1. Eukaryotic genomes contain nuclear DNA as well as organelle DNA from mitochondria and chloroplasts. Genome size, or C-value, varies greatly between species from 106 bp in prokaryotes to over 1011 bp in some amphibians. 2. Renaturation kinetics can be used to measure genome complexity based on how quickly denatured DNA strands reanneal, with more common sequences reassociating faster. A COT curve plots the percentage of renatured DNA over time at different DNA concentrations. 3. Eukaryotic genomes contain genes, repetitive sequences like satellites and transposons, and non-coding DNA. While genes and complexity generally increase together in lower e

1. MOLECULAR GENETICS

2. GENOME ORGANIZATION IN EUKARYOTES
• Genome – totality of all of the DNA of an organism
• Eukaryotic genome – nuclear genome and Organelles genome (mitochondrial & chloroplast).
C value – Amount of DNA present in haploid genome of species. It is characteristic of each
species.
• The value ranges from <106 bp (prokaryote, mycoplasma) to >1011 bp for amphibians.
• Genome of the higher organisms contains large excess of DNA.
C – value paradox :
• DNA content of the haploid genome is related to morphological complexity of lower
eukaryotes but varies extensively in higher eukaryotes.
• Yeast, amount of DNA increases with increasing complexity of organisms. In higher
eukaryotes there is no correlation between increased genome size & complexity.
• This lack of correlation between genome size and genetic complexity – C-value paradox.
• Ex: man is more complex than amphibians (genetic development) but amphibian cells
contain 30X more DNA than human cells.

3. GENOME SIZE IN EUKARYOTES
Organism Genome size
(Mb)
S.cerevisiae (yeast) 12
A.thaliana (mustard
plant)
120
D.melanogaster
(fruit fly)
170
H.sapiens (humans) 3,300
H.vulgare (barley) 5,300

4. RENATURATION KINETICS
• Genome complexity – total length of different sequences of
DNA. It can be measured through the renaturation kinetics
of denatured DNA.
• Renaturation occurs through complementary base pairing.
• The rate at which heat-denatured DNA sequences in
solution will renature is dependent on DNA concentration,
re association temperature, cation concentration, and
viscosity.

5. COT CURVE
• It is concerned with the measurement of the degree of reannealing of DNA
strands.
• It is a curve drawn with X-axis having DNA concentration unit multiplied
by time. Since the initial concentration is considered represented as Co and
when multiplied with time t, it becomes "Cot" and the graph is known as
Cot curve.
• The graph is drawn against % reanealled versus Cot.
• Cot = DNA conc. (mol/L) x renaturation time in sec x a buffer factor that
accounts for the effect of cations on the speed of renaturation.
For example:
• Nucleotide concentration = 0.050 M
• Renaturation time = 344.000 sec
• Buffer factor, 0.5 M SPB = 5.820
Cot =100.000

6. • The rate at which a particular sequence will reassociate is
proportional to the number of times it is found in the genome.
• Given enough time, nearly all of the DNA in a heat-denatured
DNA sample will reassociate.
Steps Involved in DNA Denaturation and Renaturation
Experiments :
1. Shear the DNA to a size of about 400 bp.
2. Denature the DNA by heating to 100oC.
3. Slowly cool and take samples at different time intervals.
4. Determine the % single-stranded DNA at each time point.
• [(Vss x Ass) x 100] ÷ [(Vss x Ass) + (Vds x Ads)] = % ssDNA
where Vss = total volume of single-strand fraction, Vds = total
volume of double-strand fraction, Ass = A260 for single
stranded fraction, Ads = A260 for double stranded fraction

7. THE SHAPE OF A "COT" CURVE FOR A GIVEN SPECIES IS A
FUNCTION OF TWO FACTORS:
- SIZE OR COMPLEXITY OF THE GENOME;
- AMOUNT OF REPETITIVE DNA WITHIN THE GENOME

8. NUCLEAR GENOME

9. FUNCTIONAL DNA SEQUENCE
 Gene – each region of a DNA helix that produces a functional RNA molecule
constitutes a gene.
Gene families :
• Protein coding genes – 1X in haploid genome – solitary genes
• Duplicated genes – constitute the gene families.
A group of genes of identical or similar sequence that code for identical or
related proteins. They remained clustered or dispersed around the genome.
mRNA
rRNA, tRNA
Introns
(coding
sequence)
Exons (non
coding
sequences)Protein
GENE

10. • Simple multigene families :
All the members have identical or nearly identical sequences. Ex:
rRNA genes.
• Complex multigene families:
Have similar sequences but differ in their gene products. Ex:
mammalian globin genes.
Pseudogenes :
Close homology to the functional genes but have been disabled by
mutations that prevent their expression.
• Conventional pseudogene :
Genes become inactive due to the accumulation of mutations
• Processed pseudogene:
A gene that results from integration into the genome of a reverse
transcribed copy of an mRNA. It contains no introns and no
promoter sequences which makes it transcriptionally inactive.

