Great updates to YFull

What is YFull?

YFull.com is an interpretation service for Y-chromosome Next-Generation Sequencing tests which include the Big Y test at Family Tree DNA and tests like the YElite 2.1 from Full Genomes Corp. YFull's interpretation includes identifying and rating new SNPs, showing hundreds of STRs that can be extracted from your file, placing you on the YFull tree, and much more. To see why you might want to use this service, see What are the benefits of YFull?

If you took the Big Y test from Family Tree DNA, for example, you will be able to submit your BAM file to YFull. Your BAM file is not automatically generated by FTDNA; you must request it first by going to your Big Y Results section and clicking Download Raw Data. Unfortunately, BAM files are not currently available because Family Tree DNA has been updating all Big Y results from the older hg19 (Build 37) to the more recent hg38 (Build 38) version of the Human Genome Reference Sequence. The BAM files should be available in early 2018.

In the meantime, YFull has been further improving the functionality of its service. These updates will greatly enhance our ability to understand our Next-Generation Sequencing results. Some of the new enhancements are shown in my post Big Changes to YFull. YFull released even more a few days ago, so let's examine them.

Great updates to YFull

On its Facebook page, YFull posted the following images of the latest updates.

Now we'll see this in practice.

Your YFull results screen

When your YFull results are returned, this will be your home screen. There are three main sections: Haplogroups and SNPs, STR results, and Novel SNP results. On the left of the screen are links to even more tools.

We will use the Novel SNPs section to examine the most recent YFull changes.

Novel SNPs

YFull's reporting of SNPs is quite a bit more extensive than Family Tree DNA's. In the Build 37 version of the Big Y, FTDNA identified a total of 26 novel SNPs in my brother's Big Y results. He had one match with a man named Cairns. 23 of the 26 new SNPs were shared with Cairns. These 23 SNPs were then named and placed on the haplotree, so they are not considered to be novel anymore. Three of the new SNPs were unnamed variants seen only in my brother's results.

When I submitted the BAM file to YFull for evaluation, YFull found a total of 48 SNPs which were categorized as best quality, acceptable quality, ambiguous quality, and low quality. Cairns had not yet submitted his results to YFull, and my brother has no other matches (yet!) in the YFull database. Therefore, all of the 48 newly-identified SNPs appear in my brother's novel SNPs section.

Low quality SNPs

When we go to the novel SNPs and check the "Low qual" tab, we can see that Yfull has identified two low-quality SNPs for this test. They are shown in the next image.

If we want to find more information about the second SNP on the list, we can click the yellow magnifier link.

Here is what the screen looked like before the latest update. We can see the hg19 and hg38 position numbers, how many times the SNP was read in the Big Y test, and what the reads showed.

In the above example, we can't tell if we are looking at information that came from the hg19 (Build 37) version of the test or the hg38 (Build 38) version, but we can see why this SNP was rated as low quality. The SNP was read only five times. Four times the test showed a T in this position, and one time it showed a G.

This SNP does not show up in the Big Y Results section of my brother's Family Tree DNA account because the results of five reads are not definitive enough to identify this as a valid SNP. However, YFull reports this SNP in the "Low qual" category because there is a possibility that this is a genuine SNP.

After the new update, here is how the same SNP position is shown:

The new screen has a red arrow pointing to the hg19 position number. This tells us that we are looking at information that was submitted under the former hg19 (Build 37) version. The black cursor is pointing to this SNP's region in the Y chromosome. The bar beneath it shows the exact position.

This position has not been identified in any database as a known SNP.

Best quality SNPs

Below are the "best quality" SNPs from the YFull report:

As you can see in the above screen, many of these "best quality" SNPs have now been named. Unfortunately, they have even been named more than once. The SNP names are shown next to the position numbers. YFull will identify a SNP as a known SNP in its database when the SNP has been shared between two or more testers at YFull. These "best quality" SNPs have not yet been seen in any other sample in the YFull database.

Here is one of the SNPs that is rated as "best quality."

It is located in the combBED (combined BED) area of the Y-chromosome where more stable SNPs are likely to be located. This position has been read consistently 103 times. It has been given two names: FGC65824 and BY20951. This SNP can be found in the ISOGG YBrowse database.

Solving a mystery with the new YFull updates

In my blog post The Big Y could be the best DNA test ever! I showed the process of evaluating SNP calls using Family Tree DNA's Big Y Chromosome Browser.

I could not find an answer to the mystery of SNP FGC46559 using this browser. I have copied the section about this SNP from my previous blog post to show why it was so confusing. The following describes how I searched for this SNP in the FTDNA system.

