is the process by which a new DNA strand is formed from an existing template
DNA strand.
● In 1953, when Watson and Crick gave the double helical structure of DNA, they also suggested a copying mechanism, in
which the two old strands would separate and each would serve as a template for the synthesis of a new strand.
● Each resulting DNA molecule has one old strand and one new strand. Since in each DNA molecule, half of the parental strands
remain conserved, it is also called the Semi-conservative mode of DNA replication.

Fig 6.27: Semiconservative mode of DNA Replication

5.1 The Experimental Proof: Meselson and Stahl Experiment

● In 1958, scientists Meselson and Stahl experimentally proved that DNA replicates by the semi-conservative mode, as theorized
by Watson and Crick.
MOLECULAR BASIS OF INHERITANCE 21

Fig 6.28: Meselson and Stahl’s experiment

● Matthew Meselson and Franklin Stahl performed the following experiment in 1958.
● They grew E. coli in a medium containing 15NH4Cl (15N is the heavy isotope of nitrogen) as the only nitrogen source for
many generations.
● The result was that 15N was incorporated into newly synthesized DNA (as well as other nitrogen containing compounds).
● This heavy DNA molecule could be distinguished from the normal DNA by centrifugation in a cesium chloride (CsCl) density
gradient as 15N is not a radioactive isotope, and it can be separated from 14N only based on densities.
● Then they transferred the cells into a medium with normal 14NH4Cl and took samples at various definite time intervals as the
cells multiplied and extracted the DNA that remained as double-stranded helices.
● The various samples were separated independently on CsCl gradients to measure the densities of DNA.
● Thus, the DNA that was extracted from the culture one generation after the transfer from 15N to 14N medium [that is after 20
minutes; E. coli divides in 20 minutes] had a hybrid or intermediate density.
● DNA extracted from the culture after another generation [that is after 40 minutes, II generation] was composed of equal
amounts of this hybrid DNA and of ‘light’ DNA.
● If E. coli was allowed to grow for 80 minutes then the proportions of light and hybrid densities of the DNA molecule will be
7:1 or 88% : 12%.

5.2 Machinery and the Enzymes

● In living cells, such as E. coli, the process of replication requires a set of catalysts (enzymes).
● The main enzyme is referred to as DNA-dependent DNA polymerase, since it uses a DNA template to catalyse the
polymerisation of deoxynucleotides. In addition to DNA-dependent DNA polymerases, many additional enzymes are required
to complete the process of replication with high degree of accuracy.
● These enzymes are highly efficient enzymes as they have to catalyse polymerisation of a large number of nucleotides in a very
short time. E. coli that has only 4.6 ×106 bp (compare it with humans whose diploid content is 6.6 × 109 bp), completes the
process of replication within 18 minutes; that means the average rate of polymerisation has to be approximately 2000 bp
per second.
● Not only do these polymerases have to be fast, but they also have to catalyse the reaction with a high degree of accuracy. Any
mistake during replication would result in mutations.

Initiation
● For long DNA molecules, since the two strands of DNA cannot be separated in its entire length (due to very high energy
requirement), the replication occurs within a small opening of the DNA helix, referred to as replication fork.
22 MOLECULAR BASIS OF INHERITANCE

● Replication begins at a particular spot called the origin of replication or Ori.

● Single-stranded DNA is stabilized by single-strand binding proteins (SSBP), so that they don’t re-anneal.
● Topoisomerases are enzymes that relieve these supercoils in the DNA, by making cuts in the DNA.

Fig 6.29: Functioning of helicase, SSBP and topoisomerase enzymes

Elongation

Fig 6.30: Elongation process in DNA replication

● DNA polymerase III can synthesize a new DNA strand only in the 5’ → 3’ direction.
● This creates some additional complications at the replication fork.
● Consequently, on one strand (the template with polarity 3'→ 5'), the replication is continuous, while on the other (the template
with polarity 5'→ 3'), it is discontinuous.
● Therefore, the synthesis of one strand is continuous and it is called the Leading strand and the synthesis of another strand is
discontinuous called the Lagging strand.
● Fragments formed on the lagging strand, also called Okazaki fragments, are then joined to each other by DNA ligase.
● In eukaryotes, the replication of DNA takes place at S-phase of the cell-cycle. The replication of DNA and cell division cycle
should be highly coordinated. A failure in cell division after DNA replication results in polyploidy (a chromosomal anomaly).

Differences between Leading strand and Lagging strand

Leading strand Lagging strand

MOLECULAR BASIS OF INHERITANCE 23

1 It is a replicated strand of DNA which grows 1 It is a replicated strand of DNA which is formed in short
continuously without any gap. segments called okazaki fragments. Its growth is discontinuous.

2 It does not require DNA ligase for its growth. 2 It is required for joining okazaki fragments.

3 The direction of growth of the leading strand 3 The direction of growth of the lagging strand is 3’→ 5’ though
is 5’→ 3’. in each okazaki fragment it is 5’→ 3’.

4 Only a single RNA primer is required. 4 Starting each okazaki fragment requires a new RNA.

5 Formation of the leading strand is quite rapid. 4 Formation of lagging strands is slower.

6 Its template opens in the 3'→ 5’ direction. 6 Its template opens in the 5'→ 3’ direction.

7 Formation of the leading strand begins 7 Formation of the lagging strand begins a bit later than that of
immediately at the beginning of replication. the leading strand.

NOTE:

Since one strand is continuously synthesized and the other isn’t, DNA replication is also semi-discontinuous or
semi-continuous.

6. Transcription

Fig 6.31: Transcription

● The process of copying genetic information from a template strand of DNA into RNA is called transcription.
● Here also, the principle of complementarity governs the process of transcription, except the adenosine complements now form
base pairs with uracil instead of thymine.
● However, unlike in the process of replication, which once set in, the total DNA of an organism gets duplicated, in transcription
only a segment of DNA and only one of the strands is copied into RNA.
● This necessitates defining the boundaries that would demarcate the region and the strand of DNA that would be transcribed.

Why are both the strands not copied during transcription?

It has the simple answer
● First, if both strands act as a template, they would code for RNA molecules with different sequences (Remember
complementarity does not mean identical), and in turn, if they code for proteins, the sequence of amino acids in the proteins
would be different. Hence, one segment of the DNA would be coding for two different proteins, and this would complicate the
genetic information transfer machinery.
24 MOLECULAR BASIS OF INHERITANCE

● Second, the two RNA molecules if produced simultaneously would be complementary to each other, hence would form a
double stranded RNA. This would prevent RNA from being translated into protein and the exercise of transcription would
become a futile one.

