DNA Compaction

Chromatin and Nucleoid Structure

The term chromosome refers to the nucleic acid molecule that is the repository of the genetic
information of a virus, a bacterium, a eukaryotic cell, or an organelle. Eukaryotic chromosomes,
in the original sense of the word, appear as sharply defined bodies in the nucleus during the
period just before and during mitosis, the process of nuclear division in somatic cells. In non-
dividing eukaryotic cells, the chromosomal material, called chromatin, is amorphous and
appears to be randomly dispersed throughout the nucleus. But when the cells prepare to divide,
the chromatin condenses and assembles itself into a species-specific number of well-defined
chromosomes.

Chromatin has been isolated and analyzed. It consists of fibers that contain protein and DNA in
approximately equal masses, plus a small amount of RNA. The DNA in the chromatin is very
tightly associated with proteins called histones, which package and order the DNA into
structural units called nucleosomes. Also found in chromatin are many nonhistone proteins,
some of which regulate the expression of specific genes. Beginning with nucleosomes,
eukaryotic chromosomal DNA is packaged into a succession of higher order structures that
ultimately yield the compact chromosome seen with the light microscope. We now turn to a
description of this structure in eukaryotes, and compare the DNA packaging in bacterial cells.

Histones

Histones Are Small, Basic Proteins. Found in the chromatin of all eukaryotic cells, histones have
molecular weights of between 11,000 and 21,000 and are very rich in the basic amino acids
arginine and lysine. Five major classes of histones are found in all eukaryotic cells, differing in
molecular weight and amino acid composition. The H3 histones are nearly identical in amino
acid sequence in all eukaryotes, as are the H4 histones, suggesting strict conservation of their
functions. Histones Hl, H2A, and H2B show a lesser degree of sequence homology between
eukaryotic species. Each of the histones can exist in different forms because certain amino acid
side chains are enzymatically modified by methylation, ADP-ribosylation, phosphorylation, or
acetylation. Such modifications change the histone molecules' net electric charge, shape, and
other properties, but the functional significance of the changes is not well understood.

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Nucleosomes Are Packed into Successively Higher-Order Structures

Wrapping DNA about a nucleosome core compacts it about sevenfold. The total compaction in a
chromosome is greater than 10,000-fold, which in itself provides ample evidence for even higher
orders of structural organization. In chromosomes isolated by very gentle methods, nucleosomes
themselves appear to be organized to form a structure called simply a 30 nm fiber. Packing
requires one molecule of histone Hl per nucleosome, although it is unclear where the Hl is
bound. Organization into 30 nm fibers does not extend over the entire chromosome, but is
punctuated by regions that are bound by sequence-specific (non-histone) DNA-binding proteins.
The structure observed also appears to depend on the transcriptional activity of the particular
region of DNA. Regions containing genes that are being transcribed are apparently in a less-
ordered state that contains little, if any, histone Hl.

A model for layers of organization in a eukaryotic chromosome. The layers take the form of coils
upon coils.

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 Bacterial DNA Is Also Highly Organized

Bacterial DNA is compacted in a structure called the nucleoid, which occupies a large fraction of
the bacterial cell's volume. The DNA of bacterial cells appears to be attached at one or more
points to the inner surface of the plasma membrane. Much less is known about the structure of
the nucleoid than of eukaryotic chromatin. In E. coli, a scaffold-like structure appears to exist
that organizes the circular chromosome into a series of looped domains, as described above for
chromatin. The local organization provided by nucleosomes in eukaryotes does not seem to be
duplicated by any comparable structure in bacterial DNA. Histone like proteins are abundant in
E. coli, and the best characterized example is a protein with two subunits called HU (Mr19,000).
However, these proteins bind and dissociate on a time scale of minutes, and no regular, stable
structure has been found. The bacterial chromosome is a relatively dynamic structure, possibly
reflecting a requirement for more ready access to the genetic information it contains. The
bacterial cell division cycle can be as short as 15 min, whereas a typical eukaryotic cell may not
divide for many months. In addition, structural genes account for a much greater fraction of
prokaryotic DNA, and high rates of cellular metabolism in bacteria mean that a much higher
proportion of the DNA is being transcribed or replicated at a given time than in most eukaryotic
cells.

