4 - Genomic

Forensic utility of mitochondrial haplogroups in Brazilian individuals
derived from coding region SNPs
Elaine C. Favaro, Denilce R. Sumita & Martin R. Whittle
Genomic Engenharia Molecular Ltda, São Paulo, SP, Brasil.
IV Congresso Brasileiro de Genética Forense
São Paulo, 2013
INTRODUCTION
The analysis of mitochondrial DNA (mtDNA) is a useful tool is studies of biogeographical ancestry, and in casework where the quality of nuclear DNA makes analysis of this genomic DNA inadequate.
Sequencing of the hypervariable regions is usually undertaken in these cases but coding region SNPs can also be used to discriminate between individuals, especially when they share hypervariable
region sequences. These coding region polymorphisms can be easily genotyped using small quantities of starting DNA. Additionally, the inference of the mtDNA haplogroup can be made directly from
this information without needing to perform comparisons between sequences.
The aim of this work was to develop a simple multiplex enabling the typing of coding region SNPs for use in forensic casework and for ancestry studies.
MATERIALS AND METHODS
Because of the significant racial admixture present in the Brazilian population, the choice of coding region SNPs was made to maximize the coverage of the most common mtDNA haplogroups, as
shown in Figure 1, and these characterize the major human population groups, namely African, Asiatic and European. The chosen SNPs were genotyped by minisequencing (SNaPshot, Applied
Biosystems) as a single panel and shown in Figure 2.
Figure 2
Figure 1
13789C
L1
3594A
10398G
L1/L2
10115C
L2
3594G
10398G
L3
*
10398A
10400G
N
663G
A
8251A
W
14470C
X
1791A
X2
10398G
10400A
M
12705C
R
3970A
R9
12406A
F
del G
B
12714C
R9b
1719A
N1
14766C
H/V
7028G
H
12308G
U
4580A
V
14798C
K
11251G
J/T
13708A
J
5417A
N9
4833G
G
13263C
C
1719A
I
5178A
D
7600A
E
Legend:
* Start
blue = african
green = asiatic
red = european
15607G
T
RESULTADOS
RESULTS AND DISCUSSION
The resultant multiplex was tested in 286 Brazilian individuals from the states of São Paulo and Rio de Janeiro, so confirming the presence of the major haplogroups in this population. The observed
haplotypes showed a good correlation with the information derived from the hypervariable region sequencing and, in some cases, the two types of data were complementary to one another. Upon
assigning phenotypes to the samples, we observed that 91% of those having African origin had the expected mtDNA haplogroup (Table 1). For individuals of Asiatic ethnicity, 89% had the expected
mtDNA haplogroup, and the same was true for 41% of individuals of European ethnicity.
Table 2
Table 1
Sample
E3.3
SNP
White
Asiatic
Number of
Individuals
Percentage
A
11
B
Back
Number of
Individuals
Percentage
8.9%
3
13
10.5%
C
6
D
Total
Number of
Individuals
Percentage
6.1%
2
5
10.2%
4.8%
1
4
3.2%
F
-
G
28600
28605
28610
29086
29088
29499
29500
30286
30289
30679
30681
31351
31396
31397
31526
31528
31937
32455
32614
32615
32618
32634
32635
34412
4
4
4
4
4
4
4
2
2
4
4
2
4
4
4
4
2
2
4
2
2
2
4
4
3534
A
G
G
G
G
G
G
A
A
A
G
G
G
G
G
G
A
G
G
G
G
G
G
A
A
10398
G
A
A
A
A
G
A
G
G
G
A
G
A
G
G
A
G
A
A
A
A
G
A
G
G
10400
G
G
G
G
G
A
G
G
G
G
G
G
G
A
A
G
G
G
G
G
G
G
G
G
G
Number of
Individuals
Percentage
1.8%
16
5.6%
13263
A
A
A
A
A
G
A
A
A
A
A
A
A
G
G
A
A
A
A
A
A
A
A
A
A
2
1.8%
20
7.0%
5178