6.1 Transcription Unit

● A transcription unit is defined as a segment of DNA between the sites of initiation and termination of transcription by an RNA
polymerase.
● A transcription unit in DNA is defined primarily by the three regions in the DNA
● A Promoter
● The Structural gene
● A Terminator

Fig 6.32: Transcription Unit

● Promoter - It is a region on a DNA molecule to which an RNA polymerase binds and initiates transcription. Transcription
takes place from the 3’ → 5’ end of the template DNA strand and a promoter is located “upstream” to the structural gene.
● Structural gene - The region of template DNA that contains the necessary information that needs to be copied into the mRNA
is the structural gene.
● Terminator - It occurs downstream to the structural gene, that is, towards the 5’ end of the template strand. Terminator is the
sequence where transcription comes to an end.
● In case of transcription, a DNA strand serves as a template for the synthesis of an RNA strand. Therefore, the enzyme involved
is called a DNA-dependent-RNA polymerase.
● There is a convention in defining the two strands of the DNA in the structural gene of a transcription unit. Since the two
strands have opposite polarity and the DNA-dependent RNA polymerase also catalyse the polymerisation in only one
direction, that is, 5'→3', the strand that has the polarity 3'→5' acts as a template, and is also referred to as template strand.
● The other strand which has the polarity (5'→3') and the sequence same as RNA (except thymine at the place of uracil), is
displaced during transcription. Strangely, this strand (which does not code for anything) is referred to as coding strand. All the
reference points while defining a transcription unit are made with coding strand.

Differences between Template Strand and Coding Strand

Template Strand Coding Strand

1 Template strand is directed in the 3’ to 5’ direction. 1 Coding strand is directed in the 5’ to 3’ direction.

2 Transcribed into mRNA. 2 Not transcribed into mRNA.

3 Contains the complementary nucleotide sequence 3 Contains the same nucleotide sequence to mRNA, except
as the mRNA. thymine at the place of uracil.

4 Hydrogen bonds are formed between the template 4 No hydrogen bonds are formed between the coding strand
strand and the synthesizing mRNA temporarily and the synthesizing mRNA during transcription.
during transcription.

6.2 Transcription Unit and The Gene

MOLECULAR BASIS OF INHERITANCE 25

● A gene is defined as the functional unit of inheritance. It is a segment of a DNA or a chromosome at a specific locus, which
carries coded information associated with a specific function.
● In 1948, Beadle and Tatum stated that a gene is responsible for synthesizing an entire enzyme. This came to be known as the
one gene-one enzyme hypothesis.
● In 1965 it was found that a structural gene can govern the synthesis of one polypeptide, and multiple such polypeptides can
unite to form a functional enzyme. Therefore, the one gene-one enzyme hypothesis was correctly replaced by the One gene-
One polypeptide hypothesis.
● It was soon discovered that genes (or segments of DNA) not only code for polypeptides, but can also synthesize different RNA
molecules. Therefore, to distinguish protein coding regions of the DNA from others, the term gene has therefore been replaced
by the term “cistron”.
● In eukaryotes, the monocistronic structural genes have interrupted coding sequences – the genes in eukaryotes are split.
● The coding sequences or expressed sequences are defined as exons. Exons are said to be those sequences that appear in mature
or processed RNA. The exons are interrupted by introns.
● Introns or intervening sequences do not appear in mature or processed RNA.
● Depending on how many polypeptide chains can be synthesized from the structural gene region, they can be classified into:

Monocistronic Gene Polycistronic Gene

Those genes from which only one polypeptide can be Those genes from which more than one polypeptide can
synthesized. be synthesized.

They are found in eukaryotes. They are found in prokaryotes

They have interrupted coding sequences, or in other words, They do not have interrupted coding sequences.
the eukaryotic genes are split.
26 MOLECULAR BASIS OF INHERITANCE

Fig 6.33: Monocistronic and polycistronic gene

6.3 Types of RNA
● In bacteria, there are three major types of RNAs:
● mRNA (messenger RNA)
● tRNA (transfer RNA)
● rRNA (ribosomal RNA)
● All three RNAs are needed to synthesize a protein in a cell.

Fig 6.34: Three types of RNA - mRNA, rRNA, and tRNA

Differences between mRNA, rRNA and tRNA

mRNA or Messenger RNA rRNA or Ribosomal RNA tRNA or Transfer RNA

MOLECULAR BASIS OF INHERITANCE 27

1. It accounts for about 5% of total RNA in 1. It accounts for about 80% of total 1. It accounts for about 15% of total
the cell. RNA in the cell. RNA in the cell.

2. It consists of 75 - 6000 bases. 2. It consists of 100 - 5000 bases. 2. It consists of 73 - 93 bases.

3. It's mol. wt. is 25000 - 2000000 daltons. 3. It's mol. wt. is 35000 - 1800000 3. It's mol. wt. is about 25000 daltons.
daltons.

4. Its sedimentation coefficient is 6 - 30 S. 4. Its sedimentation coefficient is 5S, 4. Its sedimentation coefficient is 4S.
5.8S, 28S and 18S in eukaryotes; 5S,
16S and 23S in prokaryotes.

5. It is moderate to large sized with moderate 5. It is smaller; moderate to large, 5. It is smallest and coiled like a
to maximum mol. weight but is least which is most abundant and highly clover leaf.
abundant. coiled.

6. It carries a coding message for many 6. It carries no coding message. 6. It carries a coding message for only
amino acids. one amino acid.

7. It is linear and never coiled. 7. It is linear and coiled. 7. It is folded.

8. It is synthesized by RNA polymerase II in 8. Its synthesis occurs in the nucleolus 8. It is synthesized by RNA
the nucleus. by RNA polymerase I. polymerase III in the nucleus.

9. It has no modification of bases in the 9. Modification of bases is very less. 9. About 5% bases are modified.
coding region.

10. It is of various types depending upon the 10. It is of 3 or 4 types. 10. It is of about 100 types.
number of genes.

11. It is short lived (3 seconds to a few days) 11. It is most stable, used again and 11. It is quite stable, used again and
and commonly degrades after protein again and does not degrade. again, and degrades very slowly.
synthesis.

12. It is called template/ nuclear/ messenger 12. It is called insoluble RNA and 12. It is called soluble or adapter RNA
or informational RNA as it carries genetic forms ribosomes. and carries amino acids to mRNA
information provided by DNA. during protein synthesis.

6.4 Transcription in Prokaryotes and Eukaryotes

6.4.1 Prokaryotic Transcription
● Transcription occurs in the cytoplasm with DNA dependent RNA polymerases that catalyze transcription of all three key RNA
types: mRNA, tRNA and rRNA in prokaryotes.
● Steps involved in prokaryotic transcription are:
● Initiation
● Elongation
● Termination
28 MOLECULAR BASIS OF INHERITANCE

Initiation Elongation Termination

RNA polymerase DNA dependent rho factor
binds to promoter RNA polymerase (termination
with the help of synthesizes a factor) stops the
sigma factor (σ) and complementary synthesis of the
initiates mRNA strand as it RNA chain.
transcription. scans the bases
present in the DNA.

