Introduction

Laboratory is a place where ideas and concepts can be tested through

experiments. Biology, like any other discipline of science, is based on
experimental work and therefore practical forms an integral part of learning.
Biology laboratory provides a unique learning environment where learners
inculcate scientific temper, develop relevant skills and get exposed to realms
of techniques and methodologies of scientific investigations. Laboratory
investigations in Biology increase the reasoning abilities, bring scientific
attitude in a learner and also help in acquisition of skills of scientific processes.
Also, observation of nature and the living organisms found in it is no less
important for the understanding of many aspects of the subject especially
the diversity of the living organisms, their systematic study, their relationships
among themselves and with the environment. Knowledge in the field of
Biology can be acquired or constructed only on the basis of correct
observations and experimentally verifiable processes.
Biology laboratory thus provides the learners an environment where the
process of learning is facilitated by hands-on experiments. Biology is a
unique discipline in the sense that it does not merely deal with the study of
morphology, anatomy, physiology and reproduction of the living organisms,
rather, understanding of the subject requires understanding of a number of
interdisciplinary areas and approaches. On one hand, a biologist needs to
be sufficiently skilled in handling the enormous diversity of the living
organisms, be it plants, animals, fungi or even microscopic bacteria, while
on the other hand, a biologist should be able to understand the biochemical,
molecular, physiological, behavioural, genetic and many other phenomena
pertaining to the living organisms. The study of intricate relationship of
different types of organisms among themselves and also with its environment
is an important concern of a biologist. Thus, experiments and exercises in
Biology train a learner about skills of observations, manipulation of the
organisms for the study of internal details, biochemical as well as molecular
composition and processes, investigation of the abiotic environment and even
analysis of phenomena like inheritance and evolution.
As far as the study of the living organism is concerned, correctness of the
method is very important. Such a study may be very simple, e.g., study of
habit, habitat and external features of the plants or animals, or, it may involve
certain manipulations like dissection and section cutting of the parts of the
organisms to study the minute details. Very often observation and study of
the magnified image of the minute parts under a microscope provides a
better insight about the features of the organisms. However, microscopic
study involves certain specific skills depending on type of the organisms/
tissues/cells to be studied. It involves specific preparations (peeling, section
 LABORATORY MANUAL: BIOLOGY

cutting, fixation, staining, dehydration, mounting, etc.) so that microscopic

examination reveals the expected details. As histological and cytological
observations give us only static pictures of the continuous processes, analysis
of biochemical, physiological and ecological aspects need certain other kinds
of skills such as preparation of chemicals and reagents, designing and
performing an experiment, observation and recording of data and ultimately
interpretation and drawing conclusions. While performing experiments,
honesty in recording of data and its correct presentation is very important
as it is not only useful in the logical interpretation but also helps in the
identification of errors.
In order to perform experiments successfully, a learner needs to go to
the Biology laboratory well prepared. This includes the following:
1. Laboratory Record Book: For maintaining all the information including
recording of data and its interpretation.
2. Dissection Box: A dissection box is required in the Biology laboratory
for various purposes like handling and manipulation of living materials,
performing experiments, preparation of slide, etc. A dissection box
should contain scissors (two pairs, one small with fine tip and one
larger), scalpels (one small and one medium sized), forceps (two, one
small with sharp fine tips and the other medium sized with blunt tips),
dissecting needles (two), razor, hand lens, dropper, fine brush, etc.
3. Laboratory Manual
4. A Laboratory Coat or Apron
5. A Hand Towel
While in the laboratory a student should be very careful and methodical.
One should listen carefully to the instructions given by the teacher/
instructor before performing an experiment. In the biology laboratory a
student has to handle a number of sharp objects and hence necessary
precaution and care should always be taken while handling objects like
scissors, forceps, needles, scalpel, razor, etc. It is also very important to
follow the safety instructions mentioned on the instruments and/or on the
label of the reagent/chemical. Students should also be aware about the
use of the First-aid Box so that in case of any accident or injury the
preliminary aid can be provided to the affected person.
While describing the experiment students are expected to follow a pattern
in which the aim of the experiment, its principle, list of the materials to be
used, procedure, observation table (if required), inference and discussion
should be given. Necessary precautions to be taken should also be
mentioned appropriately in the procedure or at the end. There are a few
experiments in which field visit is essentially required. For this all the
necessary preparations (materials, equipments, reagents and chemicals)

2
E XERCISE 1
I NTRODUCTION

should be made in advance. Drawing of illustrations is also an important

component of the practical in Biology. Students are expected to follow certain
fundamental rules while drawing the illustrations so that it reflects your
observations correctly.
• Make your illustrations using pencil only and always use white drawing
sheet. Illustration should be in the centre of the page.
• Drawing of an object (plant, animal or experimental set-up) should be
proportionate in size.
• Draw your illustrations keeping the object before you.
• Drawing must be clear with simple outlines.
• Appropriately label your drawing. Parts of the drawing should be
indicated by straight horizontal line or arrow. Two lines or arrow should
never cross each other. As far as possible, labelling should be done on
the right side of the drawing. An appropriate legend or heading of the
drawing should also be given below it.
About the Manual
The main objective of the manual is to introduce the students of higher
secondary stage to the fascinating world of plants, animals and microbes
and their complex biological phenomena. The manual covers a complete
description of the experiments and exercises. The suggested experiments
cover almost all the units/topics including those on diversity in living world,
plant, animal and human physiology, genetics, bio-technology and human
welfare and environment. A standard format has been used to describe each
experiment which includes
• Aim: It gives a brief title of the experiment under investigation.
• Principle: It is a very brief introduction of the experiment under
investigation and explains the biological phenomenon involved. It gives
brief but comprehensive ideas about the design of the experiment and
explains the significance of the phenomenon being studied.
• Materials required: This includes the names of plants/animals to be
used as 'samples', the type of apparatus, the type and quantity of
glasswares required, reagents, chemicals and solutions needed, their
concentration and other specifications, method of preparations of
solutions and reagents. If a particular material/chemical/glassware is
not available, sufficient alternatives have been suggested.
• Procedure: This section includes full details of experimental procedure
explained stepwise, including special precautions necessary to be taken
while the experiment is being conducted. Drawings of the samples,
apparatus and the experimental setup, wherever found necessary, have
been included to facilitate the students to perform the experiment as
accurately as possible.

3
 LABORATORY MANUAL: BIOLOGY

• Observation and Results: This section deals with the recording of all
observations made during the experiment. Students are advised to
consider the entire data. Data can be represented in the form of tables,
graphs and histograms wherever possible. Use of units in which various
quantities are measured has been indicated in the manual.
• Discussion: Included in this heading is a statement of the conclusions
drawn from the experimental results and compared thesis (wherever
possible) with any comparable data from other sources. The relevance
of the conclusions drawn from the experimental results to the various
processes under investigation and to the life of plant, animal and microbes
has been prompted out.
• Precaution: This section contains all the necessary precautions to be
taken during experimentation to obtain results free of errors. However,
attempts have been made to mention required precautions along with
the procedure also.
A great emphasis has been laid on a student getting valid results and
interpreting them. It is essential that the teacher should properly explain
each experiment so that inexperienced students will be able to obtain accurate
results within a reasonable time. Teachers are also expected to help students
in identifying errors and mistakes committed during experiments and ways
for correcting them. It is possible that some of the students may undoubtedly
be capable of doing more sophisticated work than that represented in the
manual. But introductory course of this sort has been designed to help all
students for some useful and joyful experience by conducting the
experiments of their own. The manual also aims that students and teachers
not be discouraged by either incomplete experiments or experiments which
yield apparently meaningless results.
With the objectives of inculcating scientific temper among learners and
providing them an opportunity to undertake independent scientific
investigation, Investigatory Project Work has been included as an integral
part of the practical curriculum of Class XII. Such investigatory projects
are expected to provide thrill in the learning process. It is also expected to
serve the real purpose of practicals, i.e., developing an ability to hypothesise
and design experiments to address certain problems, to make observations
methodically and to draw conclusions out of the experimental data. A
comprehensive idea about undertaking investigatory project has been given
in the book with a list of a few problems on which investigatory project
work can be undertaken. However, the list is only suggestive and considering
the wider scope students can undertake any kind of investigatory project
work depending on their region, its climatic condition, availability of
resources, etc.

