FLOWERING PLANT REPRODUCTION
This week you'll study the flower structure and reproductive biology of flowering plants with the
goal of understanding more about sexual reproduction and its place in evolution. Bring your
textbook to lab.
REPRODUCTIVE BIOLOGY OF FLOWERING PLANTS
Most of the cells in a flowering plant, including most cells of the flower, are diploid. The diploid
body of cells making up shoots and roots is called a sporophyte, because it is the bearer of the sporemaking tissues. The spores, which are haploid, are the result of meiotic divisions that occur in just
two places, the stamen and the pistil of the flower. These haploid spores undergo two to four
additional mitotic divisions to yield small haploid organisms called gametophytes. It is these small
gametophytes that yield the gametes (egg and sperm) that are involved in syngamy.
Two critical events in angiosperm (flowering-plant) reproduction are pollination (the movement of
pollen to the stigma), and dispersal of seeds. The structures that you will encounter in your
observations and experiments all relate to one of these two functions.
START THE POLLEN IN SECTION E FIRST
A.
Basic Flower Structure - Alstroemeria
1.
Select a flower of Alstroemeria to dissect. Alstroemeria is a scraggly climbing plant
that grows near treeline in the southern Andes, in Bolivia and Peru. Do your
dissection under a dissecting microscope. Have two probes at hand.
2.
In theory the flower is a kind of shoot -- that is a stem with leaves attached. However
the leaves are highly specialized and the internodes are so short that all of the flower
parts appear to be attached to a single point on the stem, called the receptacle.
3.
Begin by looking at the two whorls (groups) of colorful structures – together called
the perianth:
a.
b.
c.
d.
The outer whorl, the sepals, is slightly different from the inner whorl, and it
encloses the whole flower when it is a bud (unopened).
The inner whorl, the petals, has a complex pattern of color and marking on
the inner surface for guiding the pollinator (ordinarily bumblebees in the
Andes).
Decide on the symmetry of the flower. Looking straight at the top of the
flower, decide whether the symmetry is radial, like a snowflake, or bilateral,
like a face.
Check out the bottoms of the sepals and petals. Are they separate from each
other, or are they partially combined together to form a tube, what we call
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fused?
4.
Now look at the stamens, which are the pollen producing organs. They consist of
two parts, an anther – the actual location of meiosis – and a filament – the support
and display axis for the anther.
a.
b.
c.
d.
5.
Count them and record your count.
Decide if the filaments are separate or partially fused to each other or to the
perianth.
Look closely at the anther under your dissecting ‘scope. You should be able
to break open the anther with a probe and see the pale yellow pollen grains
inside.
Remove some of these and mount them on a slide. Under the compound
‘scope, look for shape and markings on the pollen.
Locate the pistil, in the center of the flower. The pistil has three parts.
a.
b.
c.
At the top is the stigma with three sticky lobes. Look for pollen: some may
have already been dumped from the stamens onto the stigma -- this is the
actual event of pollination (the arrival of pollen at the stigma.)
Beneath each stigma is a stem-like display axis called the style.
At the base of the style, is a swollen part of the pistil called the ovary. Inside
this portion of the pistil is the other place where meiosis takes places in
flowers and where seed development takes place.
Tear open the ovary with your probe to expose a group of small, spherical whitish
green things inside. These are the ovules -- each one the site for a single meiosis.
Syngamy itself takes place inside of these ovules -- and the ovules develop into the
seeds that have the potential to establish a new generation of diploid sporophyte
plants.
B.
Basic flower structure and variation – Petunia
Now study a second flower to start to get a grip on the kinds of features that vary in flowers.
1.
Select a flower of Petunia to dissect. Petunia is also from the Andes, but from
lower in the mountains, and not so far south. Again, do your dissection under a
dissecting ‘scope using the probes provided.
2.
This time the sepals are very distinctive since they are green. Record the number,
fusion, and symmetry.
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3.