Fig 6.35: Prokaryotic transcription

● RNA polymerase binds to the promoter and initiates transcription (Initiation). It uses nucleoside triphosphates as substrate and
polymerises in a template-dependent fashion following the rule of complementarity.
● It somehow also facilitates opening of the helix and continues elongation.
● Only a short stretch of RNA remains bound to the enzyme.
● Once the polymerase reaches the terminator region, the nascent RNA falls off, so also the RNA polymerase. This results in
termination of transcription.
● An intriguing question is how the RNA polymerases are able to catalyze all the three steps, which are initiation, elongation
and termination.
● The RNA polymerase is only capable of catalysing the process of elongation.
● It associates transiently with initiation-factor (σ) and termination-factor (ρ) to initiate and terminate the transcription,
respectively. Association with these factors alter the specificity of the RNA polymerase to either initiate or terminate.
● In bacteria, since the mRNA does not require any processing to become active, and also since transcription and translation take
place in the same compartment (there is no separation of cytosol and nucleus in bacteria), many times the translation can begin
much before the mRNA is fully transcribed.
● Consequently, the transcription and translation can be coupled in bacteria.

6.4.2 Eukaryotic Transcription

● Transcription takes place in the nucleus of a eukaryotic cell.
● Transcription in eukaryotes is slightly more complicated due to two key reasons:
MOLECULAR BASIS OF INHERITANCE 29

● There are at least three RNA polymerases in the nucleus (in addition to the RNA polymerase found in the organelles).
There is a clear-cut division of labour.
The RNA polymerase I transcribes rRNAs (28S, 18S, and 5.8S), whereas the RNA polymerase III is responsible for
transcription of tRNA, 5srRNA, and snRNAs (small nuclear RNAs). The RNA polymerase II transcribes a precursor of
mRNA, the heterogeneous nuclear RNA (hnRNA).

Fig 6.36: Types of RNA polymerases in eukaryotes

● The second complexity is that the primary transcripts contain both the exons and the introns and are non-functional. Hence,
it is subjected to a process called splicing where the introns are removed and exons are joined in a defined order.
30 MOLECULAR BASIS OF INHERITANCE

Fig 6.37: Splicing in eukaryotes

● The hnRNA undergoes additional processing called capping and tailing. In capping an unusual nucleotide (methyl guanosine
triphosphate) is added to the 5'-end of hnRNA.
● In tailing, adenylate residues (200-300) are added at 3'-end in a template independent manner.
● It is the fully processed hnRNA, now called mRNA, that is transported out of the nucleus for translation

Fig 6.38: Eukaryotic transcription

MOLECULAR BASIS OF INHERITANCE 31

7. Genetic Code
● The process of translation requires transfer of genetic information from a polymer of nucleotides to synthesizing a
polymer of amino acids. Neither does any complementarity exist between nucleotides and amino acids, nor could any
be drawn theoretically.
● This led to the proposition of a genetic code that could direct the sequence of amino acids during synthesis of proteins.
● It was George Gamow, a physicist, who argued that since there are only 4 bases and if they have to code for 20 amino acids,
the code should constitute a combination of bases. He suggested that in order to code for all the 20 amino acids, the code
should be made up of three nucleotides. This was a very bold proposition, because a permutation combination of 43 (4 × 4 × 4)
would generate 64 codons; generating many more codons than required.
● Providing proof that the codon was a triplet, was a more daunting task.
● The chemical method developed by Har Gobind Khorana was instrumental in synthesising RNA molecules with defined
combinations of bases (homopolymers and copolymers).
● Marshall Nirenberg’s cell-free system for protein synthesis finally helped the code to be deciphered.
● Severo Ochoa enzyme (polynucleotide phosphorylase) was also helpful in polymerising RNA with defined sequences in a
template independent manner (enzymatic synthesis of RNA).
● Finally a checker-board for genetic code was prepared which is given in Table.

Fig 6.39: The codons for the various amino acids

32 MOLECULAR BASIS OF INHERITANCE

7.1 Features of Genetic Code

● Salient features of the genetic code include:

01 UNIVERSAL The genetic code is the same across species, with rare exceptions.

02 NON-AMBIGUOUS One codon does not specify more than one amino acid

03 DEGENERACY One amino acid can be coded for by more than one codon
04 NON-OVERLAPPING The triplet codes are all discrete and do not overlap
05 POLARITY The genetic code is always read in the 5’-3’ direction

7.2 Mutation and Genetic Code

Fig 6.40: Mutated gene in the genetic material

● The relationships between genes and DNA are best understood by mutation studies.
● Effects of large deletions and rearrangements in a segment of DNA are easy to comprehend. It may result in loss or gain of a
gene and so a function.
● A point mutation or substitution is a genetic mutation where a single nucleotide base is changed, inserted or deleted from a
DNA or RNA sequence of an organism's genome.
● A classic example of point mutation is a change of single base pair in the gene for beta globin chain that results in the change
of amino acid residue glutamate to valine. It results into a diseased condition called as sickle cell anemia.
● Insertion or deletion of one or two bases changes the reading frame from the point of insertion or deletion. However, such
mutations are referred to as frameshift insertion or deletion mutations. Insertion or deletion of three or its multiple bases insert
or delete in one or multiple codons hence one or multiple amino acids, and the reading frame remains unaltered from that
point onwards.
MOLECULAR BASIS OF INHERITANCE 33

Fig 6.41: Point mutation

7.3 tRNA- the Adapter Molecule

● Francis Crick said that there has to be a mechanism to read the code and also to link it to the amino acids, because amino acids
have no structural specialities to read the code uniquely. He postulated the presence of an adapter molecule that would on one
hand read the code and on other hand would bind to specific amino acids.
● The tRNA, then called sRNA (soluble RNA), was known before the genetic code was postulated. However, its role as an
adapter molecule was assigned much later.
● The tRNA molecule has a clover-leaf 2-D structure and a 3-D structure resembling an inverted L.

Fig 6.42: Clover leaf model & Inverted L model of tRNA

Functions of tRNA
● Adaptor molecule - supplies amino acids during translation to ribosomes.
● Recognizes the codons on mRNA through the anticodon loop.
● Recognizes and places specific amino acids at particular points as per the codons in an mRNA.
● Activates specific aminoacyl synthetase enzymes for recognising amino acids.
34 MOLECULAR BASIS OF INHERITANCE

8. Translation
● Translation refers to the process of polymerisation of amino acids to form a polypeptide.
● Translation is the process by which a protein is synthesized from the information contained in a molecule of messenger
RNA (mRNA).
● Translation occurs in a structure called the ribosome, which is a factory for the synthesis of proteins.

8.1 Translation Machinery

The translation machinery of a cell or the raw materials required for protein synthesis include:
● Ribosomes - The cellular factory responsible for synthesising proteins is the ribosome. The ribosome consists of structural
RNAs and about 80 different proteins. In its inactive state, it exists as two subunits; a large subunit and a small subunit. When
the small subunit encounters an mRNA, the process of translation of the mRNA to protein begins. The ribosome also acts as a
catalyst (23S rRNA in bacteria is the enzyme- ribozyme) for the formation of peptide bonds.
● Amino acids - They get added in a chain-like manner according to the codons present on mRNA
● mRNA -A translational unit in mRNA is the sequence of RNA that is flanked by the start codon (AUG) and the stop codon
and codes for a polypeptide. An mRNA also has some additional sequences that are not translated and are referred to as
untranslated regions (UTR). The UTRs are present at both 5' -end (before start codon) and at 3' -end (after stop codon). They
are required for an efficient translation process.
● tRNA - It transfers its amino acid to the growing polypeptide chain
● Aminoacyl tRNA synthetase enzyme -helps in combining an amino acid to its particular tRNA

Fig 6.43: Translation machinery

8.2 Process of Translation

The order and sequence of amino acids are defined by the sequence of bases in the mRNA. The amino acids are joined by a bond
which is known as a peptide bond. Formation of a peptide bond requires energy.
It involves the following steps:
MOLECULAR BASIS OF INHERITANCE 35

● Activation of amino acids

In the first phase itself amino acids are activated in the presence of ATP and aminoacyl tRNA synthetase enzyme to form
amino-acyl-AMP-enzyme-complex. It is an activated amino acid.