4
Exercise 3
Aim: To study pollen tube growth on stigma

Principle: Pollen grains germinate and form pollen tubes after they get deposited by the process
of pollination on compatible stigma. Pollen tube, made up of cellulose, is an extension of the
inner wall of pollen grain (intine). It emerges through one of the germ pore and passes through
tissues of stigma and style to reach the ovule. The growing pollen tube is observed by staining
with cotton blue.

Requirement: 5–6 excised styles with stigma of Petunia/grass/maize/sunflower/Abelmoschus

(Lady's finger), beaker, water, slides, cover slips, cotton blue stain, microscope, brush, needle.

Procedure
(i) Place the stigmas in boiling water in a beaker
for softening the tissues for 5–10 minutes. Pollen grains
(ii) Stain with cotton blue for 3–5 minutes and
wash with water to remove excess stain.
(iii) Mount one stigma in a drop of glycerine on a Pollen tube
slide. Place a cover slip on the stigma and gently
press the cover slip on the material. Observe Style
the slide under a microscope.
(iv) If you fail to observe pollen tubes mount
another stigma.

Observation
Look for long blue-coloured tubular structures
traversing through the tissues of stigma and Fig.3.1 Growth of pollen tube
style (Fig. 3.1). in the style of a carpel
EXERCISE 3

Discussion
Pollen tubes are seen amidst the stylar tissue. Many pollen tubes may be
seen. Trace the origin of pollen tubes to the pollen grains present around
the surface of the stigma.

Questions
1. Can pollen grains of one plant species germinate on stigma of other species?
Give reasons.
2. Do all pollen tubes reach the ovules?
3. Are all the pollen tubes of equal length? If not, why?

15
Exercise 4
Aim: To study the discrete stages of gametogenesis in mammalian testis and ovary

Principle: In all male and female organisms gamete formation takes place in their gonads, i.e.,
testis and ovary respectively. The process of gamete formation, called gametogenesis involves
meiotic cell division. The gametogenic development in testis is called spermatogenesis and in
ovary it is oogenesis. They exhibit marked differences and can be examined in transverse section
(T.S.) of these organs.

Requirement: Permanent slides of T.S. of testis and ovary, compound microscope,

lens-cleaning paper and cleaning fluid

Procedure
(i) Clean the slide and microscope’s eye and objective lenses with the
help of lens cleaning paper using any cleaning fluid.
(ii) Place the slide on the stage of the microscope and observe first under
lower magnification and then in higher magnification. Observe various
stages of gamete development.
(iii) Record your observations in the notebook and draw labelled diagrams.

Observation
T.S. of testis
(i) You will observe a large number
of seminiferous tubules under
Seminiferous tubule
lower magnification. Observe a
complete tubule in higher
magnification and view various
stages of gamete development Spermatozoa
from periphery towards lumen
(Fig. 4.1) and identify the
following types of cells namely, Germinal
Epithelium
Germinal epithelium,
Spermatogonial cells, Primary Spermatogonia
spermatocytes, Secondary
spermatocytes, Spermatids and
Spermatozoa. Fig. 4.1 T.S. of mammalian testis
Exercise 6
Aim: Preparation and study of mitosis in onion root tips

Principle: Somatic growth in plants and animals takes place by the increase in the number of
cells. A cell divides mitotically to form two daughter cells wherein the number of chromosomes
remains the same (i.e., unchanged) as in the mother cell. In plants, such divisions rapidly take
place in meristematic tissues of root and shoot apices, where the stages of mitosis can be
easily observed. In animals, mitotically dividing cells can be easily viewed in the bone marrow
tissue of a vertebrate, epithelial cells from gills in fishes and the tail of growing tadpole larvae
of frog.

Requirement: Onion bulbs, wide mouth glass tubes/jar/bottle, glacial acetic acid, ethanol
2-4% acetocarmine/acetoorcein stain, N/10 HCl, spirit lamp/hot plate, slide, cover slips,
blotting paper, molten wax/nail polish and compound microscope

Procedure
Growing of root tips
Select a few medium-sized onion bulbs. Carefully remove the dry roots
present. Grow root tips by placing the bulbs on glass tubes (of about 3–4
cm. diameter) filled with water. Care should be taken so that the stem portion
of the bulb (basal part) just touches the water. A few drops of water may be
added periodically to compensate evaporation losses. New roots may take
3–6 days to grow. Cut 2–3 cm long freshly grown roots and transfer them to
freshly prepared fixative, i.e., aceto-alcohol ([Link] glacial acetic acid : ethanol).
Keep the root tips in the fixative for 24 hours and then transfer them to 70%
ethanol (for preservation and use in future). Onion root-tip cells have a cell
cycle of approximately 24-hour duration, i.e., they divide once in 24 hours,
and this division usually takes place about two hours after sunrise. Therefore,
roots grown on water should be cut only at that time to score maximum
number of dividing cells.

Preparation of slide
Take one or two preserved roots, wash them in water on a clean and grease-
free slide. Place one drop of N/10 HCl on the root tip followed by 2–3 drops
of aceto-carmine or aceto-orcein stain on it. Leave the slide for 5–10 minutes
EXERCISE 6

on a hot plate (or warm it slightly on spirit lamp). Care should be taken that
the stain is not dried up. Carefully blot the excess stain using blotting paper.
Now cut the comparatively more stained (2–3 mm) tip portion of the root
and retain it on the slide and discard the remaining portion. After
(10–20 seconds) put one or two drops of water and blot them carefully using
blotting paper. Again put a drop of water on the root tip and mount a cover
slip on it avoiding air bubbles. Place the slide in between the folds of blotting
paper using the fingers in such a way that the cover slip mounted on the
slide is properly held. Now slowly tap the cover slip using the blunt end of a
pencil so that the meristematic tissue of the root tip below the cover slip is
properly squashed and spread as a thin layer of cells. Carefully seal the
margins of the cover slip using molten paraffin wax or nail polish. This
preparation of onion root tips cells is now ready for the study of mitosis.

Study of slide
Place the slide on the stage of a good quality compound microscope. First
observe it under the lower magnification (10 X objective) to search for the
area having a few dividing cells. Examine the dividing cells under higher
magnification of the microscope to observe the detailed features of mitosis.

Observation
The stages of mitosis can be broadly categorised into two parts: karyokinesis
(division of nucleus) followed by cytokinesis (division of cytoplasm, and
ultimately of the cell). Those cells, which are not in the phases of cell division
are considered to be in interphase. You may observe that most of the cells
in a microscope field are in interphase

Interphase
The cells are mostly rectangular, oval or even circular in shape, with almost
centrally situated densely stained nucleus. The chromatic (coloured) material
of the nucleus is homogeneous and looks granular. The boundary of the
nucleus is distinct. One or few nucleoli (sing: nucleolus) can also be observed
inside the nucleus (Fig. 6.1a).

Stages of Mitosis
(a) Prophase
Intact nuclear outline is seen. The chromatin (seen as a homogeneous
material in the nucleus at interphase) appears as a network of fine threads
(chromosomes). Nucleoli may or may not be visible (Fig. 6.1b).

