P-151
Laser
microdissection and pressure catapulting with PALM® to assist typing
of target DNA in dirt samples
Lambie-Anoruo BL1,2, Prince DV1, Koukoulas I1, Howells DW3, Mitchell RJ2, van Oorschot RAH1
1Victoria Police Forensic Services
Department, Victoria
3085, Australia
2Department of Genetics and Human Variation, La Trobe University, Victoria
3086, Australia
3Austin Hospital, Department of Medicine, University
of Melbourne, Victoria
3084, Australia
Obtaining a DNA profile from a subset of cells within
a mixture of cells where the predominant cells are of a different type and
source becomes problematic once the proportion of the target cells becomes very
low. This can be difficult even when the total number of minority cells is
theoretically sufficient to generate a good DNA profile. Differential extraction
methods to separate sperm from epithelial cells are commonly used to assist
with mixtures of such cell types (as frequently encountered in sexual assault
cases). These methods, however, are inadequate when dealing with mixtures of
other cell types, such as saliva and blood, or saliva and shed skin cells. It
can also be troublesome to retrieve profiles from small biological samples in
debris such as saliva in dirt. Use of the PALM laser microdissection and
pressure catapulting process may assist in the retrieval of target DNA and
subsequent DNA profiling in these situations.
We tested the capability of PALM to isolate saliva
cells (12µl saliva) from mixtures with dirt (8µl of humus rich dirt). DNA was
extracted from replicate samples using Chelex and organic extraction methods
and compared to DNA retrieved from cells isolated from replicate mixtures using
PALM isolation followed by Chelex extraction. Each test was repeated four times
on mixtures dried for one day and on mixtures dried for seven days. Extracted
DNA was quantified using the Quantifiler™ kit and when found to be positive
also amplified and typed with Profiler Plus™ using an ABI PRISM®
3100 Genetic Analyser in conjunction with GeneMapper™ ID.
Results from the one-day-old series of samples
demonstrated that typeable DNA from the saliva component of the dirt sample was
not retrievable from the samples extracted using either of the standard Chelex
or organic extraction methods. In contrast, PALM assisted isolates of 200
saliva derived cells from amongst the dirt of one-day-old samples provided DNA
from which the expected full DNA profiles were generated. The DNA extracted
from the seven-day-old series of samples, using either the Chelex or the
organic method, was also not typeable. The seven-day-old sample examined using
PALM revealed that there were fewer whole cells observable and that the
retrieval of recognisable cells took significantly longer. The 47 cells
(representing a portion of the total available cells) that were isolated from
the seven-day-old sample provided a partial profile.
The use of PALM should be considered to identify and
isolate target cells from debris that may prevent the generation of DNA
profiles using standard DNA extraction methods.
P-152
Allele frequencies of
fifteen STR loci in an Italian Population
Lancia M , Coletti A ,
Margiotta G , Lottanti L , Carnevali E , Bacci M
Section
of Legal Medicine, University
of Perugia, Terni, Italy.
The study of the
STR loci is important for the creation of local human identification databases.
In the latest years, European countries have begun to plan studies whose
purpose is to create national and/or local databases and to be known with the
expression frequency of a great number of these DNA loci. Our research has the
aim of creating a local database, according to the recommendations published by the International
Society for Forensic Haemogenetics.
The study was carried out on
specimens taken from 100 healthy and not related individuals, who were born in Terni and have been
living there for at least two generations. The biological material consisted of
blood in 96 cases, and of oral swab in the remaining 4. Extraction of the DNA
from the blood specimens was carried out by using QIAamp DNA miniKit of the
Qiagene company.
The DNA extraction from oral
swabs was executed by the Chelex1 method.
The DNA polymorphism analysis
was carried out by enzymatic amplification (Polymerase Chain Reaction - PCR).
Allelic frequency
determination becomes very important in forensic use when you need to calculate
the probability that two DNA specimens derives from the same individual.
In our case,
after obtaining the typization of the fifteen STRs studied for the 100
specimens we analysed, we calculated the allelic frequency of every single
system. In order to verify if the considered population was in equilibrium with
an ideal one having a Gaussian-type distribution, we applied Hardy-Weinberg’s
law, Pearson’s test and p-value calculation.
In our case Terni population turned
out to be in equilibrium in all the fifteen systems we studied. The allelic
frequencies of the population were compared to the corresponding data of
Italian population in a generalized way.
P-153
DNA recovery
from semen swabs with three different extraction methods
Lazzarino
F., Laborde L. , Lojo M.M.
