DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers
mtDNA Part VII - The Cambridge Reference Sequence
mtDNA Part VIII - mtDNA Test Types
mtDNA Part IX - mtDNA Haplogroup Determination
mtDNA Part X - mtDNA Subclades
mtDNA Part XI - mtDNA Haplogroup H
mtDNA Part XII - Subclades of mtDNA Haplogroup H  <<– you are here
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In the previous blog, we talked about mtDNA Haplogroup H and introduced the “Subclades of H”.  In this blog, we will discuss what is currently known about the subclades of H. 

Let’s take a closer look at the Subclades of H

Due to the large size of Haplogroup H and its wide distribution, there has been much research recently on the sub-clades of H, which surprisingly shows more restricted and regional geographic distributions. 

A study of the mtDNA of Haplogroup H individuals by examining the HVR1, HVR2 and control region of the mtDNA reveals a very large number of independent sub-branches, giving rise to subclades which have several further sub-branches themselves.  Studies to date reveal defined geographical patterns for Subclades H1 and H3.  All other subclades of H are found at a lower frequency and studies to reveal detectable geographic patterns are still ongoing. 

The following table summarizes what is known today about the subclades of Haplogroup H: (this table is based on a summary of current research published in peer reviewed journals and will be updated as more scientific data becomes available for the subclades of H)

In the next blog, we will begin to wrap up our discussion of mtDNA Haplogroup H by providing a detailed Haplogroup H distribution map.

DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers
mtDNA Part VII - The Cambridge Reference Sequence
mtDNA Part VIII - mtDNA Test Types
mtDNA Part IX - mtDNA Haplogroup Determination
mtDNA Part X - mtDNA Subclades
mtDNA Part XI - mtDNA Haplogroup H  <<– you are here
mtDNA Part XII - Subclades of mtDNA Haplogroup H
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In the last blog, we introduced “Subclades”, which are the finer branches of the Haplogroup tree.  One of the most well studied is mtDNA Haplogroup H.  In the next few blogs, we will discuss the latest research findings for mtDNA Haplogroup H and its subclades.

Who’s who in the field of Haplogroup H Research

Studies have shown that Europeans fall into one of several main mtDNA haplogroups:  H, I, J, K, N1, T, U2e, U3, Ur, X, W, U5, and V.  Haplogroup H is one of the most dominant family groups in Europe, representing approximately 40% of the mtDNA gene pool in populations in various parts of Europe and extending as far as western Asia.  Recent publications by the following researchers have provided significant advances in our understanding of Haplogroup H and its subclades:

• Roostalu et al. from the University of Tartu and Estonian Biocentre, Estonia
• Brandstatter et al. from the Innsbruck Medical University, Austria
• Pereira et al. from Universidade to Porto, Portugal
• Grignani et al. from Universita di Pavia, Italy
• Loogvali et al. from University of Tartu, Estonia
• Achilli et al. from Universita di Pavia, Italy

Table 1: Top peer reviewed research publications for mtDNA Haplogroup H 

This table lists the most significant papers for Haplogroup H in peer reviewed journals, with links to access the original publications.  These papers have provided significant advances in the current understanding of Haplogroup H and form the basis for the Haplogroup H subclade test panel. 

Let’s summarize the peer reviewed findings to date for Haplogroup H:

Several of the recent papers aim to provide resolution for the distribution of Haplogroup H and its subclades and begin to answer the fundamental questions of the origins of Haplogroup H:  where did it come from and where is it most concentrated in Eurasia?

To follow is a summary of what is currently known about Haplogroup H.  As more details are confirmed, this list will be updated: 

Despite the broad geographic distribution pattern of Haplogroup H, further investigation of each Subclade of Haplogroup H provides further resolution and reveals a much more specific and distinctive distribution pattern for each Subclade of Haplogroup H.

Haplogroup H is the most prominent European Haplogroup, and the papers were able to successfully sub-classify members in Haplogroup H into subclades H1 to H16 based on characteristic SNPs in the mtDNA, many of which are located in the coding region.  Next, let’s talk about the Subclades of Haplogroup H.

Subclades of Haplogroup H

The sub-clades of Haplogroup H surprisingly show more restricted and disctinctive regional geographic distributions. 

Based on the recent papers for defining the subclades of Haplogroup H, a subclade test panel is now available to test for sub-clades H1 to H16.  Individuals who are confirmed to belong to the Haplogroup H family can now take the H Subclade SNP Test to find out which sub-clade of Haplogroup H they belong to. 

Subclades H1 to H16 account for over 70% of individuals who belong to Haplogroup H.  The remaining 30% of individuals who belong to Haplogroup H belong to yet unidentified subclades of Haplogroup H.  As the studies progress, more subclades of H will be identified and will provide further classification.

The subclades of Haplogroup H can be defined by the following panel of coding region SNPs:

The mtDNA Haplogroup H Subclade Tree:

The 16 subclades of Haplogroup H can be summarized in the following phylogenetic tree.  Click here to view a more detailed version of the mtDNA Haplogroup H Subclade Tree.

In the next blog, we will discuss the features and distribution pattern of each subclade of mtDNA Haplogroup H.

DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers
mtDNA Part VII - The Cambridge Reference Sequence
mtDNA Part VIII - mtDNA Test Types
mtDNA Part IX - mtDNA Haplogroup Determination
mtDNA Part X - mtDNA Subclades  <<– you are here
mtDNA Part XI - mtDNA Haplogroup H
mtDNA Part XII - Subclades of mtDNA Haplogroup H
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In mtDNA Part IX, we discussed how to use your mutations to determine your Haplogroup.  In this section, we will show you how you can trace your ancestry further by using your mutations to determine your “Subclade”. 

What are Subclades?

All people living today can trace their maternal ancestry back to one of 26 core mtDNA Haplogroups.  Haplogroups are the main “trunks” of the mtDNA phylogenetic tree and represent extremely ancient family groups which arose tens of thousands of years ago.  Over time, the descendents of each Haplogroup formed further subgroups, called “Subclades”.  Once you discover which Haplogroup you belong to, you can further fine tune your results by tracing which sub-branch of your Haplogroup you belong to through “Subclade” analysis. 

Subclades are named using numbers and letters.  For example, Subclades of Haplogroup H include H1, H2, H3, H4,…. and so on.  The Subclade H2 can be further classified as H2a, H2b, etc.  Similarly, the Subclade H5 can be further classified as H5a, H5b, etc.

Which mtDNA Subclade tests are available? 

Subclade testing will be launched over the next few months.  Refer to the following chart for the launch dates of upcoming mtDNA Subclade test panels.  Once the subclade test is available, it will appear automatically as an upgrade option from your control panel.

Please remember the following rules for subclade testing:

1. You can only take a subclade test if your Haplogroup is known (either confirmed, or strong prediction)
2. You can only take a subclade test for your own Haplogroup.  For example, if you belong to Haplogroup T, you will not qualify for Haplogroup H subclade testing. 
3. If a Subclade test is not yet available for your subclade, check back occasionally as the control panel is updated as the science progresses.  Once more information is known about your Haplogroup and its Subclades, it will automatically be announced here.

Coming up next, H Subclades!

Since the next Subclade test to become available is the mtDNA Haplogroup H Subclade SNP Test, in the next blog, we will discuss Haplogroup H, and provide an overview of current scientific research and discoveries for Haplogroup H. 

In preparation for the next blog, we recommend downloading and printing the Haplogroup H Subclade Map.  You will need this map when we talk about Haplogroup H subclades in the next blog. 

DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers
mtDNA Part VII - The Cambridge Reference Sequence
mtDNA Part VIII - mtDNA Test Types
mtDNA Part IX - mtDNA Haplogroup Determination  <<– you are here
mtDNA Part X - mtDNA Subclades
mtDNA Part XI - mtDNA Haplogroup H
mtDNA Part XII - Subclades of mtDNA Haplogroup H
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In mtDNA Part VI, we discussed two ways that mtDNA testing can be used to trace ancestry: 

1. Direct Comparisons - search for matches, confirm or refute findings from ancestral studies
2. Ancestral Tracking - tracing your deep ancestry through “haplogroup” determination

 In today’s blog, we will focus on #2 “Ancestral Tracking”, and show you the science behind how the mutations in your mtDNA are used to determine your haplogroup “deep ancestry”.  We will always automatically predict your haplogroup for you after you take the mtDNA test.  However, the more you understand the science behind the technology, the more you will get out of your genetic genealogy experience.  New studies and data become available all the time in this fast moving and exciting field, so it is a good idea to know the basics behind how the technology works.