● Charging or aminoacylation of tRNA

The amino-acyl-adenylate enzyme complex reacts with a tRNA specific for an amino acid to form an amino-acyl tRNA
complex, where the -COOH end of the amino acid is linked to the 3’-OH end of the tRNA. This tRNA, complexed with an
amino acid is a “charged tRNA”. The process is commonly called as charging of tRNA or aminoacylation of tRNA.

Initiation

Fig 6.44: Initiation of translation

36 MOLECULAR BASIS OF INHERITANCE

● During initiation,the small ribosomal subunit binds to the start of the mRNA sequence.
● Then a transfer RNA (tRNA) molecule carrying the amino acid methionine binds to the start codon of the mRNA sequence.
● The start codon in all mRNA molecules has the sequence AUG and codes for methionine.
● Next, the large ribosomal subunit binds to form the complete initiation complex.

Elongation

Fig 6.45: Elongation of translation

● While Methionine-tRNA occupies the P site, the aminoacyl-tRNA binds to the A site, using energy yielded from the
hydrolysis of GTP.
● Methionine moves from the P site to the A site to bond to a new amino acid there, starting the growth of the peptide.
● The ribosome then translocates along the mRNA molecule to the next codon and the growing peptide lies at the P site and the
A site is open for the binding of the next aminoacyl-tRNA, and the cycle continues.
● Thus the complexes composed of an amino acid linked to tRNA, sequentially bind to the appropriate codon in mRNA by
forming complementary base pairs with the tRNA anticodon. The ribosome moves from codon to codon along the mRNA.
Amino acids are added one by one, translated into Polypeptide sequences dictated by DNA and represented by mRNA.

Termination

Fig 6.46: Termination of translation

MOLECULAR BASIS OF INHERITANCE 37

● When a stop codon (UAA, UAG, UGA) reaches the A-site, no new amino-acyl tRNA is added. The P-site tRNA gets
hydrolysed and the completed polypeptide is released.
● Once a termination codon is encountered, the ribosome moves over the nonsense codon (a codon that doesn’t code for any
amino acid) and slips off the mRNA chain. The two ribosomal subunits separate or undergo dissociation in the presence of a
Dissociation Factor (DF) or Release factor.
● A polyribosome (or polysome or ergosome) is a group of ribosomes bound to an mRNA molecule like “beads” on a “thread”.
It consists of a complex of an mRNA molecule and two or more ribosomes that act to translate mRNA instructions
into polypeptides.
● Polysomes are formed during the elongation phase when ribosomes and elongation factors synthesize the encoded polypeptide.
● Multiple ribosomes move along the coding region of mRNA, creating a polysome.

Fig 6.47: Formation of polyribosome

9. Regulation of Gene Expression

● The mechanism by which a gene is able to express itself in the form of a phenotype in an organism is called gene expression.
● Some genes are expressed all the time but other genes do not need to be expressed all the time.
● Based on this expression pattern, genes can be divided into:

Housekeeping genes Regulated genes

Continuously expressed in all the cells Not always expressed. Activity is
of the body. Also called constitutive regulated. Also called luxury genes
genes or non-constitutive genes.

E.g.: Genes responsible for metabolizing E.g.: Genes responsible for

glucose (glycolysis) metabolizing lactose

9.1 Types of Gene Regulation

● The mechanism by which genes are switched on and off depending on the current requirement of the cells and the state of
development of an organism, is called gene regulation.
● Gene regulation is of two types :
38 MOLECULAR BASIS OF INHERITANCE

Positive gene regulation Negative gene regulation

The process that turns on gene expression is known as
positive gene regulation.
The genes remain non-expressed until and unless they are
induced to do so.
It is also called inducible regulation.
The type of gene regulation which prevents gene
expression is known as negative gene regulation.
The genes continue expressing their effect till their
activity is suppressed.
It is also called repressible regulation.

9.1.1 Gene Regulation in Eukaryotes

In eukaryotes, the regulation could be exerted at
● transcriptional level (formation of primary transcript),
● processing level (regulation of splicing),
● transport of mRNA from nucleus to the cytoplasm,
● translational level.

Fig 6.48: Gene regulation in eukaryotes

MOLECULAR BASIS OF INHERITANCE 39

9.1.2 Gene Regulation in Prokaryotes

● In prokaryotes, gene expression is regulated mostly during transcription initiation. Since binding of RNA polymerase at a gene
promoter is key for initiating transcription, regulatory molecules can affect this polymerase-promoter binding and thus
control transcription.
● Molecules that can affect regulation of gene expression in prokaryotes are:
● Inducers - Activate transcription
● Repressors - Suppress or turn off transcription

Fig 6.49: Operon concept

9.2 Lac Operon
● The lactose operon (lac operon) is an operon required for the transport and metabolism of lactose in E. coli and many other
enteric bacteria. Although glucose is the preferred carbon source for most bacteria, the lac operon allows for the effective
digestion of lactose when glucose is not available through the activity of beta-galactosidase.
● Gene regulation of the lac operon was the first genetic regulatory mechanism to be understood clearly, so it has become a
foremost example of prokaryotic gene regulation.
● This lactose metabolism system was used by François Jacob and Jacques Monod to determine how a biological cell knows
which enzyme to synthesize. Their work on the lac operon won them the Nobel Prize in Physiology in 1965.
● In lac operon, a polycistronic structural gene is regulated by a common promoter and regulatory genes. Such arrangement is
very common in bacteria and is referred to as operon. To name a few such examples, lac operon, trp operon, ara operon, his
operon, val operon, etc.
40 MOLECULAR BASIS OF INHERITANCE

Fig 6.50: Lac Operon

● The lac operon consists of one regulatory gene (the i gene – here the term i does not refer to inducer, rather it is derived from
the word inhibitor) and three structural genes (z, y, and a).
● The i gene codes for the repressor of the lac operon.
● The z gene codes for beta-galactosidase (β-gal), which is primarily responsible for the hydrolysis of the disaccharide, lactose
into its monomeric units, galactose and glucose.
● The y gene codes for permease, which increases permeability of the cell to β-galactosides.
● The a gene encodes a transacetylase. It is an enzyme that transfers an acetyl group from acetyl-CoA to β-galactosides,
glucosides and lactosides.
● Hence, all the three gene products in lac operon are required for metabolism of lactose. In most other operons as well, the
genes present in the operon are needed together to function in the same or related metabolic pathway.
● Lactose is the substrate for the enzyme beta-galactosidase and it regulates switching on and off of the operon. Hence, it is
termed as inducer.
● In the absence of a preferred carbon source such as glucose, if lactose is provided in the growth medium of the bacteria, the
lactose is transported into the cells through the action of permease.
The lactose then induces the operon in the following manner.
● The repressor of the operon is synthesised (all-the-time – constitutively) from the i gene.
● The repressor protein binds to the operator region of the operon and prevents RNA polymerase from transcribing the operon.
● In the presence of an inducer, such as lactose or allolactose, the repressor is inactivated by interaction with the inducer.
● This allows RNA polymerase access to the promoter and transcription proceeds.
● Regulation of lac operon by repressor is referred to as negative regulation.

