Transcript for the Single-Cell Genomic Sequencing Podcast

Ann Nguyen:
Hi, everyone. Welcome to this podcast from Cambridge Healthtech Institute for the Next Generation Dx Summit, which runs August 19th through the 21st in Washington, DC. I’m Ann Nguyen, associate conference producer. We have with us today one of our featured speakers from the Single-Cell Sequencing Conference, Dr. Jan Vijg, Professor and Chairman of Genetics at Albert Einstein College of Medicine. Jan, thank you so much for joining us today.

Jan Vijg:
Sure, pleasure.

Ann Nguyen:
What's it like to do research with the Genetics Department at Albert Einstein College, your current environment, activities and resources?

Jan Vijg:
First I must tell you that I'm biased here, of course, because I'm the chairman of the department. I took over the department about six years ago, 2008, and it was a small department basically focused on doing developmental genetics with a number of model organisms. I first greatly extended it and then moved more into human disease-oriented genetics. There's lots of sequencing, computational genomics and epigenomics and an ever stronger focus on aging as a sort of a unifier of all those major diseases that occur during the aging process so that's really now cause a major genetic problem here. Of course, the theme of this whole meeting, single-cell approach, is becoming more and more important. To tell you how we are actually doing with respect to resources and current environment but everybody knows, of course, that the NIH situation is really bad, so basically the amount of money available for research is really decreasing. This makes it even more important to team up with each other and establish sort of small groups with a particular focus and then have joint grant applications. I think we are pretty successful in doing that. We are still reasonably well funded. Of course, I also have the benefits of having an endowment which also allows me to here and there provide people with some pilot money so overall I think we do quite well.

Ann Nguyen:
How has single-cell sequencing evolved and improved over time and what key challenges do researchers still face in this area?

Jan Vijg:
When I started to take an interest in single-cell analysis, that was I believe around 2004, 2005 or so in that time. Of course we didn't have any next-generation sequencing, so the end points were basically microarrays. Everybody knows that they are quite noisy. That's really a problem because when you take single cells and then you have to amplify either the whole genome or the whole transcriptome, you also introduce lots of noise, experimental noise and then, of course, it's sort of amplified then again when you have to read out on the microarrays. So I think there are two major improvements here. One is really that now we can use this digital endpoint, digital readout and that's of course, next generation sequencing, so we can do whole-genome sequencing and we can do RNA-seq and I think that's really very important. The second major improvement I think, is amplification. At a time when I started to do this, those were hardly available. It was really very difficult. You had to make it up yourself more or less, but now you can see more and more companies who provide them in kit form and actually work quite well. It still doesn't mean they are perfect but definitely there has been dramatically improved. I think those are really the two main technical improvements and then, of course, important when you want to start up a single-cell lab is that the instrumentation has become so much better. There are beautiful instruments available. Companies like Fluidigm, they really provide this whole infrastructure for single-cell analysis. I have nothing to do with Fluidigm, by the way, no interest in the company at all, but they obviously are like one of those companies who are zooming on this whole single cell [inaudible 00:03:13]. It is important.

What I think is key in the near future are two things. First is miniaturization so to get more high-throughput to do many, many single cells and then second would be the cost of the actual sequencing. That's still very important. That's still very high. This is really the areas that we have to zoom into. Important also, of course, is the front end part of it that's really basically to accurately capture single cells in the way you want them or a single nuclei from cells. The back end is the computation. That's of course also very critical but I think both the front and then the back end part will at the end of the day, quite easily improve. It's really like to make amplification better, as I said, and try to get the high cost of the sequencing better.

 

Ann Nguyen:
Finally, what will be the main theme of your presentation at the Single-Cell Sequencing Conference on August 21st and why this focus?

Jan Vijg:
My own focus is actually to use single-cell approach ... because, of course there are many reasons for using single-cell approach. Most people would say, "Well, we simply want to know what the heterogeneity is in a particular tissue or a cell population." That can be pretty dramatic [inaudible 00:04:13] when people seem to think that cells are sort of behaving all the same in a tissue but that is far from true. First of all, we don't even know if all those cells really are the same and not really different cell types. You can only figure that one out when you do single-cell approach. Second, there can be enormous variation even if cells are the same cell type, they can still vary like sometimes a thousandfold in particular responses to challenges. To really understand the impact of that, you really have to take a single-cell approach. In our research, I'm forced to take a single-cell approach because I'm interested in the accumulation in tissues of animal and humans of DNA mutations during the aging process and I want to know if that is functionally important. That seems like a fairly simple problem because the sequence, the tissue of an old animal and the same tissue from a young animal and then you find it out but that's not the case. The reason is that mutations are, of course, occurring more or less randomly so when you take the tissue as a whole, the frequency of a particular mutation can be very low, like 1 in 1000 to 1 in 10,000 so you will never see it. Of course, this is only because each single cell has its own unique spectrum of mutations. That's why when you take single cells and you sort of do whole-genome sequencing and then hold the variants, then you will be able to make a reliable estimate of the total load of somatic mutations in a tissue. The same, of course, is true for a tumor. In a tumor there's enormous variation between different cells. We all know that when you sequence a whole tumor then what you find is really the tip of the iceberg. You find 1/1000 or 1/10,000 of the mutations that are really there. That's why you actually do need to take the single-cell approach. We want to do that. What we're really interested in is to characterize the total landscape of particular tissues during aging with respect to somatic mutations. You really know what type of mutations you accumulate during aging, where do they accumulate. A very important question is how many per cell. If it's very, very low, then probably it will not affect function but if the total number of mutations in a very old tissue is really fairly high, it's likely that at the end of the day, it starts to affect functional pathways. It doesn't need selection to exert an adverse function but it can also directly cause a particular functional decline because it starts to affect the complex network that provides function in the cell. That's really what I will mostly discuss and I will discuss that in different organisms from drosophila to humans.

Ann Nguyen:
Excellent. Jan, thank you again for your time and insights today. We’re looking forward to hearing more of them in a couple of months. That was Jan Vijg from the Albert Einstein College of Medicine. He will be giving his featured presentation at the Single-Cell Sequencing Conference during the session, "Tips ‘N Tricks for a New Sequencing Frontier" at the upcoming Next Generation Dx Summit, taking place August 19th through the 21st at Washington, DC. I'm Ann Nguyen. Thank you for listening.

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