Let’s begin with a basic step-by-step guide on how to use your mutations in your mtDNA to determine your mtDNA haplogroup:

Step #1:  Download and print the Haplogroup Reference Guide	Click here to download and print the mtDNA Haplogroup Reference Guide.  This guide is a valuable reference tool for you.
Step #2:  Gather your mutations	Determine which mtDNA tests you have taken and which ones you did not take yet. The HVR1 Test will show you all mutations between 16000 to 16400 (shown in blue). The HVR2 Test will show you all mutations between 1 to 400 (shown in red). The SNP Haplogroup Backbone Test will show you all relevant mutations in the Coding Region (shown in black).
Step #3:  Identify your mutations on the Reference Map	Your mtDNA results report will always include a mutation table.  Examine your mutations table and circle your mutations on the Reference Map. Starting from the CRS, circle the mutations that you have.  Follow the path of your mutations away from the CRS.  Ignore the markers that you did not test. The final destination of the path mutations is the haplogroup that you belong to.

Let’s take a look at an example for a sample individual, Scott Mckenzie:

Let’s assume that Scott has only taken the HVR1 Test.  Scott has not yet taken the HVR2 or SNP Haplogroup Backbone Tests.  This is an example of an actual mutation table for Scott’s HVR1 Test:

Let’s see if we can tell which haplogroup Scott belongs to by examining the mutations in his HVR1 region.

1.  Find the CRS on the Reference Guide.  The first marker you encounter when moving away from CRS is “263″.  263 is located in the HVR2 region (HVR2 includes locations 1 to 400), so it is included in the HVR2 Test.

• Tip #1:  If you took the HVR2 Test, then look at your HVR2 mutation results and see if you have a mutation at location 263.  If you have a mutation at 263, then you can trace your ancestry away from CRS towards “Haplogroup H”.  If you do not have a mutation at 263, then you stay within CRS, and your haplogroup is likely the same as CRS.
• Tip #2:  If you did NOT take the HVR2 Test, then ignore 263 and move on to the next marker.

Scott did not take the HVR2 Test, so we can skip this marker for now because we do not know whether Scott has a mutation at 263.

2.  The next marker is “7028″.  7028 is located in the Coding Region, so it is included in the SNP Haplogroup Backbone Test.

• Tip #1:  If you took the SNP Haplogroup Backbone Test, look at your results and see if you have a mutation at location 7028.  If you have a mutation at 7028, then you can trace your ancestry away from Haplogroup H towards “Haplogroup HV”.  If you do not have a mutation at 7028, but you do carry a mutation at 263, then you belong to Haplogroup H. 
• Tip #2:  If you didn’t take the SNP Haplogroup Backbone Test, then ignore 7028 and move on to the next marker.

Scott did not take the SNP Haplogroup Backbone Test yet, so we can skip this marker for now because we do not know if Scott has a mutation at 7028.

3.  Next are markers 14766, 16067, 16298 and 72:

14766 leads to Haplogroup Pre-HV.  14766 is located in the Coding region and included in the SNP Haplogroup Backbone Test.

• Tip:  If you carry the 14766 marker, but not 16067, 16298 or 72, then you can trace your ancestry away from Haplogroup HV towards Haplogroup Pre-HV.

Scott did not take the SNP Haplogroup Backbone Test yet, so we do not know if he has a mutation at 14766.

16067 leads to Haplogroup HV1.  16067 is located in the the HVR1 region (HVR1 includes locations 16000 to 16400), so it is included in the HVR1 Test.

• Tip:  If you carry the 16067 marker, but not 14766, 16298 or 72, then you can trace your ancestry away from Haplogroup HV towards Haplogroup HV1.

Scott’s mtDNA HVR1 results indicate that he does not have a mutation at location 16067, so he is unlikely to belong to Haplogroup HV1.

16298 and 72 lead to Haplogroup Pre-V.  16298 is located in the HVR1 region so it is included in the HVR1 Test.  72 is located in the HVR2 region so it is included in the HVR2 Test. 

• Tip:  If you carry a mutation at 16298 and 72, but not 14766 or 16067, then you can trace your ancestry away from Haplogroup HV towards Haplogroup Pre-V.