10. Human Genome Project

Fig 6.51: Human genome project

MOLECULAR BASIS OF INHERITANCE 41

● The Human Genome Project (HGP) launched in the year 1990, was an international scientific research project with the goal of
determining the base pairs that make up human DNA, and of identifying, mapping and sequencing all of the genes of the
human genome from both a physical and a functional standpoint.
● The Human Genome Project was a 13-year project coordinated by the U.S. Department of Energy and the National Institute of
Health. During the early years of the HGP, the Wellcome Trust (U.K.) became a major partner; additional contributions came
from Japan, France, Germany, China and others. The project was completed in 2003.
● It remains the world's largest collaborative biological project. The enormous amount of data expected to be generated also
necessitated the use of high speed computational devices for data storage and retrieval, and analysis. HGP was closely
associated with the rapid development of a new area in biology called Bioinformatics.
● Many non-human model organisms, such as bacteria, yeast, Caenorhabditis elegans (a free living non-pathogenic nematode),
Drosophila (the fruit fly), plants (rice and Arabidopsis), etc., have also been sequenced.

Some important facts about the Human Genome Project are:

Goals of HGP
● Identify all the approximately 20,000-25,000 genes in human DNA
● Determine the sequences of the 3 billion chemical base pairs that make up human DNA
● Store this information in databases
● Improve tools for data analysis
● Transfer related technologies to other sectors, such as industries
● Address the ethical, legal, and social issues (ELSI) that may arise from the project

Methodologies of Human Genome Project (HGP)

In the course of the Human Genome Project, following two approaches were undertaken in order to sequence the entire
human genome.
● Expressed Sequence Tags - In genetics, an expressed sequence tag (EST) is a short sub-sequence of a cDNA sequence. ESTs
may be used to identify gene transcripts, and were instrumental in gene discovery and in gene-sequence determination
42 MOLECULAR BASIS OF INHERITANCE

● Sequence Annotation - Sequencing the entire genome, including both coding and non-coding regions and later assigning
functions to each region. For sequencing, the total DNA from a cell is isolated and converted into random fragments of
relatively smaller sizes (recall DNA is a very long polymer, and there are technical limitations in sequencing very long pieces
of DNA) and cloned in a suitable host using specialised vectors. The cloning resulted in amplification of each piece of DNA
fragment so that it subsequently could be sequenced with ease. The commonly used hosts were bacteria and yeast, and the
vectors were called BAC (bacterial artificial chromosomes), and YAC (yeast artificial chromosomes). The fragments were
sequenced using automated DNA sequencers that worked on the principle of a method developed by Frederick Sanger. These
sequences were then arranged based on some overlapping regions present in them. This required generation of overlapping
fragments for sequencing. Alignment of these sequences was humanly not possible. Therefore, specialised computer based
programs were developed. These sequences were subsequently annotated and were assigned to each chromosome.
● The sequence of chromosome 1 was completed only in May 2006. This was the last of the 24 human chromosomes – 22
autosomes and X and Y – to be sequenced.

10.1 Salient Features of Human Genome

● The human genome contains 3164.7 million bp.
● The average gene consists of 3000 bases, but sizes vary greatly, with the largest known human gene being dystrophin at 2.4
million bases.
● The total number of genes is estimated at 30,000–much lower than previous estimates of 80,000 to 1,40,000 genes. Almost all
(99.9 per cent) nucleotide bases are exactly the same in all people.
● The functions are unknown for over 50 per cent of the discovered genes.
● Less than 2 percent of the genome codes for proteins.
● Repeated sequences make up a very large portion of the human genome.
● Repetitive sequences are stretches of DNA sequences that are repeated many times, sometimes hundred to thousand times.
● Chromosome 1 has the most genes (2968), and the Y has the fewest (231).
● Scientists have identified about 1.4 million locations where single base DNA differences (SNPs – single nucleotide
polymorphism, pronounced as ‘snips’) occur in humans.

10.2 Applications and Future Challenges

MOLECULAR BASIS OF INHERITANCE 43

11. DNA Fingerprinting

Fig 6.52: DNA fingerprinting data on a paper

● DNA profiling (also called DNA fingerprinting) is the process of determining an individual's DNA characteristics.
● DNA profiling is a forensic technique in criminal investigations, comparing criminal suspects profiles to DNA evidence so as
to assess the likelihood of their involvement in the crime It is also used in parentage testing, to establish immigration
eligibility, and in genealogical and medical research. DNA profiling has also been used in the study of animal and plant
populations in the fields of zoology, botany, and agriculture.
● DNA fingerprinting involves identifying differences in some specific regions in a DNA sequence called repetitive DNA,
because in these sequences, a small stretch of DNA is repeated many times. These repetitive DNA are separated from bulk
genomic DNA as different peaks during density gradient centrifugation.
● Repeated sequences (also known as repetitive elements, repeating units or repeats) are patterns of nucleic acids (DNA or
RNA) that occur in multiple copies throughout the genome.
● Repetitive elements found in genomes fall into different classes, depending on their structure and/or the mode of
multiplication. The disposition of repetitive elements consists either in arrays of tandemly repeated sequences, or in repeats
dispersed throughout the genome.
● Tandem repeats occur in DNA when a pattern of one or more nucleotides is repeated and the repetitions are directly adjacent
to each other.
● Tandem repeats can be
Satellite DNA - typically found in centromeres and heterochromatin.
Minisatellite - repeat units from about 10 to 60 base pairs, found in many places in the genome, including the centromeres.
Microsatellite - repeat units of less than 10 base pairs; this includes telomeres, which typically have 6 to 8 base pair
repeat units.
● Debates regarding the potential functions of these elements have been long standing. Controversial references to ‘junk’ or
‘selfish’ DNA were put forward early on, implying that repetitive DNA segments are remainders from past evolution or
autonomous self-replicating sequences hacking the cell machinery to proliferate.
● These sequences normally do not code for any proteins, but they form a large portion of the human genome. These sequences
show a high degree of polymorphism and form the basis of DNA fingerprinting.