Using Family Tree DNA's Big Y Chromosome Browser

SNP FGC46559 appears on the Cairns and Thompson list of Non-Matching Variants. I searched for this SNP in my brother's account:

As you can see above, Thompson is not derived for SNP FGC46559. The reference and genotype are both listed as A which means that Thompson has the ancestral value and does not have a SNP here. Since it is a non-matching variant, Cairns must have this SNP.

This one is puzzling. When we click the SNP name and go to the Chromosome Browser, we see the following:

There are many more calls for this SNP that are not shown. You have to scroll up and down to see them all. This position was read a total of 76 times. The Reference value is A and is highlighted in red directly below the black arrow. All of the Genotype calls beneath it (in pink) are G except for that one blank space on the seventh line from the top. Yet the Genotype and Reference are both stated to be A.

What can we find about this SNP with the latest YFull update?

It doesn't appear to make any sense that the Big Y results show that my brother has an A at Position 19714591 when he clearly has a G.

At YFull, Position 19714591 appears in my brother's list of "acceptable quality" SNPs. We will click on the yellow magnifier to see what YFull reveals about this SNP.

This is one of the only SNPs in my brother's YFull account that has the blue YF icon. The blue YF icon indicates that this position is a known SNP in the YFull database.

At YFull we are looking at the hg19 version of the Big Y results. Here the number of reads is 94, and the hg38 Big Y results showed 76. This difference may be due to mapping to the new hg38 reference sequence. 

Notice that two different mutations have occurred in this position. SNP FGC46559 is a mutation from A to C at this position. This is rated as a high quality, five-star SNP in the YFull database. This SNP also appears in the YBrowse database.

My brother's results, however, indicated that at Position 19714591 he had a mutation from the ancestral value A to his derived value of G.  My brother's mutation from A to G has been named FGC65832.

Finally we can see why my brother's results are so puzzling in the Big Y Chromosome Browser. When we went to Position 19714591 in the Big Y browser, we were shown SNP FGC46559 which we now know is a mutation from A to C. Since my brother has the mutation A to G, he does not have SNP FGC46559. My brother has SNP FGC65832 which occurred at the same location. The Big Y browser does not yet show that two different SNPs have occurred at that position.

If we search for Position 19714591 in YBrowse, we will see the following screen showing the two SNPs FGC65832 and FGC46559.

But we can interpret this much easier in the new YFull browser because it clearly shows that FGC46559 is the mutation A to C and FGC65832 is the mutation A to G.

For this reason, I was particularly excited by the new YFull enhancements.

Verified by Sanger Sequencing

Position 12144810 has an orange check mark not shown in the other images.

When we hover the mouse over this orange check mark, we see that it indicates that the SNP has been verified by Sanger sequencing at the company YSeq. The term "verified" can be somewhat confusing. In this case, "verified" does not mean that it has been verified that my brother has this SNP.  All the term means here is that the SNP is available for Sanger sequencing at YSeq and that one person has been tested for this SNP. However, he did not have this SNP; it was reported negative.

In the example below, you can see that there is a green check mark next to one of the mutations.

The mutation G to A has been identified as SNP M241. It has been truly verified as a genuine SNP because 1008 people have tested for this SNP, and in 233 of these tests the results were positive. Because it has been proven that some people actually have a SNP at this location, the green check mark indicates that it is "verified."

A SNP is truly "verified" only when we can see a green check mark.

Why is Sanger Sequencing important?

Next-Generation Sequencing (the kind of sequencing used for the Big Y test) scans the Y chromosome and can find known SNPs as well as variants that have never before been discovered. But as we have seen, some positions are read many more times than others, and not all of the reads produce the same results.

With Sanger Sequencing, we can zero in on a particular position to see whether or not we actually have a variant at that location. We can verify the validity of many of our newly-discovered SNPs using Sanger Sequencing.

Furthermore, we can test any number of SNPs using this technology. For example, if someone wants to find out if he shares SNP FGC65832 with my brother, he can test that one SNP.  If he is wondering about several SNPs, he can test just those.

How do I make my newly-discovered SNPs available for Sanger Sequencing?

You can submit any named SNPs or unnamed variants to YSeq using the Wish A SNP option. Go to yseq.net and create an account, or log into your existing account.

Once you have an account, click Shopping Cart.

Now click "Wish A SNP" and follow the instructions on the next screen.

I will write more about the process of submitting SNPs to YSeq in an upcoming blog post.

I already have YFull results, what do I do next?

Examine your results with the new updates to YFull. If, for example, you do not see this icon for YBrowse, begin submitting your SNPs to YSeq.

When we don't see the icon for YFull, this means that we need to encourage more people to do Next-Generation Sequencing tests and to submit their results to YFull when the BAM files are available.

Thank you, YFull, for the great updates!

