Lab Post #24 (11/16/16)

Today we observed our starbursts that we inoculated last class. All three YPD amounts (325uL, 350uL, and 375uL) had nearly equal amounts of yeast growth.

IMG_7140.JPG
One of our plates which displayed yeast growth. Going clockwise they are 325uL, 325uL, 350uL, 350uL, 375uL, and 375uL.

Because each YPD density experienced similar growth, today we will be replicating the experiment with solely 325uL of YPD on each arm of the starburst. We chose this amount because it was the easiest to plate on the starbursts and therefore optimal. We also will be testing the functionality of the design by inoculating 20uL of yeast in the center of the starburst in hopes that it will experience outward growth in all channels.

IMG_7139.JPG
Plating two new starbursts with 325uL of YPD in each arm and 20uL of yeast in the center.

We also looked at our 3D print today which sadly did not have as good results. The acrylic liquid weld was not successful in dissolving the PLA in the PDMS mold. Because of this we considered cutting it open and manually taking out the 3D print and the gluing it together but the PDMS was harmed in the process and therefore we had to end this portion of our experiment.

IMG_7138.JPG
Our PDMS mold with the PLA manually cut out.

 

Lab Post #23 (11/14/16)

Due to some issues with growing the yeast, we decided that we would rather focus our research project on trying to find out if yeast can be grown in our analytical template. In order to do so, we are using the same model of the starburst to test different amounts of agar: 375 uL, 325 uL, and 350 uL with four replicates each.

As observed in the images below, we tested each amount twice, so as to obtain as much accurate results as we possibly can.

We inoculated 15 uL of yeast in each arm of the starburst.

IMG_7127.JPGIMG_7128.JPG

In regards to the 3D design, the PLA (plastic polymer used to make the 3D print) within the PDMS has not yet been dissolved by the acrylic liquid weld because the surface area of the channel is way too small for the acrylic liquid weld to enter and dissolve it. Because of this, submerged the PDMS into a few microliters of acrylic liquid weld and let it sit for 2 days.

Lab Post #21 (11/07/16)

We got our yeast starburst plates back today, and there was some growth in the Wild type plate in channel 6 and a little bit in channel 8. Note that there was only one replicate for the wild type due to contamination while plating the YPD.

img_1009

In this TOS3 deficient replicate, there was mold growth between channels 3 and 4. Also, there is some yeast growth within channels 4 and 6.

img_1010

In this TOS3 Deficient replicate, there are multiple spots of yeast growth in channels 4 and 8.

img_1012

In this SAK1 Deficient replicate, there is mold growth between channels 7 and 8, as well as no visible growth.

 

In this SAK1 Deficient replicate, we are not sure whether the red-ish dots between channels 5, 6, and 7 are yeast or mold. Note the lack of YPD in channel 4, due to resource restrictions.

img_1011

 

We also received our new 3D prints. However, while cleaning out the print to make it smoother, the manometer became detached. So, we fixed it using Gorilla Glue. In order to easily see if the mold is set, we broke the top part of one of the halves and wrapped the entire print in Saran Wrap.

img_1017
Here is our new mold, with the manometer glued to the box.

However, there was a problem with the PDMS leaking through the Saran Wrap. We fixed it by taking the top we broke off and gluing it to the open ended part of the box. The PDMS did not leak through, which was great, and we vacuumed the entire mold for 15 minutes to prepare for placing it in the oven for 30 minutes.

img_1019
This is where we vacuumed the mold (which is in the clear cylinder).

 

We also determined today how we will dissolve the PLA in order to remove the PDMS from the mold. Based on the test dissolves we started on Wednesday, we decided to go with the acrylic liquid weld because it removes the PLA is 20 minutes without destroying the PDMS, whereas the NaOH dissolved everything and the DRAINO had some effect, but was very slow.

 

img_1016
Result of the acrylic liquid weld; the crystallization is due to almost a week of exposure.

 

img_1015
This is the NaOH. There is nothing left behind.

 

img_1014
This is the DRAINO. There is some dissolved PLA, however it is very little.

 

Lab Post #20 (11/02/16)

Today we started lab by checking on our PDMS mold that we set during last lab. When we got back the mold. All PDMS was gone from inside of our 3D print and the outside of the print had a new slimy texture. We believe that for some reason, our 3D printed mold was slightly porous, enough to allow the PDMS to soak into the walls and move out of the mold.

That being said, our new 3D mold print design should solve that problem. The new design is to print the mold into two separate halves. That will allow us to line the inside of our mold with cellophane so when we pour the PDMS, it should not be able to seep through.

We also wanted to test a new novel idea at dissolving the 3D printer polymer. We placed a piece of scrap polymer and PDMS into a beaker of DRAINO and allowed it to sit. We did the same thing with 3M NaOH, and acrylic liquid weld instead of DRAINO to compare how each would affect the scrap polymer and PDMS from the time of now till next lab. After 20 minutes the acrylic liquid weld almost completely dissolved the polymer. By 25 minutes it was completely gone and the pdms was still the same as it was before.

Displaying IMG_1916.JPG
Acrylic Liquid Weld

Our goal for the rest of the day is to complete set up for our 2D assay. We printed 6 wax paper starbursts (2 replicates) and placed parafilm and solid ypd onto each. We then inoculated the yeast and put the wax papers in petri dishes and then it all went into a ziplock bag with wet paper towels to prevent drying out. The plan is to incubate every starburst and check growth on Monday.

Optical density of yeast today :  .5413 SAK1

Optical density of yeast today: .7345 TOS3

Optical density of yeast today: Unknown Wild Type

 

Lab Post #19 (10/31/16)

Today we checked upon our starburst and discovered that the yeast had dried out. We infer that the long period of time had caused the water to evaporate which did not allow for any observable yeast growth. We plan to repeat the process of making the starburst in the same environment, but for a shorter period of time that will not dry out the yeast. We also printed out our 3D-models and used acetone to smoothen the surface of our 3D-printed manometers. Furthermore, we poured PDMS into our mold.