Biology Laboratory Investigation V: Onion Root Tip Mitosis Lab

Hi, welcome back to www.educator.com. This is the laboratory investigation 5, onion root tip mitosis lab.0000

Here is the introduction, the purpose of this lab is to calculate the amount of time0008

that onion root tip cells spend in the phases of mitosis.0013

If you are wondering why an onion root tip cell?0016

You have these meristems, meristematic tissue in plants, whether it is the apical meristems,0020

at the top and bottom, or the lateral meristems that widen plants.0027

Those are the more actively dividing tissues.0032

They are going through more mitosis events per day.0035

If we want to see a lot more mitosis, we are going to look at those regions.0039

If we are looking at other parts of the plant, like right in the middle of the leaf, you would not see as much mitosis.0043

Looking at this area in an onion root tip, where you can find the root apical meristem, you will see a lot of mitosis.0049

Materials, preserved onion root tips in microscope slides, stained for visibility.0058

As a teacher, that is my favorite thing to do, it was just easy, they are reliable.0063

You know they are going to be visible to students, if they know how to use a microscope.0067

Other teachers, I have actually seen them making the young onions have root growth and then making cross sections themselves.0073

Mounting those tissues on the slide and staining, that takes a lot more work.0082

I guess it would be more rewarding if you do correctly,0087

but I tend to go with the preserved onion root tips that have been there for years.0089

You need a compound light microscope that can go to times 400 magnification, so you get around this close.0096

Time required, about 45 minutes.0102

If you use your time wisely, that is all you need.0106

Part 1, mount the slide, actually get it there.0109

If you are using slices from an actual onion root tip that is alive or get the preserved slide,0115

put it on stage, and zoom into the root apical meristem.0121

The apical meristems, there is the shoot apical meristem and there is a root apical meristem.0125

We are looking at the one at the bottom.0129

Here is what you would see when you actually around times 100.0131

Times 40 that first view to the objective lens, you will be able to more zoom out.0137

Here is what is known as the root cap, this is what you do not want to zoom into.0142

You do not want to zoom into that area.0150

That is more protective cap, it is not actively dividing.0153

You want to zoom into the area up here, there is actually multiple regions,0158

multiple views in these areas, that are going to have some active mitosis.0165

If you look very carefully in the slide, you can see some phase of mitosis.0169

Later on in this lesson, where we zoom in times 400, it is much more visible.0174

You are going to zoom in until you get to that times 400 magnification,0180

that is where you have the appropriate view, you are close enough to really see those phases of mitosis.0185

We may not actually count all the phases, count the cells that are in each phase.0189

Interface, the cells in interphase look like plain old cells.0193

They just have a solid nucleus with a visible nucleolus.0197

During interface, you are between mitotic division, you do not have those visible chromosomes.0202

Prophase is when you have these chromosomes, you can see them condensed.0208

It is still in that nuclear area, they are still bundled together in the center.0212

Metaphase, they are lined up along the equator, the metaphase plate.0217

Anaphase is where they are being pulled apart.0221

Telophase is where you would have those groups of chromosomes towards the poles of the cells, about to be boned up into new nuclei.0224

You are going to record results along with the total number of cells that you actually counted.0232

Here is what we are going to be looking at.0240

When you are zoomed in times 400, you are going to be seeing something like this.0241

This is a typical view, you see about 80 cells.0244

I had a student see anywhere from 70 to 140 cells when they are zoomed in times 400.0249

It really depends on what specific area of the root apical meristem, you are on that specific region because the farther up,0257

the more superior you go, the slightly more elongated the cells are that they look physically bigger and take up more room.0265

This is a good view, you see a lot of things going on.0272

Let me highlights some cells, in terms of what phases they are definitely in.0274

If you look at this, this is definitely an interphase cell.0281

You can see the cell nucleus with a nucleolus inside of it.0291

Prophase, this is a classic prophase.0295

Some people would actually consider this prometaphase.0302

If you are in a more advanced biology class, there will be a lot more talk about prometaphase.0306

It is the phase that is halfway between prophase and metaphase.0309

You can see they are on their way to going to that metaphase plate, to be lined up along the center or the equator.0313

This, I would still consider it prophase for the purposes of this lab.0321

Here is metaphase, you can see they are lined up along the center.0326

Here is metaphase, here is anaphase, you can see how these chromosomes are being pulled to the side, there is anaphase.0330

There is a fine line between anaphase and telophase.0344

Sometimes, when you look get a view, you might not even see any telophase.0351

You may not see any anaphase.0355

It is just a coincidence and what was frozen in time, at that moment.0357

If I flash forward this a little bit further, and you have this bundle kind of more condensed over here but not looking like a new nucleus.0361

This one is a little condensing here but not looking like a complete nucleus, I would say that is definitely telophase.0368

This is close to telophase as well.0373

Actually, this may be a good candidate for telophase.0376

It looks like it is at its end, it is really a judgment call.0379

That is why this lab is only reliable, depending on who is looking into view and who is doing the counting.0385

