Plant growth regulators (auxins, cytokinins, gibberellins, and abscisic acid) are crucial for plant development, differentiation, and organogenesis, particularly in tissue culture. Solidifying agents like agar are also essential for creating a stable medium for plant growth.
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plant growth regulators. We are telling
that there are four important
group of chemicals that is your oxen,
cytoinine, gibberlins
and abicesic acid. So these are our
growth regulators. And why is these
growth regulators necessary during a
tissue culture is only the growth
regulators can bring about the
differentiation and organogenesis of the tissue.
tissue.
If my tissues have to grow or if my cell
has to grow, it can be brought about
only by the growth regulators. That is
why we add two or three growth
regulators which are necessary
in the media and also remember the ratio
of hormones required for root or shoot
induction varies considerably.
So this is why we require the plant
growth regulators. We shall define what
are plant growth regulators. Plant
growth regulators are compounds which
are required in low concentration. [Music]
Okay, these are required in low concentration
concentration
and they capable of modifying the plant growth.
growth.
Okay. Either growth or even the
So what do you mean by morphogenesis? It
is the development of the uh tissues or organs
organs
into a particular shape that is called
as the morphogenesis. So my plant growth
regulators are required in small
quantity though they required in small
quantity they are very essential because
they are important for the growth and
morphogenesis of my plant. So usually
into a media we require at least two to
three growth regulators we add and also
remember the ratio what we add is also
important. what is the ratio of the
growth regulators we are adding that
also is a important.
So just a small
uh differentiation we actually we
interchange plant hormone plant growth
regulators. Even throughout my class
also I'll be using these two
interchanging and using but actually
plant hormones are naturally occurring
in the plant. Plant growth regulators
means they are both naturally occurring
and synthetic.
That is when we do it, we start
synthesizing in the laboratory that is
called as synthetic and that actually
refers to the plant growth regulators
but usually we keep interchanging it.
All plant hormones are growth
regulators. So all your plant hormones
can be your growth regulators but not
all growth regulators are your plant hormones.
hormones.
Why? because I can do different type I
can synthesize a diff different variety
of compound which is called as growth
regulators which may not be available
naturally. So remember all plant
hormones are growth regulators but all
growth regulators are not plant
hormones. So in my class also I just
Now we shall go to our first growth
regulator that is oxins.
So we have already studied that vent
isolated oxen from the tips of the
colopet of the oat leaves. Colopelis
means the tip of the region. If this is
the plant okay this region if this is a
plant the tip of this region okay it is
called as the colocopin. So from there
it was isolated.
So where are these oxins produced? Or
oxins are the natural hormones of the
plant. So where are they produced? They
are produced in the epesses of the stem
that is in the tip of the stem and even
in the roots also they are produced. So
from there wherever they are produced
okay in the tip of your roots or in the
axis wherever they produced they migrate
to the region wherever they are
and oxins. No, it is not soluble in
water. You have to dissolve it in
So what is the main function of our
oxen? The main function of our oxen is elongation.
elongation.
And we also came across a word known as
indoleacetic acid. This is one of the
common oxins which are used.
So as we told that it is present in the
shoot tips and even in the root tips and
they are transported to all other region.
So if you look at the function
or let me go into this. If you look at
the types of oxins, there are natural
oxins and also there are synthetic
oxins. Natural means they are present
naturally in the plant. Synthetic oxins
means it is man-made oxins.
So the most common is the indolacitic
acid, indolutric acid. So they are
isolated from the plants because they
present naturally. But synthetic is what
man has created that is you have
napylene acetic acid and 24D.
This 24D is very important that is it is
24 dchloropoxy acetic acid which has
been asked for two marks question. So
these are what they are they can be used
as herbicide.
24D is used to killot weeds.
Weeds means we don't require that plant
unnecessary growth of certain plants.
These are called as wheat. So weeds. So
we can use this 24D which is an
synthetic form of oxen to kill the
diacot weeds but it is not effective
against the monocot plant. Most of our
monocott plants are what are crop plants.
So the function is say that it
establishes the epical basal polarity of
the seed embryo.