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 Karyotype Analysis and Chromosome Banding

Chromosomes in metaphase can be identified using certain staining techniques, so called

banding. Cells are cultured and then stopped in metaphase to maximize the number of suitable
cells. They are then spread on a slide, stained with a suitable dye and visualized in the
microscope. Most conventional cytogenetic analyses depend on the karyotyping of banded
metaphase chromosomes.

A band is defined as that part of a chromosome which is clearly distinguishable from its adjacent
segments by appearing darker or brighter with one or more banding techniques. The
chromosomes are visualized as consisting of a continuous series of bright and dark bands.

The banding techniques fall into two principal groups:

1) those resulting in bands distributed along the length of the whole chromosome, such as G-, Q-
and R-bands and

2) those that stain a restricted number of specific bands or structures. These latter include
methods which reveal centromeric bands, C-bands, and nucleolus organizer regions.

C-banding methods do not permit identification of every chromosome in the somatic cell
complement, but can be used to identify specific chromosomes. G- and R- bands can be bright
field or fluorescent.

Bright field G-bands

These G-bands are most commonly used. They take their name from the Giemsa dye, but can be
produced with other dyes. In G-bands, the dark regions tend to be heterochromatic, late-
replicating and AT rich. The bright regions tend to be euchromatic, early-replicating and GC
rich.

Bright field R-bands

These R-bands are approximately the reverse of G-bands (the R stands for "reverse"). The dark
regions are euchromatic and the bright regions are heterochromatic.

Fluorescent G- and R-bands

These bands are the photographic negative of the bright field versions. i.e. the reverse of the
bright field G-bands and R-bands.

Q-bands are like fluorescent G-bands, but certain heterochromatic regions are more brightly
stained with Q-banding.

Methods of Chromosome Banding

Nearly all methods of chromosome banding rely on harvesting chromosomes in mitosis. This is
usually achieved by treating cells with tubulin inhibitors, such as colchicine or demecolcine
(colcemid), that depolymerize the mitotic spindle and so arrest the cell at this stage. Excessively
long incubations with Colcemid result in overcondensed chromosomes that band poorly and

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moreover some cell types, especially those from the mouse, eventually escape the Colcemid
block and proceed through the cell cycle.

Chromosome banding methods are either based on staining chromosomes with a dye or on
assaying for a particular function. The most common methods of dyebased chromosome banding
are G-(Giemsa), R-(reverse), C-(centromere) and Q-(quinacrine) banding.

Bands that show strong staining are referred to as positive bands; weakly staining bands are
negative bands. However the staining patterns are not black and white, different bands stain to
different intensities.

G-positive bands are usually just called G-bands and likewise for R-positive (R-) bands. Positive
C-bands contain constitutive heterochromatin. Q-bands are considered equivalent to Gbands.

The most widely used function-based banding method is replication banding and is based on the
fact that different bands replicate their DNA at different times during S phase of the cell cycle.
Generally, R-band DNA is replicated earlier than G-bands. G-bands also correspond to the
condensed chromomeres of meiotic chromosomes and R-bands to the interchromomeric regions.

Uses of Chromosome Banding

G-and R-banding are the most commonly used techniques for chromosome identification
(karyotyping) and for identifying abnormalities of chromosome number, translocations of
material from one chromosome to another, and deletions, inversions or amplifications of
chromosome segments. This has had an invaluable impact on human genetics and medicine and
the power of this approach has been augmented by combining cytogenetics with fluorescence in
situ hybridization (FISH). The detection of chromosome deletions associated with disorders,
very often contiguous gene syndromes, provided some of the first disease gene localizations in
humans.

Similarly, translocations have been important in pinpointing the location of disease-associated

genes and the characteristic translocations associated with some leukaemias is important, not
only for understanding the molecular basis of these cancers, but also for their diagnosis and
prognosis. One of the best examples of this is the translocation between human chromosomes 9
and 22 t(9;22)(q34:q11) or the Philadelphia chromosome diagnostic of chronic myelogenous
leukaemia (CML). Comparisons of chromosome banding patterns can confirm evolutionary
relationships between species and also reveal changes in karyotype that may have been important
in speciation.

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The banding patterns of human, gorilla and chimpanzee chromosomes are almost identical,
though human chromosome 2 is the result of a fusion between two great ape chromosomes.
There are also extensive similarities between human chromosome bands and those of lower
primates.

