A START TO BETTER UNDERSTANDING DNA SEQUENCING
WHAT'S THE POINT IN ALL THIS DNA SEQUENCING?
Now that I've finished growing my starters and I've turned in samples of each of them for DNA sequencing, its time to get an even closer look at what made these starters behave in the way they did. Before we can hop into this though, it's important to understand why DNA sequencing is important.
One of the broader purposes of this experiment is to gather data to determine if certain fruit starters grow faster/slower than our control starters and/or other groups of fruits.
Now, it's important to understand that a starter isn't made up of a single organism, rather it a whole community of microbial organisms, meaning that the smell, texture, etc. that were describe isn't for just one organism, but rather a plethora of organisms.
DNA sequencing will enable us to determine what types of microbes are the most common in which starters. Once we are able to determine the species of microbes present, we will have a better chance at understanding their genotypes and how their genotypes ultimately make up their phenotypes (a.k.a. the way they look, smell, etc.).
Not only will DNA sequencing allow us to determine the microbial species of the different starters, but it will also enable almost an entire class of GN312 students to better compare their DNA starters to each other. By comparing the different DNA sequences of the different starters, we can have an even better understanding of which microbial species are prevalent in which starters (for example, I grew a peach starter, meaning I can compare the DNA sequence of my peach starter to other classmates' who also grew a peach starter and determine which one is the most common for their starters as well and if they match up with my data).
WHAT DO I EVEN KNOW ABOUT DNA SEQUENCING?
Now that I've covered our main purpose(s) of DNA sequencing our sourdough starters in this class, you may be wondering, what did I even know about DNA sequencing before this? The truth is, not an entire lot. I mean, I've taken BIO183, which is a cellular and molecular biology class at NC State and for half the semester, we covered genetics, which included topics such as the structure of DNA, DNA sequencing, etc..
My memory is definitely a little hazy on the topic considering these topics were covered right around the time that classes got moved online and I didn't handle the adjustment as well as I would've hope to have.
I do remember performing sequencing with gel electrophoresis in lab for BIO181 during the fall semester of last year though. We sequenced different colonies of bees to determine whether they were an African honey bee, a western honey bee, or a mixture of both. At the time, I didn't understand what we were doing at all, considering BIO181 is a biology class that covers more of the broader biological subjects, such as ecology, evolution, and biodiversity; however, after taking BIO183 and learning about it more in genetics, I've find I've come to have a better understanding of it (although, I'm still not completely sure if I could provide the best explanation).
Gel electrophoresis - Above is a picture of Gel electrophoresis from the Nuts and Volts website.
UNDERSTANDING THE DIFFERENT TYPES OF DNA SEQUENCING:
So, I've talked a lot about what DNA sequencing is and my understanding of it, but I really haven't covered much on the many different ways DNA can be sequenced (I mean, I touched on gel electrophoresis, but there are so many more options for DNA sequencing due to advancements in technology).
Some of the main types of DNA sequencing technology are Next-Generation Sequencing (NGS) and Sanger Sequencing. When it comes to these two technologies, its important to note that NGS is more efficient due the fact that it can sequence DNA faster and it can sequence more of it at a time. You see, while Sanger is only able to sequence only DNA fragment at a time, NGS is able to sequence MILLIONS of fragments simultaneously in one sequencing run.
Now, since NGS is able to sequence millions of fragments at once, whereas Sanger sequencing is only able to sequence one fragment at a time, NGS is much faster than Sanger (and therefore cheaper). Since the more time a task takes to complete, the more money it will cost, NGS is cheaper because its able to do more work in a shorter period of time.
NGS is also more powerful in the sense that it can do a lot more work a lot faster than Sanger sequencing and it is able to detect novel or rare variants by using deep sequencing, whereas Sanger sequencing does't have the ability to do so.
WHICH PROCESS WILL IT BE?
Since I've covered the ways in which DNA can be sequenced and the importance of DNA sequencing in this research project, it's now time to reveal which DNA sequencing we'll be using. Drum roll, please!
It's...
NEXT GENERATION SEQUENCING (NGS).
After watching a short video by Illumina Sequencing, I came to quickly understand that we are using NGS to sequence our sourdough starters.
Now, I'm going to be quite honest, I didn't understand a majority of this video, however there were a few things I picked up on. I did learn that the there are four steps to NGS sequencing, which involve 1.) sample prep, 2.) cluster generation, 3.) sequencing, and 4.) data analysis.
I honestly am confused about most of the steps for NGS; however, I did understand that DNA sequencing is occurring due to clusters being excited by a light source, which ultimately produces a fluorescent signal that enables us to detect different strand of DNA. Based on previous articles I've read, I also understand that this method allows us to sequence millions of fragments simultaneously.
One other important portion I picked up on was the fact that we use flow cells, which are glass slides with lanes, to carry out the process of sequencing DNA (this is introduced in the cluster generation stage of NGS).
However, while there was some information I picked up on, there is still A LOT I'm confused on. For instance, what are oligos? The video talked about these a few times and I'm still not completely sure what they are or what their role in DNA sequencing is.
TO BE CONTINUED...
In conclusion, there is still A LOT I need to learn about DNA sequencing before I can say I have a really grasp on the subject. However, I've come to understand that DNA sequencing is important in the fact that it'll help us identify the different microbial species present in our sourdough starters and that it'll allow us to compare our starter's DNA to that of other classmates. We're are able to do this through the process of Next Generation Sequencing (NGS), in which case millions of fragments of DNA from our starters will be sequenced at once. Overall, there's still a lot to learn, but I'm starting to slowly understand the big picture behind DNA sequencing.
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