We know that whenever there is the seed
it has a polarity to develop into a shootute
shootute
and then
it is bipolar
induces vascular tissue differentiation.
It mediates photorophism. Very
important. Promotes the formation of
adventicious roots.
This is also important. Stimulates fruit
development and it is used for cloning
in plant tissue culture. So remember
whenever we are growing a plant in a
tissue culture. Okay, remember this is
your expplant and you are growing the
plant in the
some media. So what happens if I want
this plants to produce the root?
I have to use what? I have to use the oxins.
oxins.
Okay. So oxins are used for the
production of the root for your tissue
culture plant and also it stimulates the
fruit development. We can see here some
A common feature of oxen is the property
even it can induce cell division also.
So in nature hormones of this group are
involved in activity like elongation of
the stem.
It is also it elongates the inter nodes.
It brings about photorophism as we have
mentioned here epical dominancy
and then it brings about the uh rooting
in the plant. So these are the important
functions. Here you can see that when
the oxins were added this type of
advantageous roots have been produced in
the plant. Similarly we can see that
when oxins were added to strawberry the
strawberry size increased and it has
photorophism. Photorophism means you
know that it is towards attraction
towards the light. So here what happens?
See this is your epical region of your
plant. Okay. If this is the plant this
region what happens whenever the light
is there it bends towards that light.
That is what we have drawn it like this.
So what is there in these epical regions
as we told in the epical regions we have
the indole acidic acid. Indole acetic
acid is nothing but your oxen. Okay. So
now when there is light now this all our
cells. Okay. All the cells will be
having the indolacitic acid. When the
light falls on this epical region, all
the indolacitic acid they'll move to one
direction. All in the opposite direction
of the light. So here less amount of
indolacitic acid there all have got transgent
of that. Why? Because what is happening
here? more oxins are there and it
inducing the growth in this region. So
there is a multiplication of your cells
in this region. So automatically what
happens the other region which is having
less oxen will bend. This is the
phenomena photoism. So this is the
normal growth which has bent here but
there is excessive growth occurring.
That is why your plant bends like this
towards the light
and also like epical dominance we mentioned.
mentioned.
So it is not clearly visible.
Okay. But this is the epical bud and
this is your lateral buds here. What
you're seeing now this is all your
lateral buds and this is your epical
buds. Now scientists did their experiment.
experiment.
What they did here you see here the
epical bud is removed and it is replaced
with the gelatin block containing indole
acetic acid. They remove this epical bud
and on this what they place they place
an gelatin. What is this gelatin
containing? The gelatin is containing
So what happened? They observed that
only this epical region started growing.
The plant started growing like this.
There was no lateral growth. No lateral
growth. It is all absent. Lateral
growths are absent. In the next
experiment, what did they do is the
epical bud was removed and replaced with
again gelatin block. Okay, I placed the
gelatin block. Now this time
the block is not having any indolacitic acid.
Now what happened? The plant did not
grow like this straight. It not grow.
Instead of that you see here lateral
So if I want a bushy tree or plant so I
can cut off this region so that always
your lateral stems can grow. So this is
known as the epical dominance of the uh
So remember the main important point is
it is inducing the root formation in
your tissue culture plant. That is what
you have to remember most important point
point
apart from these whatever I mentioned.
actually uh
uh
I'll tell you about this later on. There
is a story about this but we'll run
short of time.
Next coming to gibberlins.
These gibberlins are also known as
gibilic acid or simply they are
mentioned as GA and they are also
produced in your epical buds in the
roots young leaf and seeds of your embryo.
embryo.
Now this gibbrilin have shown to
stimulate blotting which is a rapid stem
I'll explain to you what this uh stem
elongation and pla in initiation
blotting all means. So,
so how was this gibberlin identified?
First it goes back to very way back in
something 1900s. What happened was in Japan
Japan
in rice plants a disease called as
backne disease or foolish seedling
caused by a fungus gibbrrella fugicuria
resulted in plants that grew very tall.
So now what happened? I have my patty plant.
plant.
