Day 47-48: I’m 95% Confident Freshman Can Do Statistics

Back again for fun and frivolity.

Monday, my students were examining their mounted slides (made previously) of their embryos from their Tep3/cyoYFP x Tep3/cyoYFP cross. They were collecting data on how many of their embryos died (obviously before we bleached and mounted them) and how many survived. This was a test to see if they saw 25% or 50% of their offspring die as embryos, which they were thinking would happen in the Tep3/Tep3 and cyoYFP/cyoYFP individuals. The 25% would come from just the cyoYFP/cyoYFP dying and the 50% would be if the Tep3/Tep3 were dying too, along with the cyoYFP/cyoYFP individuals; the other 50% surviving would be from the Tep3/cyoYFP individuals. So students took to their microscopes and recorded their data in a table, seen below:

Individual A/P Normal Denticle Belts Present Early Lethal Notes Slide Number:
1 Date:
2 Scored by:
3
4
5

The A/P normal refers to organisms the appeared to have normal development of anterior and posterior portions, a first step for determining lethality. Then they checked for denticle belts (see Figure A and B below) which are easily seen in developmentally normal Drosophila and could also determine defects. The early lethal column was where students determined if after those 2 checkpoints if the embryos were dead before mounting. Those organisms looked like opaque embryos (compared to dark) and did not display normal signs of A/P or denticle belt development (such as Figures C-J below).

After they determined this we created a class-wide spreadsheet that students imputed the number of alive vs. dead embryos they started with/counted. That was Monday.

Then Monday night I got a weird idea…

…what if I tried teaching my freshman some statistics?*

Obviously our question from Monday, “Are 25% or 50% of our embryos dying?” begged for some statistics, and pretty common ones too. We have data of how many embryos lived or died and we have a question that asks do our results closely match 25% or 50% lethality? Is our 35% close enough to 25% or 50%? (There were differing opinions in the room: talk about entry points, they needed no help randomly suggesting whether they felt 10% or 15% off from a value was of significance to them) I then said we have a test to determine significance in our data. We need Chi-Square!** (At this point, I would like to acknowledge, that given time I would have loved to run through Brad’s chi-square excel model with them [check the AP Discussion Board], but we did not have the chance. Maybe later. That was just a plug for a great student exercise)

So, I presented the problem to them again, this time with our data. (Let’s say 350 dead out of 1,000 embryos just as an example) I asked them if 50% were dying due to the Tep3/Tep3 and cyoYFP/cyoYFP mutation, how many of the 1,000 embryos would we expect to be dead? Of course, 500. Then I asked, “Ok, how off or “different” from that 500 are we for both the dead and alive organisms?” At this point I told them it might be easiest to set up a table. So they set up a table with a 3 rows and initially 3 columns. Eventually, they created this.

Outcomes Observed (O) Expected (e) o-e (o-e)^2 (o-e)^2) (o-e)^2)/e
Alive
Dead

They knew observed and expected pretty easily and finding the difference between the two. Because this gives them negative values I asked them are we looking for magnitude of difference or general significant vs. non-significant? Again, we decided negative values aren’t telling us too much, so I introduce the squaring idea. We then divide by e (I had no idea how to explain this principle to students. I think e is also variance squared but this concept would destroy their minds, so I’m not sure how much of a disfavor I did to students by not explaining this)

Either way, we got significant results for both the 50% and 25% (Yes, I made some students who didn’t pass out do both***) which told us that our data was not close enough to 25% or 50% to be just random coincidence. Something else is at play in our research. We know cyoYFP/cyoYFP individuals die, because inheriting two balancer chromosomes is lethal. So what’s up with our Tep3/Tep3 mutation? Apparently, it is causing death in embryos, but not all, and larva, but not all.

Interesting…sounds like an experiment.

*Turns out if you Google “teaching freshman statistics” you get a bunch of college results back. Changing it to “teaching high school freshman statistics” only gave me more college results. Google never listens to me.

**They reacted something like this:

***Hey, the ones who did it twice loved it.

Day 35-38: Counting is Hard

Remember when I said Inquiry is hard? Apparently, I was getting ahead of myself. Counting is hard. No, not that counting. Like, the 1, 2, 3 kind of counting.

Some days I need to breath and realize the hardest things for high schoolers can be the most mundane and seemingly simple tasks.

So Wednesday and Thursday we took a look at our flies and removed any larva that had hatched and placed them in a culture tube. This requires students to examine their petri dishes closely, to see if hatched larva are crawling around. They then picked them up with a toothpick (all if them at once if they could) and placed the toothpick in the tube. This makes it easy for the larva to crawl off instead of trying to make them get off.

Larva Tube
Toothpicks were put larva end down.