Using Karyograms to Detect Chromosomal Abnormalities

Today, G-banded karyograms are routinely used to diagnose a wide range of chromosomal
abnormalities in individuals. Although the resolution of chromosomal changes detectable by
karyotyping is typically a few megabases, this can be sufficient to diagnose certain categories of
abnormalities. For example, aneuploidy, which is often caused by the absence or addition of a
chromosome, is simple to detect by karyotype analysis. Cytogeneticists can also frequently
detect much more subtle deletions or insertions as deviations from normal banding patterns.
Likewise, translocations are often readily apparent on karyotypes. When regional changes in
chromosomes are observed on karyotypes, researchers often are interested in identifying
candidate genes within the critical interval whose mis-expression may cause symptoms in
patients.

This search process has been greatly facilitated by the completion of the Human Genome
Project, which has correlated cytogenetic bands with DNA sequence information. Consequently,
investigators are now able to apply a range of molecular cytogenetic techniques to achieve even
higher resolution of genomic changes. Fluorescence in situ hybridization (FISH)
and comparative genomic hybridization (CGH) are examples of two approaches that can
potentially identify abnormalities at the level of individual genes. Molecular cytogenetics is a
dynamic discipline, and new diagnostic methods continue to be developed. As these new
technologies are implemented in the clinic, we can expect that cytogeneticists will be able to
make the leap from karyotype to gene with increasing efficiency.

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 Role of the Xist Gene in X Chromosome Choosing
XIST (X Inactive Specific Transcript (Non-Protein Coding)) is an RNA Gene, and is affiliated
with the non-coding RNA class.

X inactivation is an early developmental process in mammalian females that transcriptionally

silences one of the pair of X chromosomes, thus providing dosage equivalence between males
and females. The process is regulated by several factors, including a region of chromosome X
called the X inactivation center (XIC). The XIC comprises several non-coding and protein-
coding genes, and this gene was the first non-coding gene identified within the XIC. This gene is
expressed exclusively from the XIC of the inactive X chromosome, and is essential for the
initiation and spread of X-inactivation. The transcript is a spliced RNA. Alternatively spliced
transcript variants have been identified, but their full length sequences have not been determined.
Mutations in the XIST promoter cause familial skewed X inactivation.

Diseases associated with XIST include X-Inactivation, Familial Skewed and Testicular Cancer.

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Eukaryotic Chromosome Structure

The length of DNA in the nucleus is far greater than the size of the compartment in which it is
contained. To fit into this compartment the DNA has to be condensed in some manner. The
degree to which DNA is condensed is expressed as its packing ratio.

Packing ratio - the length of DNA divided by the length into which it is packaged

For example, the shortest human chromosome contains 4.6 x 107 bp of DNA (about 10 times the
genome size of E. coli). This is equivalent to 14,000 m of extended DNA. In its most
condensed state during mitosis, the chromosome is about 2 m long. This gives a packing ratio
of 7000 (14,000/2).

To achieve the overall packing ratio, DNA is not packaged directly into final structure of
chromatin. Instead, it contains several hierarchies of organization. The first level of packing is
achieved by the winding of DNA around a protein core to produce a "bead-like" structure called
a nucleosome. This gives a packing ratio of about 6. This structure is invariant in both the
euchromatin and heterochromatin of all chromosomes. The second level of packing is the coiling
of beads in a helical structure called the 30 nm fiber that is found in both interphase chromatin
and mitotic chromosomes. This structure increases the packing ratio to about 40. The final
packaging occurs when the fiber is organized in loops, scaffolds and domains that give a final
packing ratio of about 1000 in interphase chromosomes and about 10,000 in mitotic
chromosomes.

Eukaryotic chromosomes consist of a DNA-protein complex that is organized in a compact

manner which permits the large amount of DNA to be stored in the nucleus of the cell. The
subunit designation of the chromosome is chromatin. The fundamental unit of chromatin is the
nucleosome.

Chromatin - the unit of analysis of the chromosome; chromatin reflects the general structure of
the chromosome but is not unique to any particular chromosome

Nucleosome - simplest packaging structure of DNA that is found in all eukaryotic chromosomes;
DNA is wrapped around an octamer of small basic proteins called histones; 146 bp is wrapped
around the core and the remaining bases link to the next nucleosome; this structure causes
negative supercoiling

The nucleosome consists of about 200 bp wrapped around a histone octamer that contains two
copies of histone proteins H2A, H2B, H3 and H4. These are known as the core histones.
Histones are basic proteins that have an affinity for DNA and are the most abundant proteins
associated with DNA. The amino acid sequence of these four histones is conserved suggesting a
similar function for all.

