2015 Oysterseed Skyline questions

Today I worked on trying to figure out what could have made my error rates from last week so awful. I re-did the Skyline DIA protocol many times and ended up realizing that some of my samples only had a few files listed in the protein, while others had all four. Asked Yaamini for clarification. I was told by Emma to make sure that I have four tabs in Skyline. Each tab represents a sample, and each sample has 4 files associated with it.

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Samples for Sequencing

Today I picked samples for sending off to get sequenced. Obviously I have to get this checked with everyone… and I’m guessing they’d rather see it in a spreadhseet form and not color-coded as I do now… have to figure out how to do this.

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June Goals

2015 Oysterseed

  • DIA error rates and move forward
  • Pubathon for 2015 Oysterseed paper
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Bioanalyzer and Pubathon

Images from my trying out the bioanalyzer. And I try again. Neither tries look great. I should try again. Pub-a-thon plan and DecaPod S1E7 published (crab mtg #3).

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May Goals

Crab Project

  • Create a visual representation of all of the successfully isolated RNA samples I have and figure out a pooling scheme for next Crab mtg
  • Publish new podcast episode every week
  • Run bioanalyzer on some samples
  • Work on google doc for project
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RNA isolation and Data org

More RNA isolation to replace those that had “out of range” RNA qubit results. Ended up contaminating them all, so unusable. Starting my journey of using R for data organization.

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Data Organization- info from Pam in excel

Today Pam showed me what she does in excel to move info from one spreadsheet into another based on a shared column. Plan for picking crabs to replace those that had “Out of range” qubit results (either no RNA in sample I pulled out; no hemolymph sampled in first place; or so little that it’s below limit of detection.

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Weekly goals

So last week was unfortunately not as productive as I had planned as a result of some dog-sitting shenanigans. So, to help myself get back on top of things, I’m making a list of what I want to and will accomplish this week. Firstly, tomorrow I have a presentation in my FISH 511 class, so I will be focusing on that today and tomorrow. The rest of the week will be as follows:

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Crab Project Updates

Updates on RNA isolation and Qubit progress and DecaPod progress and plan. Finally post S1E1 on project background!

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April Goals

  • RNA Isolation
    • Isolate RNA from all crabs within the subset I selected (2 more sets of 9 to go)
    • Run Qubit on ALL of the isolated RNA samples from the subset I selected
  • Podcast
    • Edit the three recordings I have that are about:
      • RNA isolation
      • Crab mtg #2
      • Background info on the project (Ep. 1)
    • Come out with an episode each week
  • Data
    • Organize into master spreadsheet - or at least start the process (morph data; qubit; EVERYTHING)
    • Add Qubit results to the spreadsheet containing the subset of samples that I’m working with to isolate RNA
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RNA iso with new protocol

Today I use the updated version of the RNA isolation protocol and find that it is so much better and isolating RNA than the generalized protocol. This one is based on what Sam did with extra samples from Sam in the fall of 2017. It adds some extra mixing and centrifuging steps that appear to be the key. Good results.

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March Goals

My goals for this month are to continue my current work. I also look back on how I did in February.

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Crab Meeting

Today we had our first crab project meeting. Our plan is to meet the first Thursday of the month (subject to change if need be).

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RNA isolation

Today I began the process of isolating RNA from the “official” crab hemolymph samples, although they can change if I mess things up, because there are many extra samples.

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Thesis Proposal Writing

This quarter I am taking FISH 521, which is the Research Proposal Writing and Professional Development course taught by Julian Olden. This class has been very helpful because it has guided me through the layout of the Crab Project and the plan of how it will all be performed. This is good becuase this is my first quarter of graduate school and I had only heard about the crab project around Thanksgiving of 2017.

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RNA isolation protocol and plan for tomorrow

Tommorrow I will be trying out RNA isolation on 2 extra Tanner crab hemolymph samples. Try protocol with 2 samples: 465 and 468 Both samples are yellow (from third sampling day) and are from crabs that are infected, immature, and held at ambient temperatures throughout the experiment.

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February Goals

I would have made this post on the actual first day of February, but I have been sick - the neverending cold. Anyway, for this short month I have some goals I’d like to achieve:

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Preliminary Crab Sample Selection for Seq

Today I chose some crab samples for RNA extraction. I made a chart with the tube numbers for the samples. All crabs are immature males. Due to a massive die-off in the warm temperature treatment groups, we unfortunately only have two infected and one uninfected sample per the final two sample days. As a result, the uninfected samples will not be “pooled”.

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