Laboratorio de Análisis Comparativo de ADN
Suprema Corte de Justicia, Buenos Aires, Argentina
Efficiently extraction of sperm cells from the solid
matrix is an important step in male DNA recovery from cotton swabs. Digestion
with proteinase K loosens the attachment
of semen to the solid support. Thus, digestion in the presence of the cotton
matrix enhance DNA yield. We used simulated samples in order to compare the
extraction efficiencies of Qiamp DNA
minikit, DNA IQ System and Chelex methods, performing all the extraction steps
in the presence of the solid support. Female oral swabs were embedded with
serial dilutions of semen samples of known cells density (sperm cells/ ml). The
experimental conditions were adjusted in order to use almost the same number of
target cells in all the three different protocols assayed. Standard proteinase
K digestion without DTT was performed by incubating a 1/4 of the cotton swab. Samples were centrifuged in a
spin basket and the following washes, as well as the full extraction procedures
were done in the presence of the solid support. DTT was added to both two
digestion buffers used in the Qiamp DNA minikit. Str amplicons
obtained by Profiler Plus PCR amplification were run on an ABIprism 310
Sequence Analyzer and the recovery of
peak intensities were compared. No significant differences were observed in the
extraction efficiency between Quiagen and DNA IQ systems, whereas the detection
limit (number of target cells) were higher in the extraction performed with
Chelex. A male DNA profile could be still recovered by Chelex extraction from
the swabs previously extracted with both Qiamp and DNA IQ methods, suggesting that both treatments were not able
to fully release the sperm out of the fiber.
mercedeslojo@hotmail.com
P-154
Selection of Y-STR loci and
development of a PCR multiplex reaction
for use in South Africa.
Leat N, McCabe M, Kleyn E,
Cloete K, Benjeddou M, Davison S
Biotechnology
Department, University of the Western
Cape, Cape
Town, South Africa
The objective
of the present study was to examine the properties of a set of single-copy
Y-STR loci to assess their suitability for forensic case work in three South
African populations. Three criteria were used to select markers for assessment.
Firstly, the single-copy markers of the minimal haplotype were selected based
on their established use in forensic studies. Secondly, eight markers were
selected on the basis of high gene diversity values reported for several
population studies, and thirdly 19 markers were chosen from a survey of
Y-chromosome sequence with selections made primarily on the basis of the number
of repeated elements present. Samples
were typed from English-speaking Caucasians, Xhosa individuals and Asian
Indians. Gene diversity values, the number of alleles identified and the
average stutter was determined for each locus. The data has been used to select
a subset of highly polymorphic Y-STR loci. A PCR multiplex reaction is
currently being refined and to facilitate the analysis of the selected loci in
forensic studies.
Contact: nleat@uwc.ac.za
P-155
Haplotypes
and mutations of 17 Y-STR loci from Korean father-son pairs
Hwan Young Lee1, Han Young Lee2, Ukhee Chung1, Myung-Jin Park1,
Ji-Eun Yoo1, Kyoung-Jin Shin1,3, Sang-Ho Cho1,3,
Woo-Ick Yang1
1Department of
Forensic Medicine, College
of Medicine, Yonsei University,
Seoul, Korea
2Department of Forensic
Medicine, National Institute of Scientific Investigation, Seoul, Korea
3Human Identification
Research Institute, Yonsei
University, Seoul, Korea
We have investigated 17 Y-STR loci-DYS19,
DYS385a/b, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, DYS438, DYS439,
DYS437, DYS448, DYS456, DYS458, DYS635 (Y GATA C4), Y GATA H4-in 365 father-son
pairs (6,205 meioses) of 355 families. Of 338 different haplotypes obtained
from 355 fathers, 326 haplotypes
were observed once, 10 haplotypes two times and the other two haplotypes were
observed 4 and 5 times, respectively. The overall haplotype diversity was
0.9996. In 365 father-son pairs, a total of 21 mutations were observed at 12 Y-STR
loci (DYS19, DYS385a/b, DYS389-I, DYS389-II, DYS390,
DYS393, DYS439, DYS437, DYS456, DYS458, DYS635 (Y GATA C4), Y GATA H4).
Sequence analysis for mutant alleles demonstrated 21 single step mutations: 8
gains and 13 losses. However, there was no significant surplus of gains or
losses. The locus-specific mutation rate estimates were between 0.0 and 8.2 X
10-3 and the average mutation rate estimates were 3.4 X 10-3
(95% C.I. 2.1~5.2 X 10-3) across all 17 Y-STR loci. Mutation rates
differed strongly between loci depending on the molecular structure of the
respective STR locus, and the locus-specific mutation rate estimates also
showed differences between populations. However, in contrast to the case of
autosomal STRs, no noteworthy correlation was observed between mutation rate
and the father’s age at child’s birth.
P-156
Retrieval of DNA and genetic profiles from swaps taken inside cars.
Lenz C, Flodgaard LR, Eriksen
B, Morling N.
Department of Forensic
Genetics, Institute
of Forensic Medicine, University of Copenhagen, Denmark.
In a survey of crime case samples collected from the
interior of cars in the period 2003-2004 the success rate of retrieving a
genetic profile from cotton swaps was estimated. A total of 241 samples were
analysed and DNA profiling was performed using the AmpFlSTR®-SGM-plusTM
kit with 28 cycles PCR. Only 23 %
of the samples showed a DNA concentration >0.02 ng/ml as quantified with a slot blot method.
STR profiles were retrieved for all but three of these samples (overall success
rate 22 %). The samples were collected by five different police units and the
success rate for the units varied from 14 % to 34 %. This indicated that the sampling technique
played a major role for the success rate.
