Trying not to wing it! – Week 7 Update

Oopsie, I forgot to post this week 7 post when I should have a month ago but here she is:

This week was definitely more rooted in dry lab as we spent most of our time in ISC researching RNA sequencing techniques and various different DNase protocols. We are almost ready to send out some samples to be sequenced but, as with most things in science (and life), sequencing costs A LOT of money so we want to make sure all of our samples are the best quality possible so we get data from all of them!

One of our main focuses of our research this week was looking into how exactly our RNA will get sequenced. Knowing all of the steps involved in the amplification and sequencing process would help us know if our samples are pure/concentrated enough to be sequenced successfully. We thought this would be a pretty straightforward protocol to read up on, but as usual, this was not the case. Many companies keep all of their reagents and protocols pretty secret, or as we like to remind ourselves in lab, everything is always “propriety”. Good for the companies, not so good for us. We managed to find some videos that described the RNA sequencing process in general terms however nothing was quite detailed enough for us to glean the information we wanted. Luckily, our advisors have some experience with getting RNA sequenced so, based on past experience, they think our numbers look decent (knock on wood), so we should get good data from these samples (hopefully). 

Our other main focus of our research this week was DNase. There is an option for us to add a step to our protocol and do a DNase treatment on our RNA samples in an attempt to get rid of extraneous DNA and leave only “pure” RNA in the tubes we send to be sequenced. It would seem that this would be an obvious choice to make, however getting rid of DNA comes with the risk of also destroying our RNA as well. Thus, we have to make the decision about whether to treat our RNA (and run the risk of getting rid of some of it in the process), or not treating it (and run the risk of sending a less pure sample to be sequenced). Again, our research came up without a solid solution but we are planning on discussing alternative solutions and other options with our advisors!

While pipettes might be fun, computers are very important research materials too!

While pipettes might be fun, computers are very important research materials too!

Apart from our daily dissections, we didn’t get into any serious wet lab until Friday. We decided to try the DNase step just to see what our yields would be if we did decide to add this step. As expected, they did end up being lower than usual but they weren’t crazy low so DNase treatment is still looking like an option from the wet lab side of things. It was also fun adding this new step because we get to use a few new reagents and new machines which is pretty nifty, but stay tuned to see if we decide to keep it up!