Teacher prep note: You’ll want to “score” the top layer of your fly food. This requires a normal amount (8-10 grains) of yeast and then 3 drops of water and using a popsicle stick, knife, of something flat you want to mash the top layer (see pic above) so that it becomes mashed and the yeast is mixed in. This is easier for the larva to eat, rather than regular sized yeast for adult flies.

Larva Directions
Water, Culture Tube, Yeast

After transferring the larva we marked on the tube how many were in there, for our data purposes. We did this for two days, which allowed for the embryos to have plenty of time to hatch if they were going to (aka – not die).

Today we took the petri dishes and counted* the remaining intact embryos, meaning the dead ones (They had been incubated for almost 4 days). This task was also not easy  because of the technique of picking up embryos with toothpicks. A quick how-to video can be viewed here, skillfully captured with one hand holding the camera over the scope lens and the other picking embryos. We placed those embryos on a microscope slide because since they are dead we want to examine them closer up to see WHY they died. The directions students got can be seen below:

Directions2
Note: After 3 steps students begin to just wing it.

Students were asked to place ANY embryo that still had an organism in it (see: dead) to be transferred to a microscope slide. They then had to record how many that was. Remember, previously they had recorded how many larva they had and of course the initial number of embryos they started with. The bleach was just ordinary bleach and was used to dissolve the egg shell. Our wicking device was cut up pieces of notecard. The water can be immediately wicked away after application, it’s just used to wash away the bleach. I then dropped in one drop of Hoyer’s Medium which is used as a mounting solution. Then to facilitate the mounting we placed them under a heat lamp that had been heating a hardtop surface to 37°C, although warmer is better, I wouldn’t go lower. That heat was applied for a couple hours.

Due to MLK day Monday we will be waiting a couple days for our larva to develop into flies and we’ll be back in business on Friday. Until then, happy trails.

*Why is this part so hard for students to do correctly? Explaining the importance of having accurate data and not quitting, even if they have a lot of embryos, seems to not make a difference to most. #FreshmanProblems?

Day 34: Inquiry is Hard

Today was supposed to be “the day”.

After all this time observing and working together to cross our flies it was the student’s turn to choose their initial research question that would guide them from here on.

*crickets*

So this is one of the hardest things for me that requires patience on my part. I noticed students beginning to turn off and quit. I had not given them a clear direction. Their direction was to “Choose your question and let that guide your observations and next steps today.” Too vague for them, freshman especially. What do I mean by “choose a question”? So I asked groups at the same lab bench (~4) to share their questions and observations to decide on a group direction. Still not much, so on the fly* I had them share their observations as lab groups on the front board so everyone could see what others had seen.

Group observations
Letters at top stand for lab group colors.
(Sorry I can’t take decent panoramas)

I spoke each group’s aloud and asked if students had questions about these observations and/or had those groups expound or clarify as well. This helped because students at tables where we were maybe less observant were able to see others had seen abnormalities/more in their flies. The questions that arose from this share out I got were generally superficial, although still important. They knew flies were dying (see: Tep3/Tep3) and that they did not make it to adults so roughly 75% of all my freshman chose “When are the flies dying?” as their direction. The ones that did not noticed something maybe a little more unique and intriguing.

Fly trachea
Can you see it? Middle-Left, the trachea are convoluted/not normal.

So some groups investigated what makes these trachea so odd and is that what is killing our larva before adulthood? They will be doing some cool biotech stuff later. Stay tuned for that.

After this we broke off into two groups, one trachea group and one “When are they Dying?” group to plan our next steps (Both AP and Freshman, who got a little more help). Each will continue the rest of the week with these experiments and I will detail them on Friday. Until then, enjoy the story below.

Two AP students were surprised to find all of their flies in their cups had died over the weekend. When I asked them what they think had happened they hypothesized old age, disease, mishandling, and undernourishment. “Interesting,” I said, “Do you remember feeding them?” (The apple juice agar plates have yeast paste on them for food.) They shook their heads. “Where was the food located for your flies?” I asked. Silence. Eventually, one of them said on the agar. “Great, so why did you flies die?” Still nothing. So I grabbed their agar plate and flipped it over so now the agar was facing the flies inside the cup instead of towards the outside. *Eyes widen* “We’re idiots” I reassured them they were not, but we discussed when experimenting with live organisms the importance of triple-checking whether we fed them and that the food is located in reach of them. Certainly made my day.

*PUN

Day 33: What Kills the Flies?

BACK!

Thanks to the Polar Vortex my flies got some extra hibernation time, but we brought them back out into the world today!

Luckily, mostly all of the culture tubes had viable offspring (minus the pink ones with strep – Awesome yet kind of scary to find in your fly cultures).

Today, I asked students to recall what the stages of the Drosophila life cycle are, how long each stage lasts, as well as the anatomical positions on the fly. For the most part, the remembered, it was the simplest way to see what they recalled of the basic terminology over break.