Scott’s mtDNA HVR1 results indicate that he does not have a mutation at 16298, so he is unlikely to belong to Haplogroup Pre-V.

Scott did not take the HVR2 test, so we do not know if he has a mutation at 72, but based on the HVR1 results, we can predict that he is unlikely to belong to Haplogroup Pre-V.

In summary, based on a process of elimination (HV1 and Pre-V are eliminated), Scott’s ancestry can be traced through to Haplogroup Pre-HV.

4.  Next are markers 11719, 73, 16126, and 16362:

11719 and 73 lead towards Haplogroup “R”.  Both 11719 and 73 are located in the coding region.  Scott did not take the SNP Haplogroup Backbone Test, so we do not know if he has mutations at 11719 and 73.

16126 and 16362 lead to Haplogroup “Pre-HV1″ and they are included in the HVR1 Test.  Scott’s mtDNA HVR1 results indicate that he has a mutation at 16126, but does not have the 16362 mutation.  It is remotely possible that Scott might belong to Haplogroup “Pre-HV1″, and had a “back-mutation” at location 16362 (we will explain back-mutations in another blog).  Back mutations are rare, but it would explain why Scott has 16126 but not 16362.  We should hold onto Pre-HV1 and continue tracing to see if there is a better match for Scott.

5.  Next are markers 11251, 16126, 16278, 16311, 16071, 151, 12308, 16189, 10310, 16304, 249:

11251 and 16126 lead to Haplogroup JT.  11251 is located in the coding region, and 16126 is located in the HVR1 region.

Scott did not take the SNP Haplogroup Backbone Test, so we do not know if he carries the 11251 mutation.  Scott’s HVR1 results indicate that he carries the 16126 mutation so we can move through to Haplogroup JT.

16278 and 16311 lead to Haplogroup R1.  Scott does not have either of these mutations, so he is unlikely to belong to Haplogroup R1.

16071 and 152 lead to Haplogroup R2.  Scott does not have a mutation at 16071 so he is unlikely to belong to Haplogroup R2.  Scott did not take the HVR2 test, so we do not know if he has a mutation at 152. 

12308 leads to Haplogroup U.  Scott did not take the Haplogroup Backbone test, so we do not know if he belongs to haplogroup U, but we can look beyond Haplogroup U to the U Subclades to see if he falls into any of the Subclades of Haplogroup U:

• 16249 and 285 lead to Subclade U1.  Scott does not have a mutation at 16249 so he is unlikely to belong to U1.
• 16051 and 16219 lead to U2.  Scott does not have a mutation at either locations, so he is unlikely to belong to U2.
• 16343 and 150 lead to U3.  Scott does not have a mutation at 16343 so he is unlikely to belong to U3. 
• 16356 and 195 lead to U4.  Scott does not have a mutation at 16356 so he is unlikely to beong to U4.
• 16270 leads to U5.  Scott does not have a mutation at 16270 so he is unlikely to belong to U5.
• 16172 and 16219 lead to U6.  Scott does not have either mutations so he is unlikely to belong to U6. 
• 16318 leads to U7.  Scott does not have a mutation at 16318, so he is unlikely to belong to U7.
• 16224 and 16311 lead to K.  Scott does not have a mutation at either 16224 or 16311 so he is unlikely to belong to K.

Thus, we can conclude that Scott does not belong to any of the Subclades of the Haplogroup U family.  If the Haplogroup Backbone Test was conducted, we could confirm that Scott does not carry 12308 and eliminate him Haplogroup U entirely. 

6.  Next are markers 16069 and 16294:

16069 leads to Haplogroup J.  16069 is included in the HVR1 Test.  Scott’s results show that he does not have a mutation at 16069, so he is unlikely to belong to Haplogroup J.

16294 leads to Haplogroup T.  16294 is included in the HVR1 Test.  Scott’s results show that he has a mutation at 16294, so we can move through to Haplogroup T.

7.  Next are markers 16163, 16186, 16189, 16304, and 16324:

16163, 16186 and 16189 lead to Subclade T1 (a branch of Haplogroup T).  All 3 markers are included in the HVR1 Test.  Scott’s results show that he has a mutation at all 3 locations, suggesting that he belongs to Haplogroup T1.