Fig 6.53: The same GATA sequence has been repeated to different degrees in different individuals
44 MOLECULAR BASIS OF INHERITANCE

● Since DNA from every tissue (such as blood, hair-follicle, skin, bone, saliva, sperm etc.), from an individual shows the same
degree of polymorphism, they become a very useful identification tool in forensic applications.
● Further, as the polymorphisms are inheritable from parents to children, DNA fingerprinting is the basis of paternity testing, in
case of disputes.
● As polymorphism in DNA sequence is the basis of genetic mapping of human genome as well as of DNA fingerprinting, it is
essential that we understand what DNA polymorphism means in simple terms. Polymorphism (variation at genetic level) arises
due to mutations.
● New mutations may arise in an individual either in somatic cells or in the germ cells (cells that generate gametes in sexually
reproducing organisms). If a germ cell mutation does not seriously impair individual’s ability to have offspring who can
transmit the mutation, it can spread to the other members of population (through sexual reproduction).
● If an inheritable mutation is observed in a population at high frequency, it is referred to as DNA polymorphism. The
probability of such variation to be observed in noncoding DNA sequences would be higher as mutations in these sequences
may not have any immediate effect/impact in an individual’s reproductive ability.
● These mutations keep on accumulating generation after generation, and form one of the basis of variability/polymorphism.
There are a variety of different types of polymorphisms ranging from single nucleotide change to very large scale changes.

DNA Fingerprinting Technique

● The technique of DNA Fingerprinting was initially developed by Alec Jeffreys. He used a satellite DNA as a probe that
showed a very high degree of polymorphism. It was called the Variable Number of Tandem Repeats (VNTR). The technique,
as used earlier, involved Southern blot hybridisation using radiolabelled VNTR as a probe.
● It included the following steps
● isolation of DNA,
● digestion of DNA by restriction endonucleases,
● separation of DNA fragments by electrophoresis,
● transferring (blotting) of separated DNA fragments to synthetic membranes, such as nitrocellulose or nylon,
● hybridisation using labelled VNTR probe, and
● detection of hybridised DNA fragments by autoradiography.

Fig 6.54: Process of DNA Fingerprinting

MOLECULAR BASIS OF INHERITANCE 45

● The VNTR belongs to a class of satellite DNA referred to as mini-satellite. A small DNA sequence is arranged tandemly in
many copy numbers. The copy number varies from chromosome to chromosome in an individual. The numbers of repeats
show a very high degree of polymorphism.
● As a result the size of VNTR varies in size from 0.1 to 20 kb. Consequently, after hybridisation with VNTR probe, the
autoradiogram gives many bands of differing sizes. These bands give a characteristic pattern for an individual DNA.
● It differs from individual to individual in a population except in the case of monozygotic (identical) twins.

DNA Fingerprinting: Applications

● It is used in forensic science to identify potential crime suspects.
● It is used to establish paternity and family relationships.
● It is used to identify and protect the commercial varieties of crops and livestock.
● It is used to find the evolutionary history of an organism and trace out the linkage between groups of various organisms.
● In case of the change in gene frequency or genetic drift, DNA fingerprinting can be used to trace the role of this change
in evolution.

Fig 6.55: Applications of DNA Fingerprinting in paternity testing and crime scene forensics

● The two alleles (paternal and maternal) of a chromosome also contain different copy numbers of VNTR. The banding pattern
of DNA from crime scenes matches with individual B, and not with A.
46 MOLECULAR BASIS OF INHERITANCE

EXERCISE (Basic Exercise) 5. Which of the following important

biochemical reactions in living systems is
1. In eukaryotes which of the following is
catalyzed by a ribozyme?
removed from initially transcribed RNA
(a) Repair of DNA
before it is transported to the cytoplasm for
(b) Electron transport chain
translation ?
(c) Formation of peptide bond
(a) Intron
(d) Organization of MTOC during cell
(b) 3' Poly A tail
division
(c) Ribosome binding site
6. VNTR are useful in DNA profiling because
(d) 5' cap
they
2. In prokaryotes, the predominant site for
I. Are hypervariable
control of gene expression is the :
II. Are inherited
(a) transcription initiation
III. Synthesize constitutive enzymes
(b) processing level
(a) I, II, III are correct
(c) transport of mRNA
(b) I and II are correct
(d) translation level
(c) I and III are correct
3. The process of splicing in eukaryotes :
(d) II and III are correct
(a) is reminiscent of antiquity
7. Which enzyme polymerizes RNA with
(b) represents dominance of RNA world
defined sequences in a template
(c) is an indicator of the complexity of
independent manner?
human genome
(a) Peptidyl transferase
(d) is a legacy of organic evolution
(b) RNA polymerase
4. In Hershey and Chase experiment, what
(c) Reverse transcriptase
finding led to the conclusion that DNA is
(d) Polynucleotide phosphorylase
the genetic material ?
8. The process of RNA splicing shows the
(a) The presence of radioactivity in the
dominance of:
bacteria when 35S was used.
(a) DNA world
(b) The presence of radioactivity in the
(b) RNA world
supernatant when 355 was used.
(c) Protein world
(c) The presence of radioactivity in the
(d) Microbial world
bacteria when 32P was used.
9. In RNA splicing :
(d) The presence of radioactivity in the
supernatant when 32P was used.
MOLECULAR BASIS OF INHERITANCE 47

(a) Exons are removed and introns are 14. A molecule that can act as a genetic
joined together material must fulfill all the following
(b) Introns are removed and exons are criteria except:
joined together (a) It should be able to generate its replica
(c) Cistrons are removed and introns are (b) It should chemically and structurally be
joined together stable
(d) Introns are removed and Cistrons are (c) It should provide scope for rapid
joined together mutations
10. The core RNA polymerase is capable of (d) It should be able to express itself in the
catalyzing which steps of transcription ? form of "Mendelian Characters"
(a) Initiation only 15. The unequivocal proof that DNA is the
(b) Elongation only genetic material was provided by:
(c) Termination only (a) Avery, Macleod and McCarty
(d) All of these (b) Hershey and Chase
11. In most prokaryotes the transcription unit is (c) Meselson and Stahl
: (d) Watson and Crick
(a) Mono-cistronic 16. Which of the following is not a feature of
(b) Poly-cistronic the double helix model of DNA ?
(c) Multi-cistronic (a) The two chains have anti-parallel
(d) Uni-cistronic polarity
12. The DNA dependent DNA polymerases (b) A purine always comes opposite to a
catalyze: pyrimidine
(a) Only in 3-5 direction (c) The pitch of the DNA is 3.4 nm
(b) Only in 5-3' direction (d) The two chains are coiled in a left
(c) In both directions handed fashion
(d) In neither directions 17. According to Erwin Chargaff, for a double
13. Reverse transcriptase is a: stranded DNA
(a) DNA dependent RNA polymerase (a) The ratios between Adenine and
(b) RNA dependent DNA polymerase Thymine, and, Guanine and Cytosine are
(c) RNA dependent RNA polymerase constant and equals one.
(d) DNA dependent DNA polymerase (b) The ratios between Adenine and
Thymine, and, Guanine and Cytosine are
constant but is not equal to one.
48 MOLECULAR BASIS OF INHERITANCE