The trumpet-like thing that is the most prominent part of the flower is thought to
be a set of petals. What about the structure might lead you to think that? Count
the number of lobes at the edge of the trumpet’s bell, and record this as the
number of petals. This group of petals, called a corolla, is the animal-attracting
device of the flower.
4.
Now use a probe to open a slit in the side of the corolla so that you can see the
inner parts of the flower. Figure out how many stamens there are, and decide if
they are fused to each other, and/or fused to the corolla. Once again remove
pollen, mount on a slide, and study under the compound ‘scope. How are the
shape and surface markings different from Alstroemeria?
5.
Locate the pistil, which looks like a fifth stamen, except that it extends all the way
to the bottom of the petal tube. Once again, distinguish stigma, style, and ovary.
Only the summit of the style gives a hint that this is actually a pistil made of two
parts fused together. Look for a slight groove in the summit of the stigma: it
divides two parts of the stigma and provides evidence of this fusion.
There is better evidence inside the ovary. If you make a cross section of the ovary with a razor
blade, you will see that there are two different chambers in which the ovules are located.
Now fill in the chart below with your data for the two kinds of flowers:
Flower Name
Sepal number,
fusion, and
symmetry
Petal number, fusion,
and symmetry
Stamen number,
and fusion
Pistil number,
and number of
parts
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C.
Microscopic Structure of the Developing Anther and Pollen of Lilium (Lily)
Inside the anther cells undergo meiosis -- as you saw last week -- to yield haploid microspores.
These microspores divide to yield a two-celled male gametophyte better known as a pollen grain. It
is these pollen grains -- immature male gametophytes -- that will be dispersed by various means to
the stigmas on pistils. Study the fine structure of the anther during the events of pollen production
by looking at slides #1 to #4 in the Lilium development series.
1.
Look at slide #1 under low power to orient yourself to a cross (transverse) section of
a lily flower. Identify the four basic flower parts from outside in -- three sepals, three
petals (each with a thick, square keel in the middle), six stamens (each with two pairs
of lobes), and one ovary (with three ovule-bearing chambers called locules).
2.
Look at slide #2 under medium power. Locate a single anther in cross section. You
should find that it has two pairs of chambers, also called locules. Each of these
locules is the space contained in a single microsporangium - that is the chamber of a
container in which meiosis yields microspores.
There are two kinds of cells in each locule, tapetum cells and microsporocytes. The
tapetum cells are recognizable because they are a set of rectangular cells that lie in a
circle around the outside of the actual space in the locule like the stones around an
old-fashioned well. These tapetum cells have an important role in manufacturing the
coat of the pollen grain.
The microsporocytes are the cells that are designated to undergo meiosis by the plant.
They are the spherical cells that lie in the locule itself, inside the ring of rectangular
tapetum cells. Many of these sporocytes are in prophase of meiosis I: you can see the
condensing strands of the chromosomes in slide #2.
3.
Orient slide #3 under low power so that you can look at a single anther locule. Under
high power, you should be able to see groups of two, three, and four cells scattered
across the locule: these are microspores -- the products of meiosis. There are, in
truth, four cells in all the groups, but the preparation does not always make it possible
to see all four. These cells in fours are called the microspores.
4.
Look at the pollen grains in slide #4 under high power. Note two differences
between pollen and microspore. First, the pollen has two nuclei, one dark and one
light. Second, the pollen grain has a thick, durable ornamented coat outside its cell
wall (produced by the cells of the tapetum, which as you can see have been used up).
The two nuclei are actually in different cells, as we'll discuss in lecture. The
elongate, darkly stained nucleus called the generative nucleus will undergo mitosis to
yield two male gametes. Each will function during double fertilization.
The other nucleus, which is round and only lightly stained, is called the tube nucleus.
It mediates the germination and growth of the pollen grain once it reaches the
stigma.
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D.
The Pollination of Flowers
The visible characteristics of flowers -- especially color and shape of parts -- have been demonstrated
over and over again by botanists and zoologists to be correlated with designs for the movement of
pollen from anther to stigma. The pollen of colorful flowers is moved by animals: observations and
inferences about the movement of pollen by animals are among the most interesting in biology. The
best part is that absolutely no prior knowledge of biology is needed to make these sorts of inferences,
so that you can get kids in a summer camp or corporate executives to make good scientific inferences
about pollination biology without any knowledge of biology. Here, you can make some of your own
inferences about the pollination of greenhouse flowers.
