Today I isolated another batch of RNA! I totally missed the isopropanol in the flammables cabinet – quite embarrassing. However, it wasn’t listed in the MyChem inventory… but I did look at that bottle multiple times and it just didn’t register in my brain that it was what I was looking for. Isopropanol can also be called “2-propanol”, which was the label on the bottle I had been using thus far.
Anyway, so I was happy to continue with the RNA isolations today.
Another successful day!
Cummulative Qubit Results
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The curious case of the catalase(s)
You thought this series was over? April Fools, it’s not. I finally addressed one of Emma’s comments on my DNR paper that’s been sitting there since February. Emma wasn’t convinced that the two catalases I used as protein targets were actually separate proteins, so she suggested that I see if the protein sequences can be aligned. To do this, I opened up my Skyline file and looked at the protein sequences.
|Figure 1. Protein sequence for CHOYP_CATA.1.3
|Figure 2. Protein sequence for CHOYP_CATA.3.3
The letters code for different amino acids. The blue highlighted letters are peptides that in the Skyline document, and the black bolded letters are other peptides in the same protein. Lo and behold, you can align the sequences. Guess they are the same catalase after all!
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This is another attempt to isolate DNA from two more of the geoduck hatchery metagenome samples Emma delivered on 20180313.
The previous attempt, using DNAzol, did not yield any DNA.
I isolated DNA from the following two samples:
I used the DNA Stool Kit (Qiagen), following the “Stool Human DNA” protocol with the following changes:
- Incubated @ 95oC for 5mins after initial addition of Buffer ASL. This is a lysis step that might help increase yields (see the “Stool Pathogen Detection” protocol)
- Did not add InhibitEX Tablet. Deemed unnecessary, since these weren’t stool samples.
- Eluted in 50μL of Buffer AE
I opted to follow the “Stool Human DNA” protocol, as it processes a larger portion of the initial sample, compared to the “Stool Pathogen Detection” protocol (600μL vs. 200μl)
Samples were quantified using the Roberts Lab Qubit 3.0 with the Qubit High Sensitivity dsDNA Kit (Invitrogen).
10μL of each sample were used.
Neither sample yielded any detectable DNA. Will discuss with Steven.
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