16304 leads to Subclade T2.  This marker is included in the HVR1 Test.  Scott’s results show that he does not have a mutation at 16304, so he is unlikely to belong to Subclade T2.

16324 leads to Subclade T4.  This marker is included in the HVR1 Test.  Scott’s results show that he does not have a mutation at 16324, so he is unlikely to belong to Subclade T4.

In summary, based on the results of the HVR1 Test, Scott most likely belongs to Haplogroup T1. 

The prediction strength is strong for the following reasons:

• There are multiple markers in the HVR1 region that lead from CRS to Haplogroup T1.  HVR1 Testing indicates that Scott carries all of the mutations leading from CRS to Haplogroup T1.
• By process of elimination, most of the other haplogroups have been eliminated, with the exception of Haplogroup B.  However, due to the large number of matching markers for Haplogroup T1, it is most likely that Scott belongs to Haplogroup T1.  This prediction can be further strengthened if Scott takes the SNP Haplogroup Backbone Test which includes marker 11251 which is specific for Haplogroup JT branch of the haplogroup tree.  If Scott is positive for a mutation at 11251, then it would further confirm that Scott belongs to Subclade T1.

The study also showed a single match at 16126 for Haplogroup Pre-HV1, but Scott does not have the 16362 marker which is usually found in Haplogroup Pre-HV1.  Due to the stronger match for Subclade T1, it is far more likely that Scott belongs to Haplogroup T1 rather than Haplogroup Pre-HV1.  To further confirm this, the SNP Backbone Haplogroup Test will confirm whether Scott carries the mutation at 11719.  The HVR2 test will confirm whether Scott carries the mutation at 73 which leads away from Pre-HV1 towards the direction of Haplogroup T. 

Haplogroup Confirmation:

If Scott indeed belongs to Subclade T1, as suggested thus far by the HVR1 test, then the HVR2 Test and the SNP Haplogroup Backbone Test will further confirm the following:

• The results of Scott’s SNP Haplogroup Backbone Test are expected to show positive mutations for the following markers:  7028, 14766, 11719 and 11251.
• The results of Scott’s HVR2 Test are expected show positive mutations for the following markers:  263, 73.

As you can see from this example, Scott’s haplogroup is predicted using his HVR1 results.  In his case, the prediction was quite strong because Haplogroup T1 contains a lot of mutations from HVR1.  The prediction can be confirmed with the HVR2 test and SNP Haplogroup Backbone Test.

We hope that this lesson will give you a good insight into how mutations are used to determine an individual’s haplogroup.  In the next blog, we will dig deeper into the use of mutations for deep ancestral analysis. 

DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers
mtDNA Part VII - The Cambridge Reference Sequence
mtDNA Part VIII - mtDNA Test Types  <<– you are here
mtDNA Part IX - mtDNA Haplogroup Determination
mtDNA Part X - mtDNA Subclades
mtDNA Part XI - mtDNA Haplogroup H
mtDNA Part XII - Subclades of mtDNA Haplogroup H
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In this blog, we will provide a broad overview of the different types of mtDNA tests available, and in the next blog, we go over some case studies to help you understand when they are used, and what each one will tell you about your ancestry.

DNA Testing 101:  There are 4 main types of mtDNA tests:

1. HVR1 Test
2. HVR2 Test
3. SNP Haplogroup Backbone Test
4. SNP Subclade Test

Let’s take a look at the main differences between these test types:

Test Type #1:  HVR1 Test

Overview:  The HVR1 Test is the most informative mtDNA test, and it is always the first test that is performed when you start tracing your maternal ancestry (the Maternal mtDNA test). The HVR1 test uses “DNA sequencing” technology to read all of the nucleotides from locations 16,000 to 16,400 of your mtDNA.  This is the entire HVR1 region, located in the D-Loop of the mtDNA.

Highlights of the HVR1 Test:  The HVR1 region is considered the most informative region of the mtDNA for ancestral studies for a number of reasons:

1. The HVR1 region contains an abundance of markers.  The HVR1 region is located in the D-Loop, so it contains an extremely high concentration of mutations (ancestral markers).  Thus, this region is highly informative.
2. The HVR1 region is easy to test.  The entire HVR1 region can be easily tested using sequencing technology.  All 400 nucleotides in the entire HVR1 region can be read from a single test.
3. The HVR1 region is well studied.  The HVR1 region is the most well studied region of the mtDNA due to its high concentration of mutations and ease of testing.  Most scientific studies to date, including indigenous data and other anthropological studies, have focused mainly on the HVR1 region.  Thus, there is more scientific data available for markers in the HVR1 region than any other region of the mtDNA, making the HVR1 region by far the most informative region of the mtDNA.