Okay, rice plant. Now it is growing but
along in the soil what is there? There
is a fungus and this fungus
it is inducing a chemical okay towards
this plant and this rice plant what
and all the patties were here. Now what
happened? It is you know how the patty
plants are okay so lean they are. And
now what happens? They started drooping
because of the weight of the patty
grains. And whenever there was breeze or
heavy wind, what happened? Because they
are very slender, they started to fall off.
off.
Now they were wondering what is happening.
happening.
Then they came to a conclusion this is
because of this fungus. Okay, this
fungus gabriela fugia which is actually
producing the chemical which is being
passed on to your plant and the plant is
increasing in the size. So later on here
you can see gibberlic acid made by the
fungus cause the rice plant to overgrow.
Then later on it was found that plants
also make gibberlic acid and that apply
it to the
to plants cost growth. So what it means
is okay they found that this fungus is
also producing ads but also later on
they came to know that our plants also
by naturally they are producing this
gibberlic acid. So there are at least
more than 100 of uh this gibberelic
acids which are present from the fungi
and higher plants which it is known. So
you can see here this plant it is very
do it is a bonus that is here what has
happened they have removed the GA
so it is short
this plant because it is having a very
reduced amount of gibberlin and that is
why it is a short plant is a boness
plant so I can treat even my grapes this
is my normal grapes. But what the
scientists have done they treated this normal
normal
grapes with the gibberlin and they saw
that these grapes became large in size
and they elongated
and most of you have seen such type of
grapes which are long grapes and
elongated in size. So what happened was
like it is agricultural boon
and one more you have to remember in
gibleins there are no synthetic forms
like in oxen we don't know man-made ones
artificial ones oxins
like lapaline acetic acid 24D and all
but here such type no synthetic form of
this hormone is present
And of all the hormones, okay, of all
the gibbrilin hormone, G83 is the most
commonly used in the plant tissue
culture. So what is its function? Stem
growth and elongation.
Then blotting and flowering.
one second. You can see here this is the
untreated mutant plant maintained as a
dove. This is a very small plant that is
what is called as drop. But see this
plant 22 days after being sprayed with
dilute gibberlin solution. This plant
reached the size of now it is quite tall.
tall.
It is no more dark. That is a plant
which was dropped. What did the
scientists do? They added
the gibberlin to this and then 22 days
after being sprayed they saw that this
dolphin was lost and the plant reached
and it is also induced in seed
germination, dormcy, sex expression,
fruit growth, partinoarpy. These are all
the important functions of the gibberlins
and one more functions you can see that
this plant no it is rosette
this shape. So when you induced
gibberelin what happened? it started
growing well spread tall it started
flowering and this is known as blotting
what I mentioned here okay blotting and
flowering if your plant is in the roset
form it's not flowering you induce
gibberlin that is your g3 which is
commonly used okay you get this
beautiful plant which is grown tall and
it just also started flowering so you
can with these two hormones hormones you
can come to know that how important the
hormone is. Without this hormone there
is the plant development the organo
genesis will not occur at all. And we
saw that um the father of tissue culture
hammerland's experiments how he it
failed and how he told that there is
some enzymes which is required for the
growth of the plant and today we know
that what are these what he meant by
enzyme he meant it is a hormone
the very easily you can remember
cytokinus. Cyto means cell kinance means
division. So cell division the main
function of your cytoanine is the cell division
division [Music]
and the cytokenine was first discovered has
has
kinet by shook and his co-workers and
this naturally occurs in the plant.
So this cytoinine
is an adinine derivative which are
mainly concerned with the cell division.
One second.
A d e n i addin 9.
So your cytokinine is derived from this
adinine and its main function is
cell division and it can also bring
about modification in the epical
dominancy and shoot differentiation. So
you have to remember this ratio whenever uh
uh
actually this oxen is always used for
your root formation.
Okay, this part you have to
remember and your cytokinine is always
So there are again different type of
cytokin okay like this most commonly
what is used is bat six benzile
aminopurine and even the cytokenine are
not soluble in water get dissolved in
HCL or NAH. So the cytokinine in plants
promote the cell division by stimulating
the production of proteins needed for mitosis
mitosis
and also they induce the callus
So some of the commonly naturally
available cytokinine is there zazatin.