For more information see Advantages of submitting to YFull

Big Changes to YFull

On October 9, I posted the Big Y Update from Family Tree DNA. The announcement stated that FTDNA was updating all Big Y results from the older human genome reference sequence, hg19, to the most recent and more accurate reference hg38. This means that YFull, which interprets Big Y results, would have to include references to both hg19 and hg38 and be ready to process new tests mapped to hg38.

A few days later, on October 13, I wrote What are the benefits of YFull? Since then, YFull not only has added the hg38 conversion, but has added more tools to make their interpretation service more valuable. A lot has happened in less than two weeks!

Understanding Next Generation Sequencing

In order to best take advantage of the new enhancements, it is helpful to understand a little about how Next Generation Sequencing tests, like the Big Y, are processed. As you may remember, your DNA consists of two strands of DNA coiled into a double helix. The strands run in opposite directions.  One is called the forward strand, and the other is the reverse strand. The strands are connected by base pairs (bp) which are the As, Cs, Gs, and Ts that form your DNA sequence. All of these can be numbered to show their position on the chromosome.

During the testing process, your DNA is not read in one continuous stretch. Instead, your DNA is broken into random fragments. The test then reads these fragments from each end. Some fragments are read many more times than others. For example, one of your fragments may have been read two times, and another 56 times. Unfortunately, not all of the reads may give the same result. So a fragment that was read consistently 92 times will be reported as a high quality SNP, while one read five times with different results will be considered to be a much less reliable SNP. We will examine one of these "low quality" SNPs below.

After all the fragments are read, they must be reassembled, mapped to the human genome reference sequence, and given a precise location. Differences between your DNA results and the reference sequence are then reported. The human genome reference sequence is continually improving. Big Y results were formerly compared against the human genome reference sequence known as hg19 which was Build 37. They are now compared against hg38 (Build 38), and many of the position numbers have changed.

So let's put this basic knowledge into practice.

Updates to YFull

In my October 13 blog post What are the benefits of Yfull? I showed the following image of Novel SNPs from my YFull results:

Less than two weeks later, the same page looked like this:

If we compare the new version of my Novel SNPs screen to the previous version above, you will notice that the first line which contained position 7285772 now says 7285772 - 7417731 Hg38.  Next, you see a blue BAM icon at the far right of every line, and finally, on the sixth line you now see an orange check mark next to the letter G.

hg19 and hg38

The new screen shows both hg19 and hg38 positions. The first number is the hg19 position, and the second is the hg38 position. Even though the hg38 positions are shown, this does not mean that my results were mapped to hg38. We will see later how we can tell that my results are mapped to hg19. However, YFull will be accepting new FTDNA hg38 results.  .

Browse Raw Data

On the right of the Novel SNP screen are blue BAM links so that I can view any of these positions in my BAM file. I will be examining one of my low quality SNPs. We are now viewing the "Low qual" tab. I want to see why the second line in the image below (position 23096690) is considered to be low quality.

Before we use the BAM link, start by clicking the magnifier button at the left of position 23096690. The yellow magnifier is a link to view information about that position in a tabular format. It is the identical information that is obtained from the "Browse raw data" link in the gray menu on the left of the screen.

Here is what you see when you click the yellow magnifier or the Browse raw data link.

In the table above, after the chromosome positions you find a line for "Reads." Here the Reads are reported as 5. This means that Position 23096690 was read five times in my Big Y test. The next line is Position data: 4T, 1G which tells us that four of the reads indicated that I had a T in this position, and one read indicated that I had a G. This gives me a probability of error of .28 which is why this SNP is considered to be low quality.

The next two lines give very important information. My Sample allele was a T in this position (this is called the derived value). The Reference allele (hg 19) was a G, so at position 23096690 the human genome reference sequence had a G (which is called the ancestral value). The Reference (hg19) allele also indicates that my sample was previously mapped to the old hg19 reference, not to the new hg38 reference.

Using the BAM browser

Now, knowing the information from the Browse raw data table, let's click on the new BAM link to view this information in a Y-chromosome browser.

You will be brought to the Y-chromosome browser screen below:

At the top of the screen you will see the range of base pairs shown in your browser.  Here the range is from base pair (bp) 23096615 - 23096765. My SNP 23096690 will be shown within this 150 bp range. After the SNP positions is a list of browser styles: compact, 1, 2, 3, 4, 5, and 6.  You are seeing the compact version on this first screen. This compact style doesn't seem to be very informative unless you remember that Y Full always uses the color green for A, blue for C, orange for G, and red for T.  So you don't even need to see the letters to know that I matched the reference sequence except for a few calls for T where the ancestral value was G.

