HOW IT WORKS
Now that you are a microbiome expert and know why the American Gut Project is important for discovering new information about our microbiome, we hope you’ll be inspired to get involved. Here’s how:
Once you make a contribution on Fundrazr, you’ll receive your sample kit in the mail. Your sample kit will contain everything you need to collect your sample and mail it back to us.
There are some important steps to follow before you collect your sample.
- First, you need to register your kit.
- Once you have done that, you’ll have to select your sample type-human, pet, or environmental.
- If you choose a human sample, you’ll digitally sign our consent form and take a diet and lifestyle survey. Then-you’re ready to go!
With all of the possibilities, you might be wondering which body site you should sample, and how to sample your chosen site. The series of videos below (available on Vimeo from Shelley Schlender) shows you how to decide which body site to sample and how to collect your sample(s)!
- What site should I sample?
- How to handle your swabs to not contaminate them
- How to sample your tongue
- Sampling the palm of your hand
- How to get that infamous stool sample
Once you’ve collected your sample, be sure to associate/log it on https://microbio.me/AmericanGut/. This step ensures that your sample is associated with the consent form that you signed; without this step being completed we are not allowed to work with your sample per the University of California’s Institutional Review Board.
Ok, you’ve signed the consent form, taken the survey, collected your sample and logged it, and mailed your sample back to us. Now what? What happens to your sample once it arrives in the Knight Lab at UCSD?
The first thing that we do when your sample arrives is to scan it with our barcode scanner. If you’ve associated your sample, this information comes out and we have the green light to process your sample-and you’ll get an email confirming that we received your sample. If your sample hasn’t been logged, an error code results and we will store your sample in the -20oC freezer. You’ll also receive an email alerting you to missing information with instructions on how to correct the issue. We’ll rescan your sample every couple of weeks to see if it has been associated.
We use liquid handling robots (yes, robots!) to help speed up the process of getting bacterial DNA out of your sample and to reduce human error-including chances of contamination. This high throughput process also helps us keep down the cost of sample processing-which in turn keeps the cost of contributing to the project low. Our liquid handling robots process 96 samples at a time. We always process eight “blanks”-wells containing only reagents but no sample material-to ensure our process is clean and not introducing contamination, so we process 88 American Gut samples at one time.
Once we have the bacterial DNA isolated from your sample, we now need to do something called Polymerase Chain Reaction (PCR) to amplify the bacterial 16S rRNA gene-the one that gives us the genetic information we need to identify which bacteria are in your sample. During this step, we add a unique DNA barcode to the bacterial DNA in your sample so that it can be sequenced together with hundreds of other samples. This barcode “tag” will allow us to determine which bacterial DNA came from your sample while we are analyzing the sequencing data. As with extracting the DNA from your sample, the PCR step is completed using liquid handling robots that process 96 samples (88 American Gut samples plus 8 blanks) at one time.
Once we’ve produced 16S rRNA gene “amplicons” from your sample, we measure how much DNA is present and combine the amplicons with amplicons from other samples and deliver them all to the Institute for Genomic Medicine at UCSD for sequencing. We usually sequence six 96 well plates-or 528 samples-at once. Again, processing things in bulk keeps the costs down for us, enabling us to offer participation in the project for a relatively low contribution amount. A few weeks later, we get sequencing information back from the sequencing center, and the fun-data analysis-begins!
The American Gut is an amazing cohort (all due to our American Gut participants; we couldn’t do it without you!)-to this date we have processed and analyzed nearly 7000 samples-and it will continue to grow. With each sample producing tens to hundreds of thousands of DNA sequences, the amount of sequencing data is already huge-and will only grow! As you can imagine, it takes a lot of compute power to work with that much data. The Knight lab has developed a data analysis software pipeline called QIIME (“chime”) to analyze American Gut data-as well as data from each of the hundreds of microbiome projects completed by our lab each year-and has access to super computers so that we can analyze your data before the next century. We have a dedicated team of developers and data analysts working behind the scenes to ensure that our analysis pipelines and super computers work smoothly so that we can chug through the American Gut data and get you your results!