It is a natural substance with cytoinine activity.
activity.
There are some synthetic compounds with
cell division promoting activity
with synthetic means it is man-made and
natural cytoines are synthesized
in regions of rapid cell division. Okay.
Wherein the root epacases, shoot bugs,
young roots and etc.
Cytokinins are produced in actively
growing tissues. That is what here also
your roots, embryos and fruits.
Cytokinins work together with oxen to
control the cell division and
differentiation. That is what I
mentioned in your previous slide.
Cytokinins slow the aging of some plants
by inhibiting the protein,
stimulating RNA and protein synthesizing
and mobilizing the nutrients from the
surrounding tissue. These are some of
the important function of the cytoinets.
But the first point what you have to
remember is that it is used for cell
division and it is also used for callus development.
This point you have to remember girls
once see this one. The ratio of oxins
and cytokenins is always important with
respect to the morphogenesis of the plant.
plant.
Whenever there is the cells and tissues
when they are undergoing division and
they're forming a particular shape of
your plants that I told you it is
morphogenesis. So for a morphogenesis of
a plant it becomes very important who
and also as I told you your cytokininess
it is involved for embryogenesis callus
initiation root initiation that is more important
and shoot proliferation. So these all
points you have to remember for your uh cytochin.
The last one what we are going to see is
the abicyic acid.
Now this abysic acid it is also called
as ABA in the when it is present in the
culture media it has dual function. It
either it can stimulate or inhibit the
callus. So you have to choose whether
this ABA is useful for the development
of the plant or not. So based on this
again trial and error method only you
can use this abyic acid. Particularly it
is an inhibitor of plant tissue culture.
But in some plants they have seen that
it has the positive effect also. Another
important function of this abbisic acid
is the major effect of abicyic acid is
its capability to signal that it is
undergoing water stress. Like whenever
the plant is go growing in some dropped
condition, it can send a signal to the
plant. This hormone can send a signal to
the plant that we are under water
stress. No water is available. So don't
lose your water. So when a plant is
undergoing water stress the ABA levels
are increased they increase. So the
water level has come down. Okay. Thereby
what happened? Your ABA level has
increased. So what does this indicates? [Music]
[Music]
The ABA levels are increased
dramatically leading to the stomatal
closure thereby preventing the water
loss. So it what it does it increases in
concentration and you know what are
stomatas the guard cells and all it
closes the stomata so that from the
stomata you don't lose any water. This
is an important function but from tissue
culture point of view it is hardly has
Next is again italene. Ethylene is also
not that very commonly used. Even these
two thisic acid and ethylene from plant
tissue culture point of view they don't
have much functions to perform.
Otherwise in the general growing plant
in outside you can see that they have
lot of function like it is useful for
water stress. Ethylene is a gas and you
all know that it causes the ripening of
the fruit. For example, if this is an
unripe fruit ethylene, it induces the
ripening of the fruit. Similarly, the
starch in presence of amias is converted
to sugar and the chlorophyll, the green
color chlorophyll because of the enzyme
hydrolace, it is converted to antoine.
So these are the some of the example
which ethyline and episic acid have in a
normal plant. But when it comes to
tissue culture level, hardly they have
when we come to our growth regulators or
growth hormones, your take-home message
is auxen. The most important function of
your oxen is the development of root.
The most important function of your
cytokinine is the chute. That is what I
told the ratio of oxen to cytoines. If
the ratio of your oxen is more and your
cytoine is less, what happens? It
induces the root formation. On the other
hand, if your cytokinine ratio is more
when it compared to your oxen,
okay, it induces chute formation.
This one you just have to imprint in
your mind never to forget. Okay.
Gibbilence, it is involved in cell
enlargement or cell elongation also.
Abbyic acid, it depends. Okay. It
depends whether it is a stimulator or in
inhibit or it can inhibit the callus
growth depending you can have to use the
abic acid otherwise it indicates the
plant stress hormone
ethyline we don't use because it is only
used for the fruit ripening.
So these are the
important hormones which are naturally
produced by the plant and we have to use
them in different concentration
or the quantity and quality
so that we induce it into the media for
the development of the plant for the
cell division of the plant or for the
formation of the root shoot cell enlargement
enlargement
cell progression. So all this is done by
So before we could wind up
we will see into one more substance
which is very important. So you were
studying about inorganic nutrient,
macronutrient, micronutrient, organic
natural materials like your coconut
water, potato. You saw the importance of
charcoal, antibiotics,
vitamins importance, amino acids
importance. You saw now the what are the
hormones which are important for my plant.
plant.
So now we shall look into the
another system called as the solidifying
agent. So you know once I tell what is
the solidifying agent what you use in
your tissue culture or even in your
microbiology you immediately what
strikes to your mind is the agar because
you have used it. So the solidifying
agents these are jelling agents. Okay,
it is called as jelling agents or
solidifying agents and it is commonly
used to prepare semiolid or solid tissue
culture media.
Now why we are using is suppose say now
if this is my tissue culture vessel and
I'm growing my plant here and it what
happens if this is filled with only the liquid
liquid
what happens to my cell it gets immersed
in this liquid.
So once it gets immersed in the li your
liquid what happens ultimately there is
lack of oxygen for this cell to perform
its function. So it dies in the liquid
media. So what we are doing we are using
the solidifying agent or jelling agent.
So what happens there is a thick so
solid form which contains all your
nutrients and your plants can be kept
here so that it will not immerse into
it. So there is an air exchange and then
it'll not die. If it is a liquid media
it'll get submerged into the liquid.
Ultimately there is no oxygen and your
plant can die.
So the most commonly what we use is the
agar 0.5 to 1% agar. This agar is a
polysaccharide which is obtained from
the CV and it is inert that is it is not
going to react with any of your media.
Its main function is to just solidify
your media. That is why agar is most
commonly used. It is very inert. It is
not going to
interfere with your media at all or nor
the growth of your plant
and also they are not digested by plant
enzyme. That is I told you this plants
know they will be secretreting certain
residues. I told you like phenolics when
the phenolics are secreted what happens
your plant this media turns black in
color these are the impurities waste
which can be removed one method of
removing this impurities is your you can
use activated charcoal or you can go for
subculturing I'll tell you this all in
detail when we talk about that chapter.
So my plant what it is doing or my
tissue culture cell what it is doing it
will be secretreting certain enzymes. So
this agar has no effect on this enzyme.
You produce the enzyme but still I'm
strong. I will be like a solidifying
agent. That is what is the function of
the agar.
Now suppose if you want to study the
nutritional content of your tissue
culture that time you cannot use this
agar because this agar will have some
impurities okay from like calcium
magnesium sodium impurities it has that
time what you have to do is you have to
remove this impurities and then you have
to do the nutritional studies which is
not it is out of scope of our syllabus
okay but just remember it can be the
impurities can be removed by Using the
double distilled water, you rinse it for
4 hours in ethanol and dry it at 60°
centigrade for 4 hours. So all the
impurities will be removed and you can
study the nutritional content also.
Apart from a you can use another
chemical agent or the solidifying agent
which is gelatin which is used at a
concentration of 10%. But this is
limited use because it starts melting at
25°. So at 25° itself if it starts
melting what happen is it a jelling
agent or is it a solidifying agent? No.
So it has a limited use apart from
your gelatin. We can use aguros, we can
use algenate and we can use phytogel pha
agar. So these are some of the
solidifying agent or jelling agent or
supporting system so that it acts as a
support for your cell or tissue or
callus to sit on the media which is
containing all the nutrients what we
spoke about.
And here the last one you can see
mechanical filter paper bridges wigs and
wraps. Sometimes what happens is this is
particularly used in when we are using
this uh liquid when your media is in the
liquid form. So when it is in the liquid
form imagine this is all the liquid
form. Now I have to place my tissue
culture plant. Now if I place what will
happen it is just going to get immersed
into it. Instead of that what I'll do is
I'll make a filter bridge
and filter paper or wigs in
chromatography. You know that how we
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