(c) The ratios between Adenine and (a) AUG

Guanine, and, Thymine, and Cytosine are (b) GUG
constant and equals one. (c) CUG
(d) The ratios between Adenine and (d) UUG
Guanine, and, Thymine and, Cytosine are 22. What is the major difference between the
constant but is not equal to one. lagging and leading strands during DNA
18. To form a nucleoside, a nitrogenous base is replication ?
linked to a pentose sugar: (a) On the leading strand, DNA synthesis
(a) Through a P-Glycosidic linkage at occurs from 5 to 3', while DNA synthesis
carbon atom number 1 occurs from 3 to 5' on the lagging strand.
(b) Through a P-Glycosidic linkage at (b) DNA polymerase is able to continuously
carbon atom number 5 add new nucleotides on the leading strand
(c) Through a N-Glycosidic linkage at while it must keep 'starting over' on the
carbon atom number 1 lagging strand.
(d) Through a N-Glycosidic linkage at (c) The lagging strand requires only a single
carbon atom number 5 primer while the leading strand requires
19. Identify the incorrectly matched pair: many.
(a) Organism - Bacteriophage 0 174; (d) Helicase opens the leading stran at a
Length of DNA - 5386 nucleotides faster rate than the lagging strand.
(b) Organism - Bacteriophage Lambda; 23. The inducer of lac operon is:
Length of DNA - 48502 base pairs (a) Beta galactosidase
(c) Organism - Escherichia coli; Length of (b) Galactose
DNA – 4×108 base pairs (c) Lactose
(d) Organism - Human beings; Length of (d) Glucose
DNA - 3.3 × 109 base pairs 24. regulatory domains of most activators
20. The amino acid acceptor arm of the tRNA interact with
is at its: (a) the transcription factor complex
(a) DHU loop (b) RNA polymerase
(b) TUC loop (c) repressors
(c) 5 end (d) the DNA binding domain
(d) 3' end 25. DNA chemically is less reactive and
21. Which of the following codons has a dual structurally more stable than RNA.
role in the genetic code? Therefore,
MOLECULAR BASIS OF INHERITANCE
(a) DNA has evolved from the RNA
(b) RNA can directly code for proteins
(c) DNA is the better genetic material than
RNA
(d) The protein synthesizing machinery has
evolved around RNA
26. A transcription unit does not contain:
(a) A promoter
(b) The structural gene
(c) A terminator
(d) An operator
27. Regulation of lac operon by repressor is
referred to as:
(a) Inducible regulation
(b) Repressible regulation
(c) Negative regulation
(d) Positive regulation
28. A synthetic mRNA of repeating sequence
5'-CACACACACACACACAC... is used
for a cell-free protein synthesizing system
like the one used used by Nirenberg. If we
assume that protein synthesis can begin
without the need for an initiator codon,
what product or products would you expect
to occur after protein synthesis?
(a) one protein, consisting of a single amino
acid
(b) three proteins, each consisting of a
different, single amino acid
(c) two proteins, each with an alternating
sequence of two different amino acids
(d) one protein, with an alternating
sequence of two different amino acids
49
29. Fredrick Griffith accidentally discovered
transformation when attempting to develop
a vaccine for pneumonia. He injected mice
with samples from S-strain (virulent) and/or
R-strain (nonvirulent) pneumococci
bacteria (Sterptococcus pneumoniae).
Which of the following results is NOT
consistent with Griffith's experiments?
(a) injected S-strain; mouse dies.
(b) injected R-strain; mouse lives.
(c) injected heat-killed S-strain; mouse
lives.
(d) injected mixture of heat-killed S-strain
and live R-strain; mouse lives.
30. Which scientists first gave experimental
evidence that DNA is the genetic material?
(a) Avery, MacLeod, and McCarty who
repeated the transformation experiments of
Griffith, and chemically characterized the
transforming principle.
(b) Garrod, who postulated that
Alkaptonuria, or black urine disease, was
due to a defective enzyme.
(c) Beadle and Tatum, who used a
mutational and biochemical analysis of the
bread mold Neurospora to establish a direct
link between genes and enzymes.
(d) Meselson and Stahl who showed that
DNA is replicated semi conservatively.
31. The function of the rho protein is
(a) to help terminate translation
(b) to help RNA polymerase bind to the
DNA
50 MOLECULAR BASIS OF INHERITANCE

(c) to help RNA polymerase find a (c) three

promoter (d) Ten
(d) to help terminate transcription 37. Consider the following statements:
32. Genetic information in a DNA molecule is I. The largest known human gene is
coded in the: dystrophin at 2.4 million bases
(a) Sequence of nucleotides II. Repeated sequences make up very large
(b) Base pairings portion of the human genome
(c) Proportion of each base present III. There are about 2.4 million locations
(d) The turning pattern of the helix where single base DNA differences
33. In DNA replication, the Okazaki fragments occur in humans
on the lagging strand are joined together by: Which of the above statements are true?
(a) DNA ligase (a) I and II only
(b) DNA polymerase (b) I and III only
(c) Primase (c) II and III only
(d) Helicase (d) I, II and III
34. The enzyme that catalyzes the peptide 38. Consider the following statements:
bonding in prokaryotes is located in the I. RNA is the genetic material of the QB
(a) Leader region of the mRNA bacteriophage
(b) Central part of tRNA II. DNA chemically is less reactive and
(c) Smaller subunit of the ribosome more stable than RNA
(d) Larger subunit of the ribosome III. Viruses having RNA genome and
35. In general, bacterial genes are regulated at having shorter life span mutate &
the time of: evolve slowly.
(a) Transcription Which of the above statements are true?
(b) Post transcription (a) I and II only
(c) Translation (b) I and III only
(d) Post translation (c) II and III only
36. The insertion or deletion of a base pair into (d) I, II and III
the genetic code will cause a frame shift 39. Consider the following statements:
mutation unless the number of base pairs I. Inheritance of a character is affected by
inserted or deleted is: promoter and regulatory sequences of a
(a) One structural gene.
(b) Two
MOLECULAR BASIS OF INHERITANCE 51

II. The split-gene arrangement represents an Which of the above statements are true?
ancient feature of the genome. (a) I and II only
III. The process of splicing represents the (b) I and III only
dominance of RNA world. (c) II and III only
which of the above statements are true? (d) I, II and III
(a) I and II only 42. Match each item in Column I with one item
(b) I and III only in Column Il and chose your answer from
(c) II and III only the codes given below:
(d) I, II and III Column I Column II
40. Consider the following statements: (NRNA codons) (Amino Acid)
I. All the reference point while defining a I. GUG 1. Tryptophan
transcription unit is made with coding II. UGG 2. Valine
strand III. UAC 3. Glutamic acid
II. It is the presence of a promoter in a IV. GAG 4. Tyrosine
transcription unit that also defines the
template and coding strands (a) (I) - 1; (II) - 2; (III) - 3; (IV) - 4
III. Exons are said to be sequences that do (b) (I) - 2; (II) - 1; (III) - 4; (IV) - 3
not appear in mature or processed RNA (c) (I) - 2; (II) - 1; (III) - 3; (IV) - 4
Which of the above statements are true? (d) (I) - 1; (II) - 2; (III) - 4; (IV) - 3
(a) I and II only 43. If there are 999 bases in an RNA that codes
(b) I and III only for a protein with 333 amino acids and the
(c) II and III only base at position 901 is deleted such that the
(d) I, II and III length of teh RNA becomes 998 bases, how
41. Consider the following statements: many codons will be altered?
I. Deoxyribonucleoside triphosphates act as (a) 1
substrates as well as provide energy for (b) 11
polymerization reaction during DNA (c) 33
replication. (d) 333
II. The replication of DNA is both 44. DNA fragments are
semiconservative and semi- discontinuous (a) positively charged
III. In bacterial DNA replication there are (b) negatively charged
multiple ori and replication fork moves bi- (c) neutral
directionally
52 MOLECULAR BASIS OF INHERITANCE