Mistakes can be made, in terms of saying all that was definitely telophase, that was anaphase, and vice versa.0390

The more data you have, the more times you do this and average out the results,0396

the more accurate your figures are probably going to be.0401

What you will do is you add up all the numbers here and I will give you an example on the next slide.0404

Part 2, in terms of getting more reliable results, you can move to another part of the root apical meristem,0411

another view, or use another root tip, a completely different root tip.0417

Zoom in times 400 again and once again, count the phases, from interphase to telophase in the second region.0421

You can do it the third time, if you want.0427

Average the numbers to maximize your accuracy.0429

Divide those numbers for each phase by the total number of cells to approximate the percentage of time you spent in each phase.0432

Let me give you an example.0439

Here is interphase, prophase, metaphase, anaphase, telophase, and here is the total.0442

Let us say with view 1, we got 52 for interphase.0454

Let us say that for prophase we get 10, metaphase we saw 4, anaphase we saw 8, and telophase we saw 6.0461

The total ends up being 80.0474

We need to get the percentages, in terms of the relative amount of time based on this data that cells are spending in a various phase.0478

If we divide 52 by 80, that gives you 65%.0485

10 divided by 80, 12.5%, 4 divided by 80 is 5%.0492

8 divided by 80 is 10%, and 6 would be 7.5.0501

It adds up to 100%.0508

If you do this again, you look at a second view and count up all those cells.0511

You may get 70 cells in interphase which would, if you average out the data0516

that would increase your percentage of interphase, closer to 70%.0522

In the second view, you may actually have less prophase, you may have less telophase or more anaphase.0527

Like I said, the more lab groups that are doing this, the more views you look at over the microscope, the more cells you are counting.0533

As you average those things out, you get more reliable data, in terms of what is actually going on with the meristematic tissue.0541

Analysis, how effective is the method of counting cells.0549

Like I hinted earlier, it depends, it depends on the student or teacher knowledge of precisely what cells are in what phases.0552

Sometimes, it is a judgment call, sometimes it is like literally in between prophase and metaphase.0560

You can count prometaphase, if you want, as a separate phase.0566

It is in between anaphase and telophase, you just have to pick one or the other.0569

In the end, you are just learning from this experience, in terms of getting a sense of0573

what the cells are doing and what they look like in this different phases of mitosis.0578

Since, these cells are dead, the ones I would be using, it will be frozen in time at the living tissue.0583

Would it be better to go with living tissue?0589

Not necessarily, there are advantages with using these preserved tissues.0592

If you were to buy all of these actual live onion root tips and plan on doing the slices and making them thin enough0598

and staining them, there is room for error, in terms of not doing it correctly or not having thin enough slices,0608

and being able to visualize the chromosomes.0614

Having them preserved, these preserved dead tissues from formally living onion root tips,0617

in terms of being able to see what is going on in there.0625

Also, you do not have to worry about movement.0630

When they have been sliced, the relative amount of movement and activity may be hampered,0634

maybe inhibited a bit because of you have cut through them and probably damaged some cells in the process.0639

There are some advantages to using the preserved tissues.0648

How can you make this more accurate or reliable?0651

Like I hinted before, more groups doing more counting or if you are doing this by yourself, do 8 views.0654

Those little errors, in terms of you did not see any telophase, maybe there was a telophase you just did not see it.0662

Or did not know that anaphase is actually telophase.0669

Those little errors will become less problematic, the more data you have.0671

The connection to mitosis as a whole, the rate of mitosis varies from species to species, and tissue to tissue.0678

There are parts of your body where cells are not dividing at all.0686

There are plenty of parts in the brain, in your skeletal muscles, or cells that I’m going to call G0.0689

A phase that is basically exit or out of the cell cycle.0694

It is not going to have active cell division throughout the life of the cell.0697

There are other parts of the body, specially your skin, the membranes within your digestive tract that have to divide,0702

specially if they are being damaged over time or subjected to friction.0709

It varies species to species, two different plants can have completely different mitotic rates.0714

Within one plant or one animal, like I said, using the meristem, more actively dividing tissue,0720

less percentage of interphase relative to mitosis.0727

There are other parts of the plants where interphase is closer to 100% of the time because the cells are not dividing.0730

Since we are using actively dividing tissue, their relative mitotic rate was higher.0736

The amount of time spent in interphase was lower.0742

That is what the meristem is doing, lots of cell division, a lot more of the m or mitotic part of the cell cycle.0745

Comparing this data to meristematic tissues from other plants, will be revealing.0753

Do onions actually have faster dividing cells than the root apical meristems of other plants?0758

I know the answer to that but it is certainly worth investigating.0768

Here is a lovely view of a plant cell from another plant.0771

Presumably, this is not actively dividing but I like this view, I think it is it is gorgeous.0775

You can see all those lovely chloroplasts.0781

With this lab and others, just reveals the intricacies and lovely things going on with living beings.0783

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