21
 LABORATORY MANUAL: BIOLOGY

a. Interphase

b. Prophase

c. Metaphase

d. Anaphase

e. Telophase

Fig.6.1 Interphase (a) and stages of mitosis (b - e) – actual microscopic

view on left side and its diagrammatic representation on the
right hand side
If the cell under observation is in the early stage of prophase then the
chromatin fibres (chromosomes) are very thin. However, in the cells
at late prophase, comparatively thicker chromatin fibres would be
visible. Besides this, in the late prophase the nuclear membrane
may not be noticed.

(b) Metaphase
The nuclear membrane disappears. Chromosomes are thick and are seen
arranged at the equatorial plane of the cell (Fig. 6.1c). Each chromosome at

22
EXERCISE 6

this stage has two chromatids joined together at the centromere, which can
be seen by changing the resolution of the microscope. Nucleolus is not
observed during metaphase.
(c) Anaphase
This stage shows the separation of the chromatids of each chromosome. The
chromatids separate due to the splitting of the centromere. Each chromatid
now represents a separate chromosome as it has its own centromere. The
chromosomes are found as if they have moved towards the two poles of the
cell. The chromosomes at this stage may look like the shape of alphabets 'V',
'J' or 'I' depending upon the position of centromere in them. Different anaphase
cells show different stages of movement of chromosomes to opposite poles,
and they are designated to represent early, mid and late anaphase (Fig. 6.1d).
(d) Telophase
Chromosomes reach the opposite poles, lose their individuality, and look
like a mass of chromatin (Fig. 6.1e). Nuclear membrane appears to form the
nuclei of the two future daughter cells.

Cytokinesis
In plants, a cell plate is formed in the middle after telophase. The plate can
be seen to extend outwards to ultimately reach the margin of the cell and
divide the cell into two. Such cell plates are characteristic of plant cells
(Fig. 6.2). However, in an animal cell, the two sides of the cell show inpushings
or constrictions formed from the peripheral region in the middle of the cell,
which grow inward and meet to divide the cell into two daughter cells.
Draw labelled diagrams of all the phases of mitosis.

Fig. 6.2 Cytokinesis

Discussion
Mitotic index (MI) is defined as a ratio of the total number of dividing cells (n)
and the total number of cells (N) in a particular focus chosen randomly under
n
the microscope and is calculated as MI = N ×100 . By randomly selecting
5 to 10 such foci, one can estimate the mitotic index for a given type.

23
 LABORATORY MANUAL: BIOLOGY

The effect of different samples of water (polluted or contaminated) can be

assayed on the mitotic-index (an indicative feature of somatic growth rate in
them).
Further, the impact of different types of pollutants on different phases of
mitosis can also be assayed.

Tabulate your observations in the tabular form given below

Features Interphase Karyokinesis Cytokinesis

Prophase Metaphase Anaphase Telophase

1. Cell morphology

2. Nuclear morphology

3. Chromosomes/chromatids

Questions
1. Suggest names of a few tissues, which are suitable for the study of mitosis.
2. Why is mitosis also known as equational division?
3. What shape would a metacentric and a sub-metacentric chromosome exhibit
during the anaphase stage?
4. How does cytokynesis differ in plant and animal cells?

24
Exercise 7
Aim: Study of stages of meiosis using permanent slides

Principle: Meiosis is a type of cell division in which the number of chromosomes is halved
(from diploid to haploid) in the daughter cells, i.e., the gametes. The division is completed in
two phases, meiosis I and meiosis II. Meiosis I is a reductional division in which the chromosomes
of homologous pairs separate from each other. Meiosis II is equational division resulting in the
formation of four daughter cells. Stages of meiosis can be observed in a cytological preparation
of the cells of testis tubules or in the pollen mother cells of the anthers of flower buds.

Requirement: Permanent slides of meiosis and compound microscope

Procedure
Place the slide on the stage of the microscope and search for the dividing
cells using lower magnification. When dividing cells are located observe them
under higher magnification.

Observation
Observe various stages of meiosis and identify them on the basis of the specific
features given in the table 7.1. A significant number of cells will be in the
Interphase. These cells have a centrally positioned densely stained nucleus.
In case of slide of animal tissue a few mitotically dividing spermatogonial
cells may also be seen.
Table 7.1 Different stages of meiosis and their features
Meiosis I
1. Prophase I Unlike the prophase of mitosis, it is a comparatively complex phase
characterised by a number of events. Five sub-phases can be
identified in it.
(a) Leptotene (leptos = slender tene = band or thread)
(i) The nuclear membrane and nucleolus are not distinctly
observable (Fig. 7.1 a).
(ii) Fine network of thin threads are seen uniformly distributed
in the [Link] are chromatin threads, which may be
observed as more prominent structures in the later stages.
 LABORATORY MANUAL: BIOLOGY

(a) Leptotene

(b) Zygotene

(c) Pachytene

(d) Diplotene-Diakinesis

Fig. 7.1 Sub-phase of Prophase I (a-d) – actual microscopic view on left

side and its diagrammatic representation on the right hand side

26
EXERCISE 7

(b) Zygotene (Zygon = paired)

This stage is characterised by the pairing of the homologous
chromosomes, which can be seen as paired chromatin threads
(bivalents) (Fig. 7.1b).
(c) Pachytene (pachy = thick)
The chromatin threads get condensed and appear shortened and
thick. Pairs of homologous chromosomes can be seen. Each
chromosome has two chromatids and thus each bivalent consists
of four chromatids. This configuration is called tetrad (Fig. 7.1c).
(d) Diplotene (diplos = double)
The homologous chromosomes (each made up of two chromatids)
show distinct separation from each other except at few regions where
attachments are seen (Fig. 7.1d). These are chiasmata (sing.
chiasma) representing the site of exchange of the parts between
two homologous chromosomes (i.e. crossing over).
(e) Diakinesis (Dia = opposite; kinesis= separation or movement)
(i) The homologous pair of chromosomes appear more shortened,
thick and prominent (Fig. 7.1d).
(ii) Chiasmata can be still observed.
(iii) All the homologous pairs appear scattered in the cell.
2. Metaphase I Homologous chromosomes are still in pairs, and are arranged along
the equatorial plane of the cell (Fig. 7.2a). At this stage, the number
of bivalents can be counted. Chiasmata may still be seen in a few
bivalents.

3. Anaphase I The chromosome pairs appear to have moved towards the two
opposite poles of the cell. At the later stage, the anaphase - I may
show the assembly of chromosomes at two poles (Fig. 7.2b). This
results into the reduction of number of chromosomes to half.
This stage can be identified by the presence of two chromatids in
each chromosome.

4. Telophase I The chromosomes present at the two poles appear decondensed

and form two distinct nuclei (Fig. 7.2c).
Note: After the telophase I stage there may or may not be cytokinesis.
Thereafter the cell enters into the second meiotic division.

Meiosis II

1. Prophase II (i) Distinct thread- like chromatin fibres or rod- shaped chromosome
are seen.

27
 LABORATORY MANUAL: BIOLOGY

(b) Anaphase I
(a) Metaphase I

(c) Telophase I

Fig. 7.2 Phases of Meosis I (a-c) – actual microscopic view on left side
and its diagrammatic representation on the right hand side.

(a) Metaphase II

(b) Anaphase II

Fig.7.3 Phases of Meosis II (a,b) – actual microscopic view on left side

and its diagrammatic representation on the right hand side.

28
EXERCISE 7

2. Metaphase II This phase is similar to that of mitotic division

(i) The chromosomes having two chromatids attached at the
centromere are observed arranged at the equatorial plane of the
cell.
Note: Metaphase II of meiosis can be differentiated from metaphase-I
on the basis of the following features:
(ii) Each chromosome of metaphase II has two chromatid
(Fig. 7.3a) whereas in metaphase I these are paired
homologous chromosomes each having two chromatids thus
forming tetrad.
(iii) In the metaphase I of meiosis, a few chiasmata are observed,
where as no chiasmata are observed during metaphase II.
3. Anaphase II The two chromatids of each chromosome after separation appear
to lie at the two poles of the cell (Fig. 7.3b).
Note: Anaphase II can also be distinguished from the anaphase I of
meiotic division on the basis of chromatids: In anaphase I, each
chromosome has two distinct chromatids, but in anaphase II, each
chromosome is represented by one chromatid only.
4. Telophase II The separated chromosomes appear decondensed and form nuclei
(Fig. 7.3c).