(d) either positively or negatively charged (b) tRNA

depending on their size (c) mRNA
45. During DNA replication, Okazaki (d) miRNA
fragments are used to elongate 50. Taylor conducted the experiments to prove
(a) The leading strand towards replication semi-conservative mode of chromosome
fork replication on
(b) The lagging strand towards replication (a) Vinca rosea
fork (b) Vicia faba
(c) The leading strand away from (c) Drosophila melanogaster
replication fork (d) E. coli
(d) The lagging strand away from 51. The equivalent of a structural gene is
replication fork (a) muton
46. Spliceosomes are not found in cells of (b) cistron
(a) plants (c) operon
(b) fungi (d) recon
(c) animals 52. Which of the following rRNAs act as
(d) bacteria structural RNA as well as ribozyme in
47. The final proof for DNA as the genetic bacteria?
material come from the experiments of (a) 5 s rRNA
(a) Griffith (b) 18 s rRNA
(b) Hershey and chase (c) 23 s rRNA
(c) Avery, Macleod and McCarty (d) 58 s rRNA
(d) Hargobind Khorana 53. DNA-dependent RNA polymerase
48. The association of histone H1 with a catalyses transcription on one strand of the
nucleosome indicates DNA which is called the
(a) transcription is occuring (a) template strand
(b) DNA replication is occuring (b) coding strand
(c) the DNA is condensed into chromatin (c) alpha strand
fibre (d) anti strand
(d) the DNA double helix is exposed 54. Which of the following is not required for
49. Which of the following RNAs should be any of the techniques of DNA
most abundant in animals cell? fingerprinting available at present?
(a) rRNA (a) Zinc finger analysis
MOLECULAR BASIS OF INHERITANCE 53

(b) Restriction enzymes cytosine. The percentages of the other three

(c) DNA-DNA hybridisation bases expected to be present in this DNA
(d) Polymerase chain reaction are
55. Which one of the following is the start (a) G/34%, A/24.5%, T/24.5%
codon? (b) G/17%, A/16.5%, T/32.5%
(a) UGA (c) G/17%, A/33%, T/33%
(b) UAA (d) G/8.5%, A/50%, T/24.5%
(c) UAG 60. Transformation was discovered by
(d) AUG (a) Meselson and Stahl
56. Which one of the following is not (b) Hershey and Chase
applicable to RNA ? (c) Griffith
(a) Complementary base pairing (d) Watson and Crick
(b) 5' phosphoryl and 3" hydroxyl ends 61. Select the correct option
(c) Heterocyclic nitrogenous bases (a) Direction of RNA synthesis – 5’ – 3’
(d) Chargaff's rule Direction of reading of the template DNA
57. Identify the correct order of organisation of strand - 3’ – 5’
genetic material from largest to smallest. (b) Direction of RNA synthesis - 3’ – 5’
(a) Chromosome, gene, genome, nucleotide Direction of reading of the template DNA
(b) Genome, chromosome, nucleotide, gene strand - 5’ – 3’
(c) Genome, chromosome, gene, nucleotide (c) Direction of RNA synthesis - 5’ – 3’
(d) Chromosome, genome, nucleotide, gene Direction of reading of the template DNA
58. Satellite DNA is important because it strand - 5’ – 3’
(a) codes for protiens needed in cell cycle (d) Direction of RNA synthesis - 3’ – 3’
(b) shows high degree of polymorphism in Direction of reading of the template DNA
population and also the same degree of strand - 3’ – 3’
polymorphism in an individual, which is 62. Commonly used vectors for human genome
heritable from parents to children sequencing are
(c) does not code for protiens and is same in (a) T-DNA
all members of the population (b) BAC and YAC
(d) codes for enzymes needed for DNA (c) Expression vectors
replication. (d) T/A cloning vectors
59. In sea urchin DNA, which is double
stranded 17% of the bases were shown to be
54 MOLECULAR BASIS OF INHERITANCE

63. Which enzymes/s will be produced in a cell 68. Removal of introns and joining of exons in
in which there is a non-sense mutation in a defined order during transcription is
the lac Y gene? called
(a) B-galactosidase (a) looping
(b) Lactose permease (b) inducing
(c) Transacetylase (c) capping
(d) Lactose permease and transacetylase (d) splicing
64. Removal of RNA polymerase III from 69. What are those structures that appear as
nucleoplasm will affect the synthesis of 'beads-on-string' in the chromosomes when
(a) TRNA viewed under electron microscope?
(b) hnRNA (a) Nucleotides
(c) mRNA (b) Nucleosomes
(d) rRNA (c) Base pairs
65. PCR and Restriction Fragment Length (d) Genes
Polymorphism are the methods for 70. Select the two statements out of the four (A
(a) study of enzymes -D) given below about lac operon.
(b) genetic transformation (A) Glucose or galactose may bind with the
(c) DNA sequencing repressor and inactivate it
(d) genetic fingerprinting (B) In the absence of lactose, the repressor
66. Which one of the following is not a part of binds with the operator region
a transcription unit in DNA? (C) The z-gene codes for permease
(a) The inducer (D) This was elucidated by Francois Jacob
(b) A terminator and Jacques Monod
(c) A promoter (a) (B) and (C)
(d) The structural gene (b) (A) and (A)
67. If one strand of DNA has the nitrogenous (c) (B) and (D)
base sequence as ATCTG, what would be (d) (A) and (B)
the complementary RNA strand sequence?
(a) TTAGU
(b) UAGAC
(c) AACTG
(d) ATCGU
MOLECULAR BASIS OF INHERITANCE 55

Answers Key
EXERCISE (Basic Exercise)
1. (a) 2. (b) 3. (a) 4. (c) 5. (c)

6. (b) 7. (d) 8. (b) 9. (b) 10. (b)

11. (b) 12. (b) 13. (b) 14. (c) 15. (b)

16. (d) 17. (a) 18. (d) 19. (d) 20. (c)

21. (a) 22. (b) 23. (c) 24. (a) 25. (c)

26. (d) 27. (c) 28. (d) 29. (d) 30. (a)

31. (d) 32. (a) 33. (a) 34. (d) 35. (a)

36. (c) 37. (a) 38. (a) 39. (d) 40. (a)

41. (a) 42. (b) 43. (c) 44. (b) 45. (d)

46. (d) 47. (b) 48. (c) 49. (a) 50. (b)

51. (b) 52. (c) 53. (a) 54. (a) 55. (d)

56. (d) 57. (c) 58. (b) 59. (c) 60. (c)

61. (a) 62. (b) 63. (a) 64. (a) 65. (d)

66. (a) 67. (b) 68. (d) 69. (b) 70. (c)