Prerequisite:  There is no prerequisite for taking the HVR1 test.  The HVR1 test is aways the first and most fundamental test that is performed when using mtDNA to trace ancestry.  The HVR1 test can be used “stand-alone” for User search and comparisons and haplogroup predictions.  All of the other test types serve to supplement the results of the HVR1 test.  There are usually enough markers and hypervariablity within the HVR1 region alone to differentiate between different individuals who are not part of the same haplogroup or family line.  Also, due to the large number of highly informative markers found in the HVR1 region, your haplogroup can often be predicted from examing just the markers in your HVR1 region. 

Test Type #2:  HVR2 Test

Overview:  Like HVR1 testing, the HVR2 test also uses “DNA sequencing” technology.  This test focuses on reading all of the nucleotides from locations 1 to 400 of the mtDNA.  This is the entire HVR2 region, the second most important region in the D-loop of the mtDNA. 

Highlights of the HVR2 Test:  Like HVR1, the HVR2 region is also located in the D-Loop of the mtDNA so it contains many ancestral markers.  The HVR2 region is always tested in conjunction with or subsequent to HVR1 Testing.  The HVR2 test will supplement the HVR1 results in the following ways:

1. Strengthen the results of User searches and comparisons.  The more regions of the mtDNA that are used for comparison, the more stringent and precise the results of the comparison.  For example, in cases where people are matching exactly at the HVR1 region, further comparison of the HVR2 region will provide greater resolution, and further confirm or refute the matches found in the HVR1 region. 
2. Assist in haplogroup prediction.  The HVR2 region contains quite a few markers which are important for haplogroup determination, and when used in conjunction with HVR1, will provide a stronger haplogroup prediction (we will examine a case study in the next blog).

Prerequisite:  The HVR1 test is a prerequisite for taking the HVR2 test.  The HVR2 region is rich in ancestral markers and HVR2 testing is an excellent way to supplement the results of the HVR1 Test.

Test Type #3:  SNP Haplogroup Backbone Test

Overview:  While the markers in the HVR1 and HVR2 Regions can be easily detected by sequencing, the Coding Region is extremely large, making sequencing impractical.  The SNP Haplogroup Backbone Test is a special panel of approximately 20 markers in the Coding Region which are specific for haplogroup determination.  By testing just the HVR1 or HVR2, you will only receive part of the “picture” of your mtDNA.  The SNP Haplogroup Backbone Test will allow you to view markers in the Coding Region of your mtDNA.

The following table illustrates all of the SNPs that are examined in the SNP Haplogroup Backbone Test:

Prerequisite:  The HVR1 and HVR2 tests are prerequisites for the SNP Haplogroup Backbone Test.  The SNP Haplogroup Backbone test examines markers in the Coding Region of the mtDNA and together with results of the HVR1 and HVR2 test, will allow you to confirm your haplogroup.   

Test Type #4:  SNP Subclade Test

Overview:  The SNP Subclade Test examines a special panel of markers in the Coding Region of the mtDNA which allows you to determine which “sub-clade” you belong to once your Haplogroup has been determined.

Prerequisite:  The HVR1, HVR2, and SNP Backbone Tests are prerequisites for the SNP Subclade Test.  Your Haplogroup must be confirmed before you can proceed with SNP Subclade testing.  At the moment, SNP Subclade Tests are available for the following mtDNA Haplogroups:

• R
• M
• H

If a subclade test is not available for your haplogroup, please check back often as new sub-clade tests are added occasionally.  Whenever a subclade test for your haplogroup becomes available, you will be able to access it from your control panel. 

The four types of mtDNA Tests are summarized in the following table:

The main goal of mtDNA testing is to gain a complete understanding of all of the important SNP mutations in your mtDNA.  The HVR1 test focuses on markers in the HVR1 region, the HVR2 test focuses on markers in the HVR2 region, and the SNP Test Panels focus on relevant markers in the Coding Region.  In the next blog, we will go over a case study to allow you to understand how these tests work together to uncover your ancestry.

DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers
mtDNA Part VII - The Cambridge Reference Sequence  <<– you are here
mtDNA Part VIII - mtDNA Test Types
mtDNA Part IX - mtDNA Haplogroup Determination
mtDNA Part X - mtDNA Subclades
mtDNA Part XI - mtDNA Haplogroup H
mtDNA Part XII - Subclades of mtDNA Haplogroup H
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In this blog, we will discuss the Cambridge Reference Sequence (aka CRS).  The CRS is a fundamental part of mtDNA data analysis.  A basic understanding of the CRS and how it is used in determining mutations will allow you to understand the role that mutations play in tracing your ancestry. 

What is the CRS?

The CRS is the first human mtDNA that was ever fully sequenced and published.  The work was performed by scientists at Cambridge University, and this groundbreaking study was officially published in 1981.  Click here to view a copy of the original publication.

This publication represents the first time that the mtDNA was sequenced.  The donor whose DNA was used for this ground-breaking project was of European descent and belonged to European Haplogroup H. 

Since this was the first mtDNA sequence ever published, this sequence was thereafter refered to as a “reference sequence” upon which all further mtDNA sequences from labs around the world was compared to.  This original sequence eventually came to be known as the “Cambridge Reference Sequence” and all mtDNA which is sequenced, even today, is compared to the CRS.

Mutations are determined based on comparison with CRS

When we state that we have mutations in our mtDNA, we are actually showing the regions of our DNA which differ from the CRS.  Let’s take a look at an actual mutation report:

In this report, the HVR1 region was tested, and 6 mutations were detected, indicating that this individual’s HVR1 region differs from the CRS at 6 different locations.  Let’s take a look at the first mutation in the list:  16126 T>c.  This means that the individual’s mtDNA is different from CRS at location 16126.  It shows that CRS has a “T” at this location, but the person tested has a “C”. 

Let’s look at the same results based on the sequencing report:

All of the letters in “black” are the same as CRS.  All of the nucleotides in “Red” are different from CRS and are considered “mutations”. 

We are all compared to the CRS, not the earliest human mtDNA!

The key point to remember is that when the results of mtDNA testing are used for genealogical purposes, the results are compared to the CRS and mutations are reported as “differences” between the results and the CRS. 

This however, can lead to confusion for beginner genetic genealogists because instinctively, people usually think that when scientists look for mutations, they should be comparing the our mtDNA to that of the earliest human DNA to see how our DNA has changed over time.  However, that is not how the research community has decided to approach the mtDNA.  The consensus within the scientific community is that mtDNA is always compared to CRS.  Since this is the case, it is important for you to become familiar with how this “reverse” method is used to analyze our mutations and determine haplogroups. 

The role of CRS in haplogroup determination:

Let’s take a look at the human mtDNA haplogroup tree.  This is a phylogenetic tree which shows how all people living today descended from a common ancestor (mitochondrial eve) who lived in Africa over 150,000 years ago.  Every person living today can trace his/her ancestry to a branch of this tree, called a “haplogroup”.  The European individual whose mtDNA sequence is famously called the CRS is located at a distant branch of the tree as shown in the diagram below:

Where is the CRS located on the mtDNA haploplogroup tree?

Now, let’s take a look at how your mtDNA haplogroup is determined. 

To determine your mtDNA haplogroup, always start with the CRS and move away. 

Example #1:  If you HAVE mutations at locations 263 and 7028, and DO NOT have mutations at locations 14766 or 16067 or 16298, then you belong to Haplogroup HV:

Example #2:  If you HAVE mutations 263, 7028, 14766, 73, 11251, 16126, and 16069, and DO NOT have a mutation at 16294, then you belong to Haplogroup J. 

Example #3:  If you HAVE mutations 263, 7028, 14766, 73, 11719, 12705, 16223, 10873, 2352, and 150, then you belong to Haplogroup L3e:

Summary of procedure for determining mtDNA Haplogroups:  To determine your haplogroup, always start from the CRS and move backwards in the tree to see which mutations you have and which ones you do not have.  Your haplogroup is determined by the difference between your markers versus CRS. 