Questions
1. What is the significance of meiosis?
2. What is synapsis and crossing over?
3. How can anaphase I and anaphase II be distinguished from each other?
4. Indicate distinguishing feature of metaphase I of meiosis and metaphase of mitosis.
5. How many daughter cells are produced at the end of meiosis?
6. The daughter cells produced at the end of meiosis are genetically different. Explain.
7. What is the significance of synapsis?

29
Exercise 8
Aim: To study the blastula stage of embryonic development in mammals, with the help of
permanent slide, chart, model or photograph

Principle: The zygote undergoes a few cycles of mitotic divisions to form a solid ball of cells called
morula. The cells continue to divide and at a later stage a cavity is formed within it. This stage is
blastula. The internal structural details of blastula can be observed in its transverse section.

Requirement: Permanent slide, chart/model of T.S. of blastula, compound microscope,

lens cleaning fluid and paper

Procedure
Observe the slide under lower magnification of the microscope. In case of
chart/models/photographs, note the feature of blastula in your practical
record and draw labelled diagram.

Observation
In transverse section, the blastula appears as a sphere with a cavity, called
blastocoel within it (Fig. 8.1). Notice an outer layer of blastomeres called
trophoblasts. A cellular mass, adhered to the trophoblast is present on one
end of the blastula. It is called inner cell mass.

Fig.8.1 Blastula stage of a mammal

EXERCISE 8

Questions
1. What are the differences between blastula and morula?
2. What are the main structures you observe in T.S. blastula?
3. Match the stages in column I with features in column II
Column I Column II
(a) Trophoblast (i) Dividing cells of the morula
(b) Morula (ii) Outer layer of blastula
(c) Blastocoel (iii) Solid ball of cells
(iv) Cavity

31
Exercise 9
Aim: To verify Mendel's Law of Segregation

Principle: When two pure lines with contrasting forms of a particular character (phenotypes) are
crossed to produce the next generation (F1 generation), all the members of the progeny are of only
one phenotype i.e. of one of the two parents. The phenotype that appears is called dominant, and
the one that does not appear is called recessive. When the F1 plants are selfed, the progeny i.e. the
F2 generation is in the ratio of 3 dominant: 1 recessive (¾: ¼ or 75%: 25%). This reappearance of
the recessive phenotype in F2 generation verifies law of segregation.

Requirement: 64 yellow and 64 green plastic beads, all of exactly same shape and size, (when
beads are not available, pea seeds may be coloured using paint, these beads represent the
gametes of a specific trait), plastic beakers/petri dishes and a napkin/hand towel

Procedure
Students have to work in pairs to perform the experiment. The following
steps are to be strictly followed in the sequence mentioned below.
(i) Put 64 yellow beads in one beaker/petridish and 64 green beads in
the other to represent respectively male and female gametes. Let the
yellow bead be indicated by ‘Y’ and green bead by ‘y’.
(ii) Take a bead from each container and place them together (it represents
fertilisation) on the napkin spread before you on the table. (One student
to take out beads and to put in the hands of the other student who will
put them on the table).
(iii) Just like the previous step, continue to pick beads and arrange them
in pairs. Thus 64 pairs of beads are obtained representing the 64
heterozygous F1 progeny.
Note that all the F1 individuals are represented by one yellow and one
green bead.
(iv) Put 32 F1 progeny in one petridish and the remaining 32 in another
petridish (representing the F1 males and females).
(v) Stir the beads of each petridish with a pencil/pen for about 10 times
taking care that no bead falls off.
EXERCISE 9

(vi) To obtain the F2 generation, one student would withdraw one bead
from one beaker labelled male and one from the other beaker labelled
female keeping his/her eyes closed (to ensure randomness), and put
them together in the stretched palm of the partner, who will put them
together on the napkin spread over the table. Continue this process till
all the beads are paired. Thus 64 offsprings of F2 are obtained.
(vii) Note the genotype (YY or Yy or yy) of each pair, and their possible
phenotype.
(viii) Have six repeats of the experiment (steps i to vii) with partners changing
their roles. Pool all the data from the six repeats together.
(ix) Calculate the genotypic and phenotypic ratios of your pooled data.
Note that larger the sample size, more accurate is the result.

Observation
Record the result in the following table:

Generation Repeat No. Total no. of Genotype (s) Phenotype (s)

individuals YY Yy yy

F1 1.
2.
3.
4.
5.
6.
Total
F2 1.
2.
3.
4.
5.
6.
Total

Phenotypic Ratio: in F1…….

in F2…….
Genotypic Ratio: in F1…….
in F2…….

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 LABORATORY MANUAL: BIOLOGY

Discussion
The results are so because each diploid individual contains two copies of
every gene - one copy on each of the two homologous chromosomes. These
two copies of the gene may be of similar type (YY or yy) or are dissimilar Yy.
The former (YY or yy) are called homozygous for that particular character,
and the Yy are called heterozygous ones. The pure lines in the above cross
are homozygous ones, which contributed only one copy of their gene (as a
result of meiosis) to their F1 progeny to restore its diploid nature with genotype
Yy (heterozygous) where only one form (allele) is expressed (dominant) and
the other form (allele) is not expressed (recessive). This is the phenomenon
of Dominance.
When the F1 individuals are crossed together to raise the F2 generation,
each F1 individual produces two types of gametes: 50% having dominant
allele, and the remaining 50% having recessive allele. These gametes undergo
random fusion during fertilisation to produce the F2 generation. According
to simple probability of mixing of opposite sex gametes (sperms and ova),
offsprings of three genotypes are likely to appear as follows: [(half of gametes
of Y type + half of remaining gamete y type) X (half gametes of Y type + half
of remaining gamete of y type)] = One-fourth of F2 individuals of YY phenotype
+ half of F2 individual Yy type + one-fourth of F2 individul of yy type. Among
these proportion of dominant phenotype would be ¼ YY+ ½ Yy = ¾ yellow
and recessive phenotype ¼ yy i.e. green phenotypes in 3:1 or 75%:25%
ratio.
This ratio of 3:1 in the F2 suggests that in the F1 heterozygotes, the
recessive allele does not get destroyed and remains only in the recessive
(dormant) state to get an opportunity to express itself when it has separated
from the influence of the dominant allele (Y). This is called Law of Segregation
of the alleles.

Questions
1. Do you expect the same results in terms of 3:1 ratio in F2 if you had started with
smaller number of beads (say 10 beads)?