11. REPETITIVE DNA SEQUENCE
• Single copy DNA sequences – found unique (i.e., there is only
1 copy in a haploid genome)
• Multicopy DNA sequences – Repetitive DNA sequences
(present in more than 1 copy)
Repetitive DNA sequences – Moderatively repetitive and
Highly repetitive.
• Highly Rep : consists of tandem repeats (short seq <100bp
repeated many times in tandem in large clusters)
• Moderatively Rep : consists of relatively short sequences that
are repeated 10-1000X in the genome

12. HIGHLY REPETITIVE DNA SEQUENCES
Satellite DNA :
• DNA fragments containing tandemly repeated sequences form 'satellite' bands
when genomic DNA is fractionated by density gradient centrifugation.
• <5 to 200bp
• Species specific
• They are not transcribed and are located most often in the heterochromatin
associated with the centromeric region of chromosomes.
• Ex : Human alphoid DNA

13. Minisatellites :
• Form clusters up to 20kb in length with 20-59 repeat units, each
containing about 15 to 100 bp.
• Ex : Telomeric DNA (in humans comprises of 100’s of motifs of 5`-
TTAGGG-3’
Microsatellites :
• Shorter, usually <150bp, and the repeat unit is usually 13bp or less. The
typical microsatellite consists of 1-, 2-, 3- or 4-bp unit repeated 10 to 20
times.
• Ex: in humans microsatellite with “CA” repeat that make upto 0.25%
of the genome.

14. MODERATIVELY REPEATED DNA SEQ
• They are interspersed at multiple sites throughout the
genomes.
• Many varieties of transposable elements are present
• Transposition -The process by which these sequences are
copied and inserted into a new site (by means of encoding
enzyme) in the genome.

15. Moderatively rep. sequence
Transposons Retrotransposons
Conservative Replicative
Viral Non viral
LTR LINES SINES

16. TRANSPOSONS
• Transpose to new sites directly as DNA.
• Ex : P element in Drosophila, Ac-Ds elements in Maize.
Conservative Transposition :
• Excision of the sequence from its original position and reinsertion
elsewhere.
• Change in the position of transposon without increase in copy number.
Replicative Transposition:
• Copy of the original transposon is inserted at the new position.
• Results – increase in copy number.

18. RETROTRANSPOSONS
• Transposable elements are first transcribed into a RNA copy, which is
then reverse transcribed into DNA and get inserted.
Viral retrotransposons : flanked by 250-600bp long terminal repeats
(LTRs) and encode reverse transcriptase and integrase.
• Ex: Ty elements in Yeast and copia elements in Drosophila
Non viral Retrostransposons :
• Lack LTRs and have A/T rich stretch at one end. The most abundant non
viral retrostransposons are,
SINES : 100 – 400 bp seq which does not contain any gene and are
transcribed by RNA polymerase III.
LINES : usually flanked by short direct repeats and contains 2 ORFs,
encodes an RNA-binding protein and the reverse transcriptases.

19. PACKAGING OF DNA INTO CHROMOSOMES

20. TRANSCRIPTIONAL REGULATION
Acetylation – addition of acetyl groups (by histone acetyl transferases) to the
lysine amino acids in the histone tails of each of histone core molecules..
• Essential for the formation of chromatin by reducing the affinity of the histones
for DNA – function as txnl activators.
• Histone deacetylases – removes the acetyl groups results in txnl repression.
Nucleosome remodeling : an energy dependent process that weakens the contact
between the nucleosome and the DNA associated. It involves a change in the
structure of the nucleosome.
Methylation : DNA methyl transferases catalyzes the methylation of cytosine
residues at CG dinucleotide .
RNA silencing – siRNA, miRNA

21. MECHANISM OF GENE SILENCING
Synthetic dsRNA RNA viruses Inverted repeats Transgene
siRNA

22. • miRNA – non coding RNA
• RNA III polymerase – drosha (dsRNA specific endonuclease + pasha
(dsRNA binding protein)
Pre miRNA
Pre miRNA
Cytoplasm – target for dicer…..

23. TRANSLATIONAL REGULATION

24. HEME CONTROL OF TRANSLATION

25. IRON CONTROL OF TRANSLATION

26. Extracellular stimuli ( e.g. insulin, EGF, angiotensin II and
gastrin)
Activation of RAS gene (by binding to its receptor)
phosphorylation and activation of MAP-interacting kinase-1 (Mnk1)
phosphorylation of eIF4E
Increased translation
4EBPs (binding or repressor protein)

27. BACTERIOPHAGE CYCLE SWITCH
CI CIICroNCIII
PRPRMPL
OR3 OR2 OR1OL1 OL2 OL3
CIN CroCIII CII
Cro, N – Lytic pathway
CI – lysogenic pathway
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