There are two major principles to pollination biology; they both relate to the adaptation of flowers to
their animal pollen carriers (called pollen vectors). These principles are technically theories in that
they are hypotheses that have stood numerous tests and explain fundamental patterns seen in living
things.
I.
Flowers are adapted to waste as little pollen as possible: evolutionarily advanced
flowers often waste much less pollen than evolutionarily primitive flowers.
II.
Flowers are designed to promote the movement of pollen from one flower to another
and usually from one individual plant to another, that is, they promote outcrossing.
(Why outcrossing is desirable is one of the most fundamental questions in evolutionary
biology and one that is being argued vigorously by specialists today).
Look at the Fuchsia and Columnea in your lab. Answer the following three questions.
1.
Observers in nature have seen hummingbirds visiting these flowers. How is each
adapted to the visits of hummingbirds?
2.
Where does pollen get deposited on the hummingbird? What features of the flowers
reduce the amount of pollen wasted?
3.
Are there any specialized features that prevent pollen from getting deposited directly
on the stigma of the same flower?
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E.
Germination and Growth of Pollen
(Start at beginning of lab)
In section B you look at meiosis and the development of pollen grains up to the point where they are
shed from the anther, and in section C you study the ways in which pollen is moved from the opened
anther to the stigma. In this section, you conduct a simple growth experiment to demonstrate the
growth of the pollen grain (immature male gametophyte) once it has reached the stigma, since pollen
grains will germinate on agar supplemented only with sugar. The rest of the nutrients needed for
growth are stored inside of the pollen grain.
1.
Your teaching fellow will put a drop of sugar agar on a clean slide for you.
2.
Wait a minute for the agar to cool and solidify. (Hot agar can cook the pollen -- and
you get no results from cooked pollen.)
3.
Flowers with pollen will be in covered dishes in the lab. Lightly tap the stamens of
these flowers over your sugar agar. Check with a microscope to be sure pollen is in
your agar. Mark your slide with your initials and the time using a wax pencil or
marker. Finally, put the slide in one of the large covered glass dishes in the lab to
keep the agar culture moist. Use a probe to dislodge pollen if it doesn't fall out when
you tap.
4.
After 15 minutes place one drop of 16% sucrose solution on top of your agar with
pollen. Return the slide to the covered dish and check it once in a while for pollen
tubes. Once you can see good pollen-tube development under the microscope (say a
pollen tube ten times as long as wide) add a small drop of toluidine blue stain and
then a coverslip to the culture. In another ten minutes, study the pollen tube closely
to try to locate the nuclei in the tube.
In real life, the pollen tube grows down a canal in the style and into the ovary, with the egg inside the
ovule as its goal. Many pollen grains can be growing down the canal at once, and there is a
competition among pollen grains that appears to be controlled by the pistil. In other words, the
female parent can choose among male candidates in the style!
5.
In slide #10 of the Lilium development slides, you can see the style of a pistil covered
with pollen. Some of the pollen has germinated and begun to grow, just as the pollen
in your sugar agar did.
6.
After one hour, count the number of pollen grains, and the number of germinated
pollen grains on each slide. (Count approximately 30 grains on each slide). Evaluate
your data to determine if there were any differences in germination between the two
treatments.
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*** The following chemicals have been shown to affect pollen tube germination. All also contain
10% sucrose.
A.
Auxin – can increase pollen tube germination at low concentrations (1x10-6 M) but can
be inhibitory at high concentrations (1x10-4M). Involved in cell (acid) growth pathway.
B.
Boric Acid – has been shown to increase germination. Involved in cell wall synthesis.
C.
Cobalt (as CoCl) – inhibits germination. Blocks calcium channels.
D.
Gibberellic Acid – can increase germination at most concentrations.
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