34
Exercise 10
Aim: To verify the Mendel’s Law of Independent Assortment

Principle: In a dihybrid cross, the segregation of one gene pair is independent of the segregation
of the other pair. It means that genes of two different traits assort independently to give a
probability ratio equal to segregration probability ratio of one allele pair X segregation probability
ratio of other allele pair, which comes to, (3:1) X (3:1) = [Link]

Requirement: Plastic beakers; 64 plastic beads each of yellow, green, red and white to represent,
yellow and green colour of seed coat and red and white flowers respectively and napkin/hand
towel

Procedure
Students are to work in pair.
The following steps are to be followed sequentially:
(i) Place 64 beads of each colour in four separate beakers.
(ii) Put the beakers containing the yellow and red beads on your left side,
and those containing the green and white beads on your right side.
The beakers on your left side represent plants bearing yellow seed and
red flower (dominant character YY, RR). Beakers on the right side
represent plants bearing green seeds and white flowers (recessive
character yy, rr). These are the two parental types having contrasting
forms of two different characters.
(iii) Stir the beads in each beaker with a pencil/pen. Each bead now
represents alleles in the male and female gametes.
(iv) Pick up one yellow, one green, one red and one white bead, and put
them together on the napkin spread on the table.
(v) Continue picking up and putting together of the beads of all colours
as mentioned in the previous step, till all the beads are utilised.
(vi) Note that in all, 64 such 4-bead clusters are obtained representing the
F1 individuals. Ascertain their genotype and phenotype.
(vii) Next step is to cross these F1 individuals to raise the F2 generation. Let
us suppose half of the 4-bead clusters (32 clusters) represent the male
parents and the remaining half (32 clusters) the female parents. Now
put the 32 red and 32 white beads together in one beaker (numbered-
I), and similarly put 32 yellow and 32 green beads together in other
 LABORATORY MANUAL: BIOLOGY

beaker (numbered-II). These two beakers represent F1 female. Similarly

put remaining 32 red + 32 white beads in beaker numbered-III, and
32 yellow and 32 green one in beaker numbered-IV to represent the F1
male. The arrangement can be presented as below

Female F1 Male F1

32 red + 32 white (Beaker I) 32 red+32 white (Beaker III)

32 yellow + 32 green (Beaker II) 32 yellow + 32 green (Beaker IV)

(viii) Stir the beads in each beaker with a pencil. In order to raise the F2
generation, pick up (with eyes closed) one bead from the beaker-I of
female and one bead from the beaker-III of the male, and put into the
palm of the partner student. Similarly, pick up one bead each from the
beaker-II of female and beaker IV of male to put in the palm of the
partner. This partner would now keep all the four beads together (to
represent the F2 individual). Continue this process till all beads are
utilised. At the end, 64 F2 individuals (each represented by a 4-bead
cluster) are obtained.
(ix) Determine the genotype and phenotype of each of the 64 F2 individuals
and write down the number of individuals of different genotypes and
phenotypes in the tabular form (given below), remembering that Y
(yellow seed colour) is dominant over y (green seed) and R (red flower)
is dominant over r (white flower).
(x) Repeat the whole procedure (steps i to ix) six times, and tabulate your
results.

Observation
Tabulate the results as follow:
Symbol (-) indicates the presence of corresponding dominant or recessive
allele e.g. Y or y and R or r.
Summarise your results (adding together the data of all the six repeats)
F1 Generation
(a) Total number of individuals: _________________________
(b) Phenotype (s) _________________________
(c) Genotype (s) _________________________

36
EXERCISE 10

Generation Total No. Genotype Phenotype

& repeat of offsprings Y-R- Y-rr yyR- yyrr Yellow Yellow Green Green
No. Red White Red white

F1
1.
2.
3.
4.
5.
6.
Total

F2
1.
2.
3.
4.
5.
6.
Total

F2 Generation
(a) Total number of individuals _________________________
(b) Phenotypes _________________________
(c) Number of individuals in each phenotypic class:
Number Phenotype
__________________ _____________________
__________________ _____________________
__________________ _____________________
__________________ _____________________
(d) Phenotypic ratio _____________________
(e) Genotypic ratio _____________________

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 LABORATORY MANUAL: BIOLOGY

(f) Number of individuals of each genotypic class:

Number Genotype
__________________ _____________________
__________________ _____________________
__________________ _____________________
__________________ _____________________
__________________ _____________________
(g) Genotypic Ratio ______________________________________

Discussion
The four phenotypic classes in the F2 generation are in ratio of [Link] as
expected from the Law of Independent Assortment. The genotypic ratio
would be ([Link]): (2:1):(2:1):1.
Note
1. In case six repeats of the experimental procedure are not feasible due
to time limitations, either the number of repeats be slashed down to
three or the data from single repeat of six different pair of students may
be pooled together to make the final calculations.
2. This Law of Independent Assortment was later found to be true only for
traits present on two different homologous pair of chromosomes, that
is, the two are not linked together. The linked traits do not assort
independently, rather they are inherited together (linked) except when
crossingover separates them.
3. It is quiet likely that you may not find your data exactly in the
expected ratio, instead almost approximate to it. The statistical
significance of this deviation from the exact expected ratio due to
probality can be checked using chi-square (χ2) test, about which
you will study in higher classes.

Questions
1. Linked traits fail to assort independently. Explain.
2. How is independent assortment of alleles important from the point of view of
variation?

38
Exercise 11
Aim: Preparation and analysis of Pedigree Charts

Principle: The Mendelian concept of dominance and segregation can also be studied in humans
by preparing and then analysing the pedigree charts. The internationally approved symbols for
indicating males and females, marriages, various generations (I, II, III), etc., are given below.

Requirement: Information about characters/traits in a family for more than one generation

Procedure
Select a family in which any one of the monogenic traits such as tongue
rolling, widow's peak, blood groups’, red-green colour blindness, dimple in
 LABORATORY MANUAL: BIOLOGY

the cheek, hypertrichosis of ear, hitch-hiker's thumb, etc., is found. Ask the
person exhibiting the trait to tell in which of his/her parents, grand parents
(both maternal and paternal), their children and grand children the trait in
question is present. Among surviving individuals the trait may also be
examined. The information made available is the basis for the preparation of
pedigree chart using the appropriate symbols. A careful examination of the
pedigree chart would suggest whether the gene for the character is autosome-
linked dominant or recessive, X - chromosome linked dominant or recessive,
Y- chromosome linked or not.

Explanation
1. Autosome Linked Dominant traits: These are the traits whose
encoding gene is present on any one of the autosomes, and the wild-
type allele is recessive to its mutant allele, i.e., the mutant allele is
dominant.
The pedigree-chart can be of the undernoted pattern (Fig. 11.2), where
the female being interviewed is exhibiting the trait, and is indicated by
an arrow-mark in the chart.

The characteristic features of inheritance of such type of traits are:

(a) Transmission of traits occurs from parents of either sex.
(b) Males and females are equally affected.
(c) The pedigree is vertical, i.e., the trait is marked to be present in each of
the generations.
(d) Multiple generations are characteristically affected.
Brachydactyly, polydactyly, dimple in the cheek are some of the common
traits of this type.

40
EXERCISE 11

2. Autosomal Recessive trait: These are the traits whose mutant allele is
recessive to its wild type allele.
The pedigree chart can be more or less of the pattern given below (Fig.
11.3), where the lady (marked by the arrow) is showing the trait. The bar

in the example represents the presence of corresponding dominant or

recessive allele for the specific trait.
Suppose the given trait is albinism. Denote its dominant allele as ‘A’
that produces pigments, and the recessive allele as ‘a’ that fails to synthesise
the pigment, melanin. The female (our subject in generation III) is therefore
of genotype aa. She must have received each of her ‘a’ allele from both the
parents (generation-II), who are therefore themselves normal but are definitely
of genotype Aa, and are carriers of the trait. The allele a must also have been
present in her grand parents too, of course in heterozygous condition also
to make them carriers (generation-I)
Albinism in the subject’s children (generation-IV) suggests her husband
too to be of genotype Aa, a carrier. Marriage of her albino daughter to an
albino man is bound to produce all her grand-children albino (gen-V).
The following are the salient features of the inheritance of such type of traits.
(a) Occur in equal proportions in multiple male and female siblings, whose
parents are normal but carriers;
(b) The siblings are homozygous for the defective allele, but their parents,
though some may appear normal, are obviously heterozygous, i.e.,
are merely carriers of the trait.
(c) Consanguinity (marriage between man and woman genetically related
to each other, such as cousins) occasionally results in the appearance
of such traits.