The length of DNA that is associated with the nucleosome unit varies between species. But
regardless of the size, two DNA components are involved. Core DNA is the DNA that is
actually associated with the histone octamer. This value is invariant and is 146 base pairs. The

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core DNA forms two loops around the octamer, and this permits two regions that are 80 bp apart
to be brought into close proximity. Thus, two sequences that are far apart can interact with the
same regulatory protein to control gene expression. The DNA that is between each histone
octamer is called the linker DNA and can vary in length from 8 to 114 base pairs. This variation
is species specific, but variation in linker DNA length has also been associated with the
developmental stage of the organism or specific regions of the genome.

The next level of organization of the chromatin is the 30 nm fiber. This appears to be a solenoid
structure with about 6 nucleosomes per turn. This gives a packing ratio of 40, which means that
every 1 m along the axis contains 40 m of DNA. The stability of this structure requires the
presence of the last member of the histone gene family, histone H1. Because experiments that
strip H1 from chromatin maintain the nucleosome, but not the 30 nm structure, it was concluded
that H1 is important for the stabilization of the 30 nm structure.

The final level of packaging is characterized by the 700 nm structure seen in the metaphase
chromosome. The condensed piece of chromatin has a characteristic scaffolding structure that
can be detected in metaphase chromosomes. This appears to be the result of extensive looping of
the DNA in the chromosome.

The last definitions that need to be presented are euchromatin and heterochromatin. When
chromosomes are stained with dyes, they appear to have alternating lightly and darkly stained
regions. The lightly-stained regions are euchromatin and contain single-copy, genetically-active
DNA. The darkly-stained regions are heterochromatin and contain repetitive sequences that are
genetically inactive.

Centromeres and Telomeres

Centromeres and telomeres are two essential features of all eukaryotic chromosomes. Each
provide a unique function that is absolutely necessary for the stability of the chromosome.
Centromeres are required for the segregation of the centromere during me iosis and mitosis, and
teleomeres provide terminal stability to the chromosome and ensure its survival.

Centromeres are those condensed regions within the chromosome that are responsible for the
accurate segregation of the replicated chromosome during mitosis and meiosis. When
chromosomes are stained they typically show a dark-stained region that is the centromere.
During mitosis, the centromere that is shared by the sister chromatids must divide so that the
chromatids can migrate to opposite poles of the cell. On the other hand, during the first meiotic

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division the centromere of sister chromatids must remain intact, whereas during meiosis II they
must act as they do during mitosis. Therefore the centromere is an important component of
chromosome structure and segregation.

Within the centromere region, most species have several locations where spindle fibers attach,
and these sites consist of DNA as well as protein. The actual location where the attachment
occurs is called the kinetochore and is composed of both DNA and protein. The DNA sequence
within these regions is called CEN DNA. Because CEN DNA can be moved from one
chromosome to another and still provide the chromosome with the ability to segregate, these
sequences must not provide any other function.

Typically CEN DNA is about 120 base pairs long and consists of several sub-domains, CDE-I,
CDE-II and CDE-III. Mutations in the first two sub-domains have no effect upon segregation,
but a point mutation in the CDE-III sub-domain com pletely eliminates the ability of the
centromere to function during chromosome segregation. Therefore CDE-III must be actively
involved in the binding of the spindle fibers to the centromere.

The protein component of the kinetochore is only now being characterized. A complex of three
proteins called Cbf-III binds to normal CDE-III regions but can not bind to a CDE-III region
with a point mutation that prevents mitotic segregation. Fur thermore, mutants of the genes
encoding the Cbf-III proteins also eliminates the ability for chromosomes to segregate during
mitosis. Additional analyses of the DNA and protein components of the centromere are
necessary to fully understand the mechanics of chromosome segregation.

Telomeres are the region of DNA at the end of the linear eukaryotic chromosome that are
required for the replication and stability of the chromosome. McClintock recognized their special
features when she noticed, that if two chromosomes were broken in a cell, the end of one could
attach to the other and vice versa. What she never observed was the attachment of the broken end
to the end of an unbroken chromosome. Thus the ends of broken chromosomes are sticky,
whereas the normal end is not sticky, suggesting the ends of chromosomes have unique features.
Usually, but not always, the telomeric DNA is heterochromatic and contains direct tandemly
repeated sequences. The following table shows the repeat sequences of several species. These are
often of the form (T/A)xGy where x is between 1 and 4 and y is greater than 1.