Then students in their pairs were asked to put their flies to sleep using FlyNap* and then count the ratio of orange-eyed, curly-winged to white-eyed, straight-winged individuals. The catch? There should have been ZERO straight-winged organisms. Why? Let’s refresh:

We crossed Tep3/cyoYFP x Tep3/cyoYFP in December, and our expected outcomes are  25% Tep3/Tep3, 50% Tep3/cyoYFP, or 25% cyoYFP/cyoYFP. We know that Tep3/cyoYFP have orange eyes and curly wings so 50% of our offspring should be curly-winged and orange-eyed. But what about the others? Turns out because the cyoYFP gene is on a balancer chromosome which means it can’t be inherited with another balancer chromosome or it’s lethal. So 25% don’t make it as embryos. But the other 25%? The Tep3/Tep3? Well, that’s our question, all we know logically is that they WON’T have orange eyes or curly wings. So if we see non-orange-eyed, curly-winged individuals, then we know the Tep3/Tep3 flies don’t live to adulthood. *DRUMROLL*

100% orange-eyed, curly-winged…“Hmmmm…”, my clever students say.

Interestingly, by telling students to look for orange-eyed flies or white-eyed flies (without connecting the dots for them like I did above) some students are tricked into thinking they must have SOME white flies. Mostly, we checked out these supposed “non-orange-eyed” flies and they always turned out to be orange-eyed. An interesting form of introducing bias, although only in the freshman, none of my AP students took the bait. Students then inputed their data into a Google Spreadsheet with their data. In AP, I asked a student to create their own spreadsheet and then share it with the rest of the class, I didn’t give them this.

Fly Data Example
Thanks, Google, I didn’t want to print gridlines anyway.

After that, the freshman transfered their adults to a new culture tube, UNLESS they asked a super research question. See we told students to listen to themselves and any question that popped up, which they should record and share with us. Normally, students see no incentive to do this, outside of their own curiosity. But there were so many dead pupa in their vials that is led to some questions like, “Why are they dying?”, “When are they dying?”, “Are they malnourished?”, “What are mostly pupa dying?” or my favorite:

What Kills the Flies Cup
student handwriting + panoramic shot = super-creepy

Ah! But those are not the normal culture tubes; they are urine-sample cups.** Students with fleshed-out questions and ways to investigate those were given a cup and an apple-juice agar plate to put their flies in…and that about provided all the jealousy needed to trigger a storm of questioning. “Why do they get a cup for their flies?!” “How do I get one?!” My response, “Well, do you have any good questions?” It seems the only thing needed to get students to take this seriously was some students getting something the others didn’t. No grades or extra credit. Not even a Jolly Rancher.

So how many good questions did I get from 130 students?

Incubator w Fly Cups
Trophies of great questions

So now with these cups, student pairs who didn’t have well-developed questions will now choose questions other students asked. So they still get a choice with what interests them and they’ll have more chances to ask questions that move them in new directions, but for now, we have an incubator full of initial questions. My AP students each got an apple-juice plate and cup themselves and will each be able to lead their own investigations.

Next time (Monday): Students will choose a question and begin observing the young growing on the new agar plates.

* Side-note: I’m not crazy about this method. It’s messy, it smells, it requires constant resetting by dipping the wands in FlyNap every hour. I know professional labs used CO2 and I’ve heard of some alternatives such as alka seltzer or other smaller CO2 sources. (SodaStream!?)

**DO NOT TELL STUDENTS THIS (or prepare for epic levels of gross-out)

Johnson County Science Cafe

Johnson County Science Cafe’

Help Us Begin Our Fifth Year of Science Cafes

Penguins, Penguins, Penguins: Now that I have your attention, let’s talk!

Speakers: Gary Wesche, Kansas City Zoo Penguin Education Coordinator
              
Date: January 7, 2014

Time: 6:30 pm

Location: Coaches Bar and Grill, 9089 W. 135th Street, one block west of 135th and Antioch, south side of 135th St.

Penguins, what can we learn from those birds as we learn about those birds? And why will people engage in difficult subjects if the talk is linked to a bird in a tux?

Gary Wesche is a local educator. In his 30 year career he has taught 4th grade in public school, taught middle school science in private school, taught worldwide classes from the West Antarctic Ice Sheet in 2009-10 from a tent with PolarTREC, and was an at home dad for 16 years to 6 children while acting and directing in musical theater. This last year Gary was an adjunct physics professor at Rockhurst University, was hired by the Kansas City Zoo under a grant from the Zoo Learning Fund to be the Penguin Education Coordinator, helped launch the international organization Polar Educators International, PEI, represented PolarTREC and Kansas City at the International Penguin Conference in Bristol, England, and found time to direct and choreograph his favorite musical for the Chestnut Fine Arts Center in Olathe.

For more information: biologycctrack@hotmail.com