After seeing the compact version of the SNP browser, we will click on each style to see the differences.  Here is Style 1:

Style 2 is much more informative showing the five forward and reverse strands. Family Tree DNA in its new Y-chromosome browser color codes the forward and reverse strands.  YFull uses capital letters or lower case letters on each strand to distinguish between forward and reverse. Remember that in the Browse raw data table above, the position 23096690 had five reads--four were T (derived) and one was G (ancestral). Here you see five lines representing the five times that position 23096690 was read. The four derived Ts are highlighted in yellow.

Style 3 is similar to style 2 with the four Ts again highlighted in yellow.

Style 4 very dramatically shows where my test result differed from the reference sequence.

Style 5 is an enlarged image of the compact version style.

In Style 6 it is very difficult to find where my results differ from the reference sequence.  I might only use this if I were to conduct a "Find the SNP" contest with someone!

Here is Style 6 again with the arrow pointing to the column where my four Ts are different from the one ancestral G.

I find only Styles 2, 3, and 4 to be useful to me, but it's all a matter of preference.

The little check mark is big news

Now that we've seen the new Hg38 reference additions and examined a SNP in the new BAM Y-chromosome browsers, let's see the final addition which is the tiny orange check mark. This could be easily ignored, but it may be the most exciting change of all. If we click the orange check mark we can see that this SNP has been tested by Sanger sequencing at YSeq. Sanger sequencing is a method by which we can verify the validity of a SNP. YSeq is a company that will conduct Sanger sequencing on your SNPs at a reasonable cost. One person was tested for this SNP at YSeq, but his result was negative (meaning he has the ancestral value at this position).

On the Anthrogenica forum, user REWM posted the following image:

Instead of an orange check mark, this one is green. This SNP has also been verified by YSeq using Sanger sequencing, but in this case nine people were tested for the SNP.  Two of them were positive meaning they had a derived value in this position. The green check mark indicates that a SNP has been found in this position using Sanger sequencing.

These new SNP verification notices are wonderful news because we can now prove whether any of our questionable SNPs are valid and show the results on YFull. Many ambiguous quality and low quality SNPs have been proven to be genuine SNPs with Sanger sequencing. Here's how to take full advantage of this:

• Submit your novel SNPs (including any of your best, acceptable, ambiguous, and low quality SNPs) to YSeq through their Wish a SNP program. Use the hg38 position numbers. It will cost you one dollar per SNP. 
• YSeq will let you know whether each SNP qualifies for Sanger Sequencing, and if so, they will make all qualifying SNPs available for testing. If you want to test several SNPs, you can then design your own SNP panel through "Wish a Panel" to bring the cost down.
• Order your test and submit a DNA sample to YSeq to verify any of your SNPs. 
• In addition to receiving your results from YSeq, the validation will then appear on YFull.  

Thomas Krahn of YSeq has stated that YSeq and YFull are examining ways to better integrate their systems.  This means there are a lot more changes in store!

Consider submitting your BAM file to YFull

I have heard some people say that you don't "need" to submit your results to YFull because you can get good interpretations from other services including haplogroup administrators. This is true, you don't "need" to submit your results to anybody. But YFull's services are hard to beat, and they just keep getting better. YFull has many useful tools including SNP dating, reporting of about 500 STRs, and more. I examined a few of these in What are the benefits of YFull? In addition, many scientific studies rely on information obtained by consulting YFull. One of the many reasons I submit results to YFull is that if scientists want to discover more about my general branch of the human Y tree, I want my specific branch to be a part of it.

As more test results are submitted to the database, the interpretations are getting even better. Please consider submitting your results, too.


Update: See Great updates to YFull

Big Y Update from Family Tree DNA

Family Tree DNA announced changes to the Big Y test. These changes take place October 10, 2017, and will bring much needed improvements. 

We will be releasing a big update to Big Y next week!

Dear Group Administrators, We're releasing a big update to Big Y on October 10th and want to give you a first look before the release goes live. Once the release is live, we will be recalculating Big Y matches. We anticipate this to take approximately 5-7 days. During this time, you will see a "Results Pending" page when you click on the Big Y section. You will be notified by email once your results are processed and ready.   Once the transition is complete, we will update you as to when BAM files will be available.   What’s New? Here’s the breakdown of what we added and how it all works   Human Genome 38 We’ve updated from hg19 to hg38. This is a more accurate representation of the human genome and is the most recent version referenced by the human genome community. Some of the advantages of hg38 are: Better mapping of NGS data to the proper location Consideration of alternative haplotypes across the genome For more information about human genome builds, click here.   Terminal SNP Guide   We've added a terminal SNP Guide that allows you to view and filter the branches closest to the tester's terminal branch on the haplotree. BIG Y Browser   We’re giving you the ability to view your SNP data from Big Y. This will allow you to personally assess all SNP call positions that are being evaluated for matching purposes. This data will be continuously updated. Update by Linda Jonas, October 10, 2017: Conversion is now underway.

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