Telomere Repeat Sequences

Species Repeat Sequence

Arabidopsis TTTAGGG
Human TTAGGG
Oxytricha TTTTGGGG
Slime Mold TAGGG
Tetrahymena TTGGGG
Trypanosome TAGGG

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 Yeast (TG)1-3TG2-3

Notice that the number of TG sequences and the number of cytosines in the yeast sequence
varies. At least for yeast, it has been shown that different strains contain different lengths of
teleomeres and that the length is under genetic control.

The primary difficulty with telomeres is the replication of the lagging strand. Because DNA
synthesis requires a RNA template (that provides the free 3'-OH group) to prime DNA
replication, and this template is eventually degraded, a short single-stranded region would be left
at the end of the chromosome. This region would be susceptible to enzymes that degrade single-
stranded DNA. The result would be that the length of the chromosome would be shortened after
each division. But this is not seen.

The action of the telomerase enzymes ensure that the ends of the lagging strands are replicated
correctly. A well-studied system involves the Tetrahymena protozoa organism. The telomeres of
this organism end in the sequence 5'-TTGGGG-3'. T he telomerase adds a series of 5'-TTGGGG-
3' repeats to the ends of the lagging strand. A hairpin occurs when unusual base pairs between
guanine residues in the repeat form. Next the RNA primer is removed, and the 5' end of the
lagging strand can be used for DNA synthesis. Ligation occurs between the finished lagging
strand and the hairpin. Finally, the hairpin is removed at the 5'-TTGGGG-3' repeat. Thus the end
of the chromosome is faithfully replicated. The following figure shows these steps.

The Replication of Telomeres

Analysis of DNA Sequences in Eukaryotic Genomes

The technique that is used to determine the sequence complexity of any genome involves the
denaturation and renaturation of DNA. DNA is denatured by heating which melts the H-

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bonds and renders the DNA single-stranded. If the DNA is rapidly cooled, the DNA remains
single-stranded. But if the DNA is allowed to cool slowly, sequences that are complementary
will find each other and eventually base pair again. The rate at which the DNA reanneals
(another term for renature) is a function of the species from which the DNA was isolated. Below
is a curve that is obtained from a simple genome.

The Y-axis is the percent of the DNA that remains single stranded. This is expressed as a ratio of
the concentration of single-stranded DNA (C) to the total concentration of the starting DNA
(Co). The X-axis is a log-scale of the product of the initial concentration of DNA (in moles/liter)
multiplied by length of time the reaction proceeded (in seconds). The designation for this value is
Cot and is called the "Cot" value. The curve itself is called a "Cot" curve. As can be seen the
curve is rather smooth which indicates that reannealing occurs slowing but gradually over a
period of time. One particular value that is useful is Cot , the Cot value where half of the DNA
has reannealed.

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.

The shape of a "Cot" curve for a given species is a function of two factors:

1. the size or complexity of the genome; and

2. the amount of repetitive DNA within the genome

If we plot the "Cot" curves of the genome of three species such as bacteriophage lambda, E. coli
and yeast we will see that they have the same shape, but the Cot of the yeast will be largest, E.
coli next and lambda smallest. Physically, the larger the genome size the longer it will take for
any one sequence to encounter its complementary sequence in the solution. This is because two
complementary sequences must encounter each other before they can pair. The more complex
the genome, that is the more unique sequences that are available, the longer it will take for any
two complementary sequences to encounter each other and pair. Given similar concentrations in
solution, it will then take a more complex species longer to reach Cot .

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Repeated DNA sequences, DNA sequences that are found more than once in the genome of
the species, have distinctive effects on "Cot" curves. If a specific sequence is represented twice
in the genome it will have two complementary sequences to pair with and as such will have a Cot
value half as large as a sequence represented only once in the genome.