41
 LABORATORY MANUAL: BIOLOGY

3. X-Linked Dominant traits: These are the traits whose encoding gene
is present on the X- chromosome, and the mutant allele of which is
dominant over its wild-type allele.
Such traits are very rare, and are almost difficult to find in the
population. One example is oral-facial-digital syndrome (Duchene
Muscular Dystrophy), which results in absence of teeth, cleft (bifid) tongue
associated with mental retardation. The pedigree chart may appear as
follows (Fig. 11.4):

The possible genotypes of the above pedigree can be written as follows

(Fig 11.5):

Fig. 11.5 Genotypes of individuals shown in Fig. 11.4

42
EXERCISE 11

Here, the dominant mutant allele is denoted by ‘D’, and its recessive wild
type allele is denoted by ‘d’. Remember that human females have two
X-chromosomes (XX), and the males have only one X and one Y chromosome.
Males receive their lone X-chromosome from their mother, and the
Y-chromosomes from their father, whereas females receives one of her
X-chromosome from her mother, and the other X from her father.
The characteristics of such inheritance are:
(a) The trait appears in almost all the generations, and the inheritance is
vertical.
(b) If the female is affected, then about half of her sons are affected.
(c) If the male is affected then all of his daughters would be affected, but
none of his sons are affected.
(d) In short, the pedigree resembles the pattern of inheritance of autosomal
dominants, except that there is no male-to-male transmission.
4. X-linked Recessive traits: These are the traits whose encoding gene is
present on the X-chromosome and its mutant allele is recessive to its
wild-type allele.
Red-green colour blindness and hemophilia, are some of its well known
examples. The characteristic features of such inheritance are:
(a) Females express the trait only when they are homozygous for the
mutant allele, whereas the males do so even when they are hemizygous
for it.
The pedigree chart would appear as the following one (Fig. 11.6):

43
 LABORATORY MANUAL: BIOLOGY

(b) About half of the sons of the carrier (heterozygous for the trait) females
are affected. In case of homozygous females showing the trait, fifty
percent of her daughters and all of her sons are likely to be affected.
Therefore, the males are most affected in the population.
(c) Affected persons are related to one another through the maternal side
of their family.
(d) Any evidence of male-to-male transmission of the trait rules out the
X- linked inheritance.
5. Y-chromosome linked traits: These are the traits whose gene is present
on the Y-chromosome. The females do not have any Y-chromosome,
whereas all the males must have a Y-chromosome to be a male, and this
Y-chromosome they get from their father. Therefore, any trait linked to
the Y- chromosome must be present only in males, and certainly not in
any of the females. This is why these traits are also called male-sex limited
traits. All the sons of the affected male would express the trait whereas
none of his daughters would do so.
The pattern of the pedigree chart would be as follows (Fig 11.7):

Hypertrichosis of the ear (presence of hairs on pinna) is one most common

example of such traits.
Note: Students may be asked to prepare the pedigree-chart from given data and analyse the
pattern of inheritance. The work may be done as a project.

Questions
1. How will you differentiate between autosome linked dominant and sex chromosome
linked dominant pedigree chart? Explain.
2. Discuss the differences in the patterns of autosome linked recessive and sex-
chromosome linked pedigree.

44
Exercise 12
Aim: To perform emasculation, bagging and tagging for controlled pollination

Principle: Conventional plant breeding programmes involve bringing under human control
reproductive processes that lead to seed and fruit formation. For this controlled pollination is
desirable using male and female parent having desired traits. One of the process that can be easily
brought under human control is emasculation. For this the knowledge of flower structure,
mechanism of pollination, fertilisation and physiology of flowering is essential for this. In
emasculation technique the stamens are removed before anthesis to obtain female parent and pollen
from the desired male parent is transferred on to its stigma.

Requirement: Ornamental plants/ wild plants bearing large bisexual flower, magnifying lens,
tweezers, small sharp scissors, brush, alcohol, rubber bands, paper bags, paper clips and tags

Procedure
(i) Select a flower in bud condition where antheses has not occurred. Open
the bud carefully and remove the stamens (Fig. 12.1). Mark this as
female parent plant.

Fig. 12.1 Showing process of Emasculation

 LABORATORY MANUAL: BIOLOGY

(ii) Cover the emasculated flower with a plastic

bag to protect it from undesired pollen
(Bagging) (Fig. 12.2). The bag should be
held securely in place with a paper clip/
string/thread. Select the size of bag in
accordance with the flower size. Bags
must be transparent with minute pores.

Fig. 12.2 Bagging of an emasculated flower

(iii) Bring into physical contact anthers of a

desired male plant containing mature
pollen grains with the stigmatic surface
of emasculated female flower (Fig. 12.3).
Use tweezers/brush if necessary to dust
the stigmatic surface with pollen.

(iv) Cover the pollinated flower again with the

bag immediately. For identification, label
the female parent (Tagging). Each
pollinated flower should bear a label
Fig. 12.3 Showing cross pollination on an containing the name of the seed parent,
emasculated flower the letter X (to signify a cross), the name
of the pollen parent, and the date on which
the cross was effected.

Questions
1. Why is emasculation performed before anthesis?
2. What are the advantages of using a bag containing minute pores?

46
Exercise 14
Aim: To identify common disease-causing organisms and the symptoms of the diseases

Principle: There are quite a large number of organisms that are parasitic/pathogenic to humans.
These organisms substantialy damage the human body and cause diseases, which may even be fatal
sometimes. These organisms exhibit characteristic features in their external morphology. Symptoms
of the diseases caused by them are also specific.

Requirement: Preserved specimens/permanent slides/photographs of Ascaris, Entamoeba,

Plasmodium, Ring-worm fungus and compound microscope

Procedure
Observe the preserved specimens/slides/photographs and note down the
features in the practical record book. Take care to observe all the minute
details and draw labelled diagrams of the pathogens.

Observation

A. Entamoeba
Observe the following features of the parasite in the slide or photograph:
(i) It is unicellular.
(ii) Shape of the cell is irregular due to
pseudopodia.
(iii) A single nucleus is present eccentrically in
the cell.
(iv) *In the nucleus a peripheral ring of granule
of nucleoprotein and central karyosome are
observed. Rest of the space in the nucleus
looks empty (Fig. 14.1).
(v) A few food vacuoles may be seen in the
cytoplasm. Contractile vacuoles are absent.
(vi) *Mature quadrinucleated cysts may be
present.
Fig.14.1 An Entamoeba
 LABORATORY MANUAL: BIOLOGY

Note: Entamoeba is an intestinal parasite in humans and causes amoebic

dysentery. The symptoms of the disease are frequent loose, mucus filled
watery stools, abdominal pain and spasms.

Systematic position
Phylum – Protozoa
Class – Rhizopoda
Type – Entamoeba histolytica

* Distinctive feature of the pathogen

B. Plasmodium vivax
(i) It is an intracellular endoparasite seen easily within the RBC of the
infected person.
(ii) It is unicellular.
(iii) The most diagnostic stage of the parasite is "signet ring" stage in the
erythrocytes, within which it appears as a rounded
body (Fig. 14.2).
(iv) It has a big vacuole inside, and the cytoplasm is accumulated at one
place containing the nucleus. Because of the above mentioned features,
the parasite appears as a ring.
Search the stage in the blood film slide, find the signet-ring stage, and
draw its labeled diagram.
Note: It is a protozoan parasite causing malaria in humans. When an infected
female anopheles mosquito bites a healthy person, it injects the infective
stage, sporozoite, into the peripheral blood vessels. The infective stage
undergoes several rounds of multiplication in liver and erythrocytes.
Symptoms: Intermittent high fever with chills followed by profuse sweating
at an interval of alternate days.