What if I don’t have any mutations?  If you do not have any mutations, that means that your mtDNA sequence (at least the part that was tested) is exactly the same as CRS.  CRS belongs to a branch of Haplogroup H, so if you belong to Haplogroup H, chances are that you will not have too many mutations in comparison with CRS.

This concludes the basic overview of the CRS.  In the Part VIII, we will start going over the different mtDNA test types available, and what each test type will tell you. 

DNA Lesson Series: The mtDNA and its role in Ancestry
mtDNA Part I - mtDNA 101
mtDNA Part II - Facts about mtDNA
mtDNA Part III - mtDNA Structure
mtDNA Part IV - Ancestral Markers
mtDNA Part V - Detecting Mutations in the mtDNA
mtDNA Part VI - mtDNA Ancestral Markers  <<– you are here
mtDNA Part VII - The Cambridge Reference Sequence
mtDNA Part VIII - mtDNA Test Types
mtDNA Part IX - mtDNA Haplogroup Determination
mtDNA Part X - mtDNA Subclades
mtDNA Part XI - mtDNA Haplogroup H
mtDNA Part XII - Subclades of mtDNA Haplogroup H
mtDNA Part XIII - Distribution of Subclades of H
mtDNA Part XIV - Descendents of Maria-Theresa
mtDNA Part XV - Luke the Evangelist
mtDNA Part XVI - Empress Feodorovna
mtDNA Part XVII - James “Earthquake McGoon” McGovern

In this blog, we will take a look at how the mutations in our mtDNA act as ancestral markers which allow us to trace our ancestry.

We all have a unique pattern of SNP mutations in our mtDNA.  Our SNP mutations can be used to trace our maternal ancestry in two ways:  1) Direct Comparisons, and 2) Haplogroup Determination.  Let’s talk about each one in more detail. 

1.  Direct Comparisons: 

By testing your mtDNA, you will discover the unique set of mutations that was passed down to you from your maternal ancestors along your direct maternal line.  Your mtDNA “profile” is the unique set of mutations that you inherited from your own mother, and it is unique to your maternal ancestry.  For example, all other descendents living anywhere in the world today who are direct descendents of the same branch of the “haplogroup tree” as you, will have exactly the same mtDNA profile as you (ie. you are linked through a common maternal ancestor).  Likewise, if someone does not have the same mtDNA profile as you, that means that he/she definitely did not descend from the same maternal line as you (ie. you are not directly linked on your maternal line).  Once you test your mtDNA markers, you can:

• Use your markers to confirm or refute findings from your ancestral studies.  Confirm possible linkages from your genealogical research and add branches to your family tree.
• Use your markers to search the Genebase database to find other Users from around the world who share the same mtDNA markers as yourself.

The more regions of your mtDNA that you test, the more precise the results of your comparison will be.  Your mtDNA contains several regions, namely, the HVR1, HVR2 and coding regions.  In the further blogs in this series, we will discuss the different types of mtDNA tests, and talk about which regions are examined by each test type. 

2.  Haplogroup Determination: 

The unique set of mutations that you carry in your mtDNA allows you to track your “deep ancestry’, ie. your ancestry from tens of thousands of years ago and discover your haplogroup. 

SNP Mutations are small “mistakes” that occur naturally in your DNA.  SNP mutations are rare, occuring at a rate of approximately one mutation every few hundred generations.  However, once a mutation occurs, it acts as a “time-and-date-stamp”, because it is passed on to all future generations.  Each mutation event can be linked to a time and place in history, and by testing the mutations in your mtDNA, you can retrace the history of your ancient ancestors. 

Let’s take a look at how mutations can allow us to trace the path of our ancestors using the following hypothetical example:

As you can see from this diagram, whenever a new ”marker” occurs, it is passed down to all future generations.  By studying all of the markers that an individual carries, we can tell them the story behind each marker, ie when did that marker first occur, and where did it occur.  By knowing when and where each marker in your mtDNA occrred, we can trace the journey of your ancestors back in time. 

In Part VII, we will begin examining some actual case studies using real mutations to see how DNA mutations are used to trace ancestry.  However, in order to understand the mtDNA mutations in the examples, you will need to have a basic understanding of the “Cambridge Reference Sequence”.  In the next blog, we will go over the Cambridge Reference Sequence and the fundamental role that it plays in mtDNA markers.