Eukaryotic genomes actually have a wide array of sequences that are represented at different
levels of repetition. Single copy sequences are found once or a few times in the genome.
Many of the sequences which encode functional genes fall into this class. Middle repetitive
DNA are found from 10s - 1000 times in the genome. Examples of these would include rRNA
and tRNA genes and storage proteins in plants such as corn. Middle repetitive DNA can vary
from 100-300 bp to 5000 bp and can be dispersed throughout the genome. The most abundant
sequences are found in the highly repetitive DNA class. These sequences are found from
100,000 to 1 million times in the genome and can range in size from a few to several hundred
bases in length. These sequences are found in regions of the chromosome such as
heterochromatin, centromeres and telomeres and tend to be arranged as a tandem repeats. The
following is an example of a tandemly repeated sequence:

ATTATA ATTATA ATTATA // ATTATA

Genomes that contain these different classes of sequences reanneal in a different manner than
genomes with only single copy sequences. Instead of having a single smooth "Cot" curve, three
distinct curves can be seen, each representing a different repetition class. The first sequences to
reanneal are the highly repetitive sequences because so many copies of them exist in the genome,
and because they have a low sequence complexity. The second portion of the genome to reanneal
is the middle repetitive DNA, and the final portion to reanneal is the single copy DNA. The
following diagram depicts the "Cot" curve for a "typical" eukaryotic genome

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The following table gives the sequence distribution for selected species.

Species Sequence Distribution

Bacteria 99.7% Single Copy
60% Single Copy
Mouse 25% Middle Repetitive
10% Highly Repetitive
70% Single Copy
Human 13% Middle Repetitive
8% Highly Repetitive
61% Single Copy
Cotton 27% Middle Repetitive
8% Highly Repetitive
30% Single Copy
Corn 40% Middle Repetitive
20% Highly Repetitive
10% Single Copy
Wheat 83% Middle Repetitive
4% Highly Repetitive
55% Single Copy
Arabidopsis 27% Middle Repetitive
10% Highly Repetitive

Sequence Interspersion

Even though the genomes of higher organisms contain single copy, middle repetitive and highly
repetitive DNA sequences, these sequences are not arranged similarly in all species. The
prominent arrangement is called short period interspersion. This arrangement is characterized
by repeated sequences 100-200 bp in length interspersed among single copy sequences that are
1000-2000 bp in length. This arrangement is found in animals, fungi and plants.

The second type of arrangement is long-period interspersion. This is characterized by 5000 bp

stretches of repeated sequences interspersed within regions of 35,000 bp of single copy DNA.
Drosophila is an example of a species with this uncommon sequence arrangement. In both cases,

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the repeated sequences are usually from the middle repetitive class. We discussed above where
highly repetitive sequences are found.

Eukaryotic Chromosome Karyotype

Whereas bacteria only have a single chromosome, eukaryotic species have at least one pair of
chromosomes. Most have more than one pair. Another relevant point is that eukaryotic
chromosomes are detected only occur during cell division and not during all stages of the cell
cycle. They are in their most condensed form during metaphase when the sister chromatids are
attached. This is the primary stage when cytogenetic analysis is performed.

Each species is characterized by a karyotype. The karyotype is a description of the number of

chromosomes in the normal diploid cell, as well as their size distribution. For example, the
human chromosome has 23 pairs of chromosome, 22 somatic pairs and one pair of sex
chromosomes. One important aspect of genetic research is correlating changes in the karyotype
with changes in the phenotype of the individual.

One important aspect of genetics is correlating changes in karyotype with changes in phenotype.
For example, humans that have an extra chromosome 21 have Down's syndrome. Insertions,
deletions and changes in chromosome number can be detected by the skilled cytogeneticist, but
correlating these with specific phenotypes is difficult.

The first discriminating parameter when developing a karyotype is the size and number of the
chromosomes. Although this is useful, it does not provide enough detail to be begin the
development of a correlation between structure and function (phenotype). To further distinguish
among chromosomes, they are treated with a dye that stains the DNA in a reproducible manner.
After staining, some of the regions are lightly stained and others are heavily stained. As
described above, the lightly stained regions are called euchromatin, and the dark stained region
is called heterochromatin. The current dye of chose is the Giemsa stain, and the resulting
pattern is called the G-banding pattern.

C-Value Paradox

In addition to describing the genome of an organism by its number of chromosomes, it is also

described by the amount of DNA in a haploid cell. This is usually expressed as the amount of
DNA per haploid cell (usually expressed as picograms) or the number of kilobases per haploid
cell and is called the C value. One immediate feature of eukaryotic organisms highlights a
specific anomaly that was detected early in molecular research. Even though eukaryotic
organisms appear to have 2-10 times as many genes as prokaryotes, they have many orders of
magnitude more DNA in the cell. Furthermore, the amount of DNA per genome is correlated not
with the presumed evolutionary complexity of a species. This is stated as the C value paradox:
the amount of DNA in the haploid cell of an organism is not related to its evolutionary
complexity.

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