![]() |
![]() |
![]() |
![]() |
![]() |
![]() |
Brian Krueger is the owner, creator and coder of LabSpaces by night and Next Generation Sequencer by day. He is currently the Director of Genomic Analysis and Technical Operations for the Institute for Genomic Medicine at Columbia University Medical Center. In his blog you will find articles about technology, molecular biology, and editorial comments on the current state of science on the internet.
My posts are presented as opinion and commentary and do not represent the views of LabSpaces Productions, LLC, my employer, or my educational institution.
Please wait while my tweets load
![]() |
![]() |
![]() |
![]() |
![]() |
![]() |
How AAAS and Science magazine really feel about sexual harassment cases in science
A week or so ago someone forgot to close the door on our enzyme freezer tightly. I had just ordered $1,000 worth of NEB enzymes to make high throughput sequencing libraries too... Before the meltdown, I made a couple of test libraries to be sure that the protocol was worked out.
I was really worried about the enzymes being thawed overnight, but I was told by a couple of different people that they'd be fine. So I went ahead and processed all three of my experimental timpoints (probably a huge mistake...), and each timepoint has 6 different libraries (one for each DNA associated protein I yanked out of the cells). So there were a total of 18 samples. Problem is they all came out looking like this.
Now I'm going through my enzyme box and testing to find out which step got messed up. This should only take a couple of days, but re-doing those timepoints is going to be a complete pain in the ass. It took two weeks the first time. I'm wondering if maybe these samples can still be salvaged though. If one of the enzymatic steps in library creation failed, then the DNA in there should still be viable, all I need to do is redo that step with new enzyme. I'm probably going to end up redoing the timepoints anyway for replication purposes, but its a total bummer that I've probably lost a month's worth of work to this.
This post has been viewed: 1179 time(s)
![]() |
![]() |
I redid the adapter ligation and PCR today on some of the samples and the smears are super bright now. So either it really was the adapter ligation that failed OR running another 18 cycles of PCR just amplified what little product was there. I didn't do a "no ligation" control, so I'll run PCR on a sample I didn't religate and see what happens. If the smear is still crappy looking then I know the adapter ligation failed and my redo is good :) But I'm kind of pessimistic, so I'm betting my new good looking smear is just because of PCR, and I'm going to have to redo everything.
Stick more passive-aggressive “please make sure the freezer is SHUT” notes on the door! Every lab I’ve been in has this problem (and multiple capitalized notes on the door). Such a simple thing causes so much damage – lost experiments, damaged enzymes, and pissy lab mates - There has to be a solution.
Wow, that totally blows.
As for signs on freezer doors...I think they are only effective for about a week or so after they are put up. After that, it just blends into the background. If you wanted it to be truly effective I think you would need to have the sign changed every couple of weeks, put in a different spot, use a different font, say something funy etc...but who the hell wants to do that as a lab job (although it's better than other lab jobs for sure!).
![]() |
![]() |
![]() |
![]() |
Jaeson, that's not true at most places. Top tier, sure, but 1100+ should get you past the first filter of most PhD programs in the sciences. . . .Read More
All I can say is that GRE's really do matter at the University of California....I had amazing grades, as well as a Master's degree with stellar grades, government scholarships, publication, confere. . .Read More
Hi Brian, I am certainly interested in both continuity and accuracy of PacBio sequencing. However, I no longer fear the 15% error rate like I first did, because we have more-or-less worked . . .Read More
Great stuff Jeremy! You bring up good points about gaps and bioinformatics. Despite the advances in technology, there is a lot of extra work that goes into assembling a de novo genome on the ba. . .Read More
Brian,I don't know why shatz doesn't appear to be concerned about the accuracy of Pacbio for plant applications. You would have to ask him. We operate in different spaces- shatz is concerned a. . .Read More