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
2.0%
4
3.5%
11
3.8%
7600
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
4833
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
19
38.8%
1
0.9%
24
8.4
5417
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
-
1
2.0%
-
-
1
0.3%
8251
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
C
-
-
1
2.0%
-
-
1
0.3%
663
A
A
A
A
A
A
G
A
A
A
A
A
A
A
A
A
A
A
A
G
A
A
A
A
A
M
-
-
9
18.4%
-
-
9
3.1%
14470
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
G
A
A
A
A
H
29
23.4%
-
-
-
-
29
10.1%
1719
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
12705
T
C
T
C
C
T
T
T
T
T
C
C
C
T
T
C
T
C
C
T
T
T
C
T
T
J
3
2.4%
-
-
-
-
3
1.0%
3970
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
L1/L2
16
12.9%
-
-
69
61.1%
85
29.7%
C
C
C
C
C
C
C
G
C
C
G
C
C
C
C
G
C
C
G
C
C
C
C
G
C
L3
22
17.7%
1
2.0%
34
30.1%
57
19.9%
∆ 82818289
12308
A
G
A
A
A
A
A
A
A
A
A
A
G
A
A
A
A
A
A
A
A
A
G
A
A
N
1
0.8%
1
2.0%
1
0.9%
3
1.0%
11251
A
A
A
A
A
A
A
A
A
A
A
G
A
A
A
A
A
A
A
A
A
A
A
A
A
N9
-
-
5
18.4%
-
-
5
1.7%
14766
T
T
T
C
C
T
T
T
T
T
T
T
T
T
T
T
T
C
T
T
T
T
T
T
T
R
3
2.4%
-
-
-
-
3
1.0%
12406
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
G
R9
-
-
2
10.2%
-
-
2
0.7%
12714
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
7
5.6%
-
-
-
-
7
2.4%
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
T
14798
13708
G
G
G
G
G
G
G
G
G
G
G
A
G
G
G
G
G
G
G
G
G
G
G
G
G
U
7
5.6%
1
2.0%
-
-
8
2.8%
-
-
-
2
0.7%
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
G
A
-
A
G
A
1.6%
15607
4580
G
7028
A
A
A
G
G
G
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
L1/L2
U
N
H
H
H
C
L1/L2
L1/L2
L1/L2
B
J
U
C
C
B
L1/L2
H/V
B
A
X
L3
U
L1/L2
L1/L2
∆82818289 G
∆82818289 G
Haplogroups
REFERENCES
V
2
Total of
Individuals
124
100%
49
100%
113
100%
286
Haplogrup
Obs.
∆82818289 G
The coding region SNPs were employed in a forensic case in which it was necessary to establish which family, out of more than 20 families analyzed, was related to considerably degraded skeletal
remains. Previous attempts at autosomal STR analysis had proved unsuccessful, although some autosomal SNPs has been typed. A strategy was developed in which it was decided to narrow down
possible related families by using the mtDNA coding region SNPs as an initial step (Table 2). Then, with a reduced subset of individuals as possible relatives, autosomal SNPs were used to identify the
related family and close the case. By this method less time was expended and the risk of false inclusions was reduced.
REFERENCES
1.
2.
3.
4.
5.
Podini D et al. (2009) SNP genotyping using multiplex single base primer extention assays. Methods Mol. Biol. 578, 379-391.
Alvarez-Iglesias V et al. (2007) Coding region mitochondrial DNA SNPs: Targeting East Asian and Native American Haplogroups. Forensic Sci. Int. Genet. 1, 44-55.
Grignani P et al.(2009) Multiplex mtDNA coding region SNP assays for molecular dissection of haplogroups U/K and J/T. Forensic Sci. Int. Genet. 4, 21-25.
Köhnemann S et al. (2009) An economical mtDNA SNP assay detecting different mitochondrial haplogroups identical HVR1 samples of Caucasian ancestry. Mitochondrian 9, 370-375.
Paneto G et al. (2011) A single multiplex PCR and SNaPshot minisequencing reaction of 42 SNPs to classify admixture populations into mitochondrial DNA haplogroups. Mitochondrian 11, 296-302.
Apoio Financeiro: