This week was spent using the CLC software to visualize and analyze genomic data from some given samples (not data from our sourdough starters).

Working with CLC wasn’t too bad. It took me around 30-40 mins to work through all the graphs and absorb what I could from the data. Of course, that’s not including the time I had to wait for the data to be uploaded and whatnot! Personally, I really enjoy looking at graphs and statistical data like this so I thought most of the stuff done using the CLC software was pretty cool.

This graph above represents the amount of OTU’s in the samples that correspond with specific groups or types of bacteria. In this case, OTU’s were only defined if they were 97%  or more similar to the reference data used. The inner ring shows that 99% of OTU’s in the sample come from bacteria and then the outer ring shows, what I believe to be the different phylum of bacteria that constitute all the bacteria in the sample.

When more rings are added to the graph (not pictured) it is important to understand just how many different species of bacteria are present in the sample. By the time the last ring is broken up there are well over 100 different pieces to represent each individual species. The diversity is staggering!

I have decided to keep the same three questions from last week because I think they’re all super interesting questions that could be answered using our data.

Does adding fruit, such as a banana, change the microbial composition of a starter?

What sorts of “outliers” will be discovered? What kind of unique or interesting microbes will be found in some classmates starter samples?

Will all of my classmates control cups be gnomically very similar? Or will they be different based on the environmental conditions they grew in while in our homes?

 

Now that the sourdough starters have started to be DNA sequenced its time to talk about what exactly is being sequenced. There are two specific genes/regions within genomes that are being sequenced.

16S rRNA gene: found in bacterial cells

ITS region: found in eukaryotic cells such as yeast

To accomplish this sequencing the cells must lysed and all the debris must be removed until only the DNA is left. Then, a specific primer that will only attach to these genes is mixed in with the DNA solution and PCR is performed to make lots of copies of the targeted gene. Next, a unique “barcode” is added to each gene so it can be later identified. Finally, all the DNA samples are added to one tube and are sequenced using a computer program to generate useful genomic data.

When comparing shotgun and amplicon sequencing I believe it would be easier to prepare DNA for shotgun sequencing because it is not necessary to worry about tagging specific genes and introducing the correct primers to only amply them. However, when it comes to actually analyzing the results it would be a lot more tedious using shotgun because it would require working with thousands and thousands of genes and trying to find a particular gene would be like trying to find a needle in a haystack.

Some questions I have that could be answered by our sourdough genomic data:

Does adding fruit, such as a banana, change the microbial composition of a starter?

What sorts of “outliers” will be discovered? What kind of unique or interesting microbes will be found in some classmates starter samples?

Will all of my classmates control cups be gnomically very similar? Or will they be different based on the environmental conditions they grew in while in our homes?

 

This week I decided it was probably time to toss my sourdough starters out. They had served their experimental purpose, but I was definitely saddened to be finished with them.

The DNA of our starters are being sequenced this week because we are trying to determine exactly what types of microbes are growing in the starters. We want to know if there’s yeast, bacteria, or both and what species of microbes they are.  The end goal of the class project is to see if different fruits have different microbes on them and if they alter the starters, which would help to explain the cultural variations of bread in whatever region these fruits are grown in.

Before this class I knew a bit about DNA sequencing from prior work in a plant genetics lab. I have actually used PCR, Gel Electrophoresis and Sanger in the lab to isolate DNA. However, I don’t believe I’ve ever heard of NGS before.

The fundamental principles of Sanger and NGS are very similar, but the primary difference in the two methods is the volume of sequencing data during one run. NGS can sequence so much data because a “library” of DNA data must be created before so that it can be referenced.

The technology used for NGS is Illumina. From what I briefly gathered this technology appears to work by having sequences attach to other complimentary sequences as they move through a flow cell. Once a sequence binds it is amplified over and over again. Nucleotide bases are added one at a time and have fluorescent tags so each base pair emits a fluorescence which can be read by a computer program to determine the DNA sequence.

https://www.thermofisher.com/us/en/home/life-science/sequencing/sequencing-learning-center/next-generation-sequencing-information/ngs-basics/what-is-next-generation-sequencing.html

 

This week was the final week for growing my sourdough starter babies. 🙁

It’s strange how attached you can get to a cup of flour, water and microorganisms!

This past week my starters did not really change much at all. I continued to feed them on a 12 hour schedule. They never had a noticeable change in smell and the texture never changed that much. I believe they got a little bit thicker by the end of this week, but overall, they were still like a soupy consistency. The starters would rise and then fall pretty consistently, but not a lot, (as seen by the results below).

Quantifying Starter Rising

In Central/South America where bananas are often cultivated and grown there are many types of bread eaten by the people that live in the region. Just a few examples of breads include: Pupusa, Hojaldres, Pan de Coco, and Quesadilla Salvadorena. Many of these types of bread are savory and buttery, often a street food or a treat. Many don’t seem to be very similar to sourdough because they are very dense, more like a pita bread.

https://www.tasteatlas.com/central-america/breads