Systematic position
Phylum – Protozoa
Class – Sporozoa
Type – Plasmodium vivax

50
EXERCISE 14

C. Ascaris
The external features of round worm are as follows: Mouth
(i) Body long (20 to 40 cm), cylindrical (5 to 6 mm
diameter) with no segmentation (Fig. 14.3).
(ii) Sexes are separate; the females are longer than
the males.
(iii) Both the ends are pointed; posterior end of male
is ventrally curved.
(iv) Mouth is situated at the anterior end, and is Female genital
surrounded by three lips, one present mid- aperture
dorsally and rest two lips are situated
ventrolaterally (for viewing these lips a magnifying
lens is needed).
Penial spicule
(v) Single longitudinal lines are present on the dorsal,
ventral and on the two lateral sides, all along the
length of the body. Out of these the lateral lines
are comparatively more distinct than the others
lines. (b)
(vi) Excretory pore is present on the ventral surface
slightly behind the anterior end.
(vii) In addition to the ventrally curved posterior tip, (a)
the male worm has a pair of penial spicules very Fig.14.3 Ascaris (a) Female (b) Male
close to the cloacal opening.
(viii) In case of female specimen a female genital
aperture is present mid-ventrally about one third
distance from the anterior end.

Systematic position
Phylum – Aschelminthes
Class – Nematoda
Type – Ascaris lumbricoides
Note: Round worm or Ascaris is one of the common parasite found in the
intestine of human beings.
Symptoms: (a) Irregular bowel, (b) Occasional vomiting, (c) Anaemia

51
 LABORATORY MANUAL: BIOLOGY

Trichophyton (Ringworm fungus)

It is a fungus that feeds on keratin of the skin of human beings. The features
as observed under the microscope are:
1. Texture of hyphae is waxy, glabrous to cotton like.
2. Unstained hyphae are white, yellowish brown to reddish brown in colour.

Systematic position
Kingdom – Fungi
Class – Deuteromycetes
Type – Trichophyton rubrum

Symptoms
Ringworm is a contagious fungal infection of the skin. Infected area of skin
is itchy, red, raised, scaly patches (with sharply defined edges). It is more
red on the periphery than in the center creating a ring like appearance.

52
Exercise 22
Aim: To determine the amount of Suspended Particulate Matter (SPM) in air at different sites
in a city

Principle: Environmental pollution is the unfavorable alteration of our surroundings wholly or

largely as a by-product of man's action through direct or indirect effects of changes in energy
patterns, radiation levels, chemical and physical constitutions of environment and abundance of
organisms. Substances that cause pollution to the environment are called pollutants. They are the
residues of things that man makes, uses and throws away. These residues pollute soil, water and air.
The atmosphere in highly populated area is very rich in dust, smoke and SPM all due to vehicular
exhausts and industrial emission.

Requirement: A few freshly cut broad leaves, Vaseline, laboratory balance, weights, brush,
paper clips and twine thread

Procedure
This experiment is an outdoor activity and may be
conducted by assigning 2–3 students into a group.
(i) Collect a few locally available broad leaves from a
nearby tree plant (Canna, Peepal, etc.).
(ii) Wash the leaves gently in running water to remove
any dust settled on their surfaces.
(iii) Blot dry the surface area of the leaves. To calculate
the area of the leaf, trace the outline of the leaf on
graph paper (Fig 22.1). Within the traced area
calculate the total number of full squares, 1/2, 1/3
and 2/3 squares and individual small squares. Add
all the squares to get the total leaf area. Multiply
their value with two to obtain total area of both the
surfaces.
(iv) Take 8–10 feet long twine thread and tie five leaves
leaving a foot distance in between. Apply an
extremely thin layer of vaseline on both surfaces of Fig. 22.1 Calculating the area of a
each leaf. Make a bundle of these leaves and pack leaf on a graph paper
 LABORATORY MANUAL: BIOLOGY

them in polythene bags. Ensure that the outer surface of polythene

bag does not have any vaseline sticking on it.
(v) Make three such bundles of smeared leaves, each bundle containing 5
leaves.
(vi) Mark bundles as A, B and C and carefully weigh each bundle of leaves
along with the polythene bags.
(vii) Select three spots (X, Y and Z) near by your school. Spots selected
should be in a manner that spot 'X' has very heavy vehicular traffic,
the spot Y has moderate traffic and spot 'Z' has little or no vehicular
traffic. At spot 'X' expose each leaf of bundle 'A' by stretching the
attached thread and tie the two ends to two poles or branches of trees
preferably at 10 feet height above ground. Keep leaves exposed for
about two hours.
(viii) After exposure at spot 'X', collect the leaves and carefully re-bundle
exposed leaves and place them along with the string in the polythene
cover 'A'.

Record your findings in the following table:

Site Leaf bundle Weight of leaves (g) Weight of Total leaf area
sample suspended (cm2) of five
particle (W2-W1) leaves
Before After
exposure (W1) exposure (W2)

X 'A'

Y 'B'

Z 'C'

(ix) Repeat the same process at spot 'Y' and 'Z' exposing leaves of 'B' and
'C' bundles respectively.
(x) At the end of the experiment, return back to the laboratory. Reweigh
each bundle of exposed leaves along with their respective polythene
cover.
- Calculate the amount of suspended particles deposited in mg cm2 of
leaf at each spot.
- Compare the results of three different spots and interpret.
Since the weight of suspended particles will be in milligrams or even less
it is advised to use a very sensitive laboratory balance.

84
Exercise 23
Aim: To study plant population density by quadrat method

Principle: Density represents the numerical strength of a certain plant species in the community
per unit area. The number of individuals of the species in any unit area is its density. The unit area
may be as small as 5 square cm to as large as 10 square metre depending on the size and nature of
the plant community under study. For herbaceous vegetation a metre square quadrat is normally
used. Density which gives an idea of degree of competition is calculated as follows.
Total number of individual(s) of the species in all the sampling unit (S)
Density=
Total number of sampling units studied (Q)
The value thus obtained is then expressed as number of individuals per unit area. When the
measured unit area is divided by the number of individuals the average area occupied by each
individual is obtained.

Requirement: Cotton/nylon thread (five meters), 4 nails and a hammer

Procedure
(i) In the selected site of study, make a 1 m X 1 m quadrat with the help of
nails and thread. Hammer the nails firmly and make sure that the
vegetation is not damaged while laying the quadrat.
(ii) List the names of the plant species seen in the quadrat (if the name is
not known mark these as species A or B etc., and the same species if
seen in other quadrats assign the same alphabet).
(iii) Count the number of individuals of each species present in the quadrat
and record the data as shown in the table.
(iv) Similarly make nine more quadrats randomly in the site of study and
record the names and number of individuals of each species.

Observations
Record the total number of species seen in the ten quadrats. This will give
an idea about the composition of the vegetation.
There will be difference in the species composition in the quadrats made in
shady areas, exposed areas with bright sunlight, dry or wet areas etc.
 LABORATORY MANUAL: BIOLOGY

Table 23.1: Density studies of the given vegetation

Plant Quadrats employed in study & no. of Total No. of Total no. Density (D)
Species individuals in each quadrat individuals (S) of Quadrats
studied (Q)
I II III IV V VI VII VIII IX X

A 2 5 7 10 3 27 10 27/10 = 2.7

Z 1 2 4 8 3 2 20 10 20/10 = 2.0

Discussion
Plants growing together exhibit mutual relationships among themselves and
also with the environment. Such a group of plants in an area represent a
community. The number of individuals of a species varies from place to
place, making it necessary to take many random sample areas for reliable
results. Density values are significant because they show relative importance
of each species. With increasing density the competition stress increases
and the same is reflected in poor growth and lower reproductive capacity of
the species. Data on population density are often very essential in measuring
the effects of reseeding, burning, spraying and successional changes.
Discuss the vegetation composition of the area (herbs/shrubs) and
comment on the dominant component species.

Questions
1. What factors influence the population density?
2. What is the significance of quadrat method?
3. What conclusion can be drawn if density of a plant species is low?

86
Exercise 24
Aim: To study plant population frequency by quadrat method

Principle: Frequency is concerned with the degree of uniformity of the occurrence of individuals
of a species within a plant community. It is measured by noting the presence of a species in random
sample areas (quadrats) which are distributed as widely as possible throughout the area of study.
Frequency is the number of sampling units (as %) in which a particular species (A) occurs. The
frequency of each species (sps. A or sps. B or sps. X etc) is expressed in percentage and is calculated
as follows.
% Frequency or Number of sampling units (quadrats) in which the species occurs
=
Frequency Index Total number of sampling units (quadrats) employed for the study

Requirements: Cotton/nylon thread of 5 metres, 4 nails and a hammer

Procedure
(i) In the selected site of study, make a 1 m X 1 m quadrat with the help of
nails and thread. Hammer the nails firmly and make sure that the
vegetation is not damaged while laying the quadrat.
(ii) List the names of the plant species seen in the quadrat (if the name is not
known mark these as species A or B etc. and if the same species is seen in
other quadrats assign the same alphabet)
(iii) Similarly lay nine more quadrats randomly in the site of study and record
the names of individuals of each species.
(iv) Calculate the percentage frequency of occurrence using the formula given.

Observations
Record the total number of species seen in the ten quadrats. This will give an
idea about the composition of the vegetation.
There will be difference in the species composition in the quadrats made in
shady areas, exposed areas with bright sunlight, dry or wet areas etc.
Observe that the frequency of occurrence is not the same for all species.
 LABORATORY MANUAL: BIOLOGY

Table 24.1: Frequency studies for the given vegetation

Plant Number of quadrats employed No. of quadrats Percentage

Species in the study (Q) in which the of frequency
species is present (N) F=N/Q X 100
I II III IV V VI VII VIII IX X

A √ √ √ √ √ 5 5/10 × 100 = 50%

B √ 1 1/10 × 100 = 10%

C √ √ √ √ 4 4/10 × 100 = 40%

Discussion
Variation in distribution of a species is caused by factors like soil conditions,
quantity and dispersal of gemmules, vegetative propagation, grazing, predation,
diseases and other biotic activities. Also frequency values differ in different
communities. They are influenced by micro-habitat conditions, topography,
soil and many other environmental characteristics. Thus unless frequency is
not correlated with other characters such as density, frequency alone does not
give correct idea of the distribution of a species.
Frequency determinations by means of sample areas are often needed in
order to check general impressions about the relative values of species. Many
species having low cover or population density also rate low in frequency, but
some may have high frequency because of their uniform distribution. Usually
if the cover and population density are high, the frequency will be high. The
plants with high frequency are wide in distribution.

Questions
1. If frequency of a plant is high, what will be your interpretation?
2. Can many micro-habitat in an area affect frequency of a species? Comment.

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Exercise 25
Aim: Study of homologous and analogous organs in plants and animals

Principle: In plants and animals there are several organs or parts thereof, apparently alike in
their function and appearance, but markedly different from each other in their origin and
anatomical structure. These organs are called analogous organs, and the seeming similarity
among them is the result of convergence, that is, adaptation to similar habitat and identical
ecological niche.
On the other hand, there are organs or parts thereof, which apparently are quite dissimilar
to each other in appearance and perform different functions, but have the same origin and
anatomy. The differences in their function and also in their appearances are the result of
divergence, due to adaptive radiation to different habit, habitat and ecological niche. These
organs are called homologous organs.

Requirement: Plant specimens showing tendrils, thorns, etc., as given in the text or any other
locally available plants, a plant with normal stem, potato and onion bulb, prickly pear, specimens
of phylloclade, cladode, wings of bird, cockroach and bat, and cervical, thoracic and lumbar
vertebrae of a mammal/lizard

Observations

1. Homologous Organs in Plants

(i) Tendrils of passion flower and thorns of pomegranate

Tendrils of passion
fruit and thorns of
pomegranate are Tendril
structurally and
functionally different
but they have similar
origin i.e. they arise
from axillary bud Thorn
(Fig. 25.1a & b).
(a) (b)

Fig. 25.1 (a) Tendrils of passion fruit (b) Thorns of pomegranate

 LABORATORY MANUAL: BIOLOGY

Tendril Thorns (ii) Tendrils of Vitis and thorns of

Carissa
Tendrils of Vitis and thorns of
Carissa originate from the terminal
bud, but they are functionally
different (Fig. 25.2 a & b).

(a) (b)
Fig. 25.2 (a) Tendrils of Vitis (b) Thorns of Carissa
(iii) Tendrils of baloon vine
Tendril (Cardiospermum) and bulbils of
Agave.
Both are modifications of floral bud,
but they perform different functions.
Tendrils help in climbing but bulbils
are meant for reproduction
(Fig. 25.3 a & b).
(a) (b)
Fig. 25.3 (a) Tendrils of baloon vine (b) Bulbils of Agave
(iv) Scale leaves of onion and spines
of prickly pear (Opuntia)
Both the scale leaves and spines are
Spines modifications of leaves but are
structurally and functionally
different. Scale leaves of onion are
thick and fleshy and store food. On
(a) (b)
the other hand spines of cactus are
Fig. 25.4 (a) Scale leaves of onion (b) Spines of cactus defensive organs (Fig. 25.4 a & b).

2. Analogous Organs in Plants

Tendril
(i) Stem tendrils and leaf tendrils
All tendrils are analogous with one
another, being structurally and
functionally similar, irrespective of
their origin.
Example: Tendrils of pea and
tendrils of Vitis. Tendrils of pea are
Tendril modification of leaf and in Vitis it is
(a) (b) the modification of terminal bud
Fig. 25.5 (a) Tendrils of pea (b) Tendrils of Vitis (Fig. 25.5 a & b).

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EXERCISE 25

(ii) Thorns and spines

Thorns and spines are analogous structures being
defensive in function. Thorns are modifications of
axillary or terminal buds, and spines are (a) (b)
modifications of leaves. Fig. 25.6 (a) Modified root of carrot
e.g: Thorns of pomegranate and spines of (b) Rhizome of ginger
prickly pear.
(iii) Modified underground stems and modified roots Spine
Modified stems (rhizome, corm, tuber) are
analogous to modified roots (carrot, radish) as they
perform similar function of storage of food but their
origin is different. Rhizome of ginger, potato tuber,
(a) (b)
Colocasia are stems and beetroot, radish etc. are
Fig. 25.7 (a) Phylloclade (b) Cladode of
roots. (Fig. 25.6 a & b)
ruscus
(iv) Phylloclade, cladode and leaves
They perform the same function i.e. they
photosynthesise but phylloclade and cladode are
modifications of stem. Phylloclade of Opuntia,
Parkinsonia, Asparagus and leaves of any local
plant like mango are analogous organs.
(Fig. 25.7 a & b) (a) (b)
3. Homologous Organs in Animals Fig. 25.8 Fore limb of (a) human (b) bat

(i) Wings of birds, and forelimb of mammals/reptiles/

frog: All have the same bony elements (humerus radio-
ulna, carpals, metacarpals and phalanges), but
perform different (flying in birds, for holding or walking
etc. in other) functions. (Fig. 25.8 a & b) (a) (b)
4. Analogous Organs in Animals Fig. 25.9 Wing of (a) dragonfly (b) bird
(i) Wings of dragonfly/cockroach/butterfly and of
birds. (Fig. 25.9 a & b)
(ii) Mandible of cockroach and mandible (lower jaw) of
a vertebrate. (Fig. 25.10 c & d)
(a) (b)
Note: Students and teachers are suggested to discuss
Fig. 25.10 Mandible of (a) cockroach
more examples.
(b) rabbit

Questions
1. Suggest examples of homologous and analogous organs other than what are
given in the manual.
2. Why are stem and leaf tendrils considered as analogous organs?

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