In a controlled experiment, we tested if the amount of
water and the storage conditions of the swaps influenced the retrieval of DNA.
One policeman made swaps from steering wheels and spokes in 14 different cars.
All swaps were taken from delimited areas. A total of 56 samples were collected
from the cars. In 89 % of the samples, a DNA concentration > 0.04 ng/ml was retrieved (range 0.01 – 5.6 ng/ml) using quantification with the
QuantifilerTM kit. DNA
analysis was performed on 25 samples and a DNA profile was obtained for all of
them. No significant difference regarding the amount of DNA retrieved from the
swaps was seen when 1 versus 4 drops of water were used for the swaps. Also, no
significant difference was seen when swaps were air dried versus frozen. The
high success rate for the samples from the controlled experiment compared to
the crime case samples could be contributed only to the sampling technique
where the swaps were taken by thoroughly wiping a delimited area.
Contact: Camilla.Lenz@forensic.ku.dk
P-157
Tsunami 2004 – experiences, challenges and strategies
Lessig R, Thiele K, Edelmann J
Institute of Legal Medicine of the University of Leipzig, Germany
The Tsunami after the sea
quake in Southeast Asia on the 26th of December 2004
represents the largest disaster in the modern World. More than 280,000 people
in the countries around the Indian Ocean have
been reported missing. Especially Thailand and Sri Lanka as
major tourist centres demand a large number of victims from different European
countries. Twenty international Disaster Victim Identification (DVI) teams were
present in Thailand
to help identifying the recovered bodies. Such a number of different teams and
the circumstances in the area of Khao Lak were a great challenge for the
organisation. It was necessary to adapt the different teams to a common
strategy of investigations. The established international centre established
the guidelines for the forensic-medical, forensic-odontological and forensic
genetic investigations. The fast decomposition of the bodies was a great
challenge. The collection of the post mortem data was done by forensic
specialists. The guidelines for the DNA analysis request a collection of
different samples from every investigated body – two healthy teeth, rib, bone
or similar tissues – for examination. The biggest problem seems to be the
expected rapid degradation of the DNA. So the suggested strategy in such cases
should be to test samples very soon to assess the suitability for genetic
typing. After knowing the possibilities the second step would be the decision
about the best markers to be used. So a small number of samples were investigated
in our laboratory. A high level of degradation of the DNA was observed and
special procedures of extraction were necessary to get a result.
So an important conclusion for
further work in this field is an agreement on international standards and also the
training of specialists who are able to coordinate the analysis. The Tsunami
shows also that the DNA analysis can be very helpful in such mass disaster case
work as a part of the forensic fields.
P-158
HVI
and HVII Sequence Polymorphisms of the Human mtDNA in the North of Portugal:
Population Data and Maternal Lineages
Lima G1, Pontes ML1, Abrantes D1,
Cainé L1, Pereira MJ1, Matos P1, Pinheiro MF1,2
1
Instituto Nacional de Medicina Legal – Delegação do Porto
2Faculdade
de Ciências da Saúde – Universidade Fernando Pessoa
The analysis of mitochondrial
DNA (mtDNA) control region is of great importance in forensic casework. The aim
of this study was to create a population database for the HVI and HVII regions
of the mtDNA in the population of North Portugal
and to analyse these two segments in maternal relatives from this population.
For the population study, the HVI and HVII segments were analysed in unrelated
and healthy individuals, chosen randomly, from the North of Portugal. For the
maternal relatives analysis, those two regions were studied in a set of
families (mother/child, grandmother/grandchild or sibling pairs) from that
region of Portugal.
The DNA was extracted from peripheral blood and oral swab samples, using
different methods (phenol-chloroform, Chelex and Chelex + phenol-chloroform).
The HVI and HVII segments were amplified by PCR using specific primers. These
two segments were direct sequenced on both strands using the universal primers
M13 and two different sequencing kits (dRhodamine and BigDye v1.1, Applied
Biosystems). The HVI and HVII sequences were studied between positions 16033bp
– 16391bp and 57bp – 408bp, respectively. Nucleotide substitutions
(transversions and transitions) and insertions / deletions were found using Anderson’s reference
sequence. Length and position heteroplasmy were observed. The genetic structure
of the population was analysed by calculating the number of different
haplotypes, nucleotide diversity, genetic diversity and mean number of pairwise
differences. The match probability and discrimination power values were
calculated. The classification into haplogroups was also made.
Contact: Biologia@dpinml.mj.pt
P-159
Polymorphisms Analysis of Mitochondrial DNA in
Coding Area
LIU YC1，HAO JP2，TANG H1，YAN JW1，WANG J1，REN JC1
1Forensic Medical Examination
Center of Beijing Public Security
Bureau
2school of Forensic Medicine,Shanxi Medical
University
Mitochondrial DNA (mtDNA) sequencing has
allowed investigators to derive genetic informations from forensic samples
where nuclear-based analyses have failed, for example degraded samples, old
bone fragments or hair shafts without roots. Currently mtDNA for forensic
testing consists primarily of portions of the control region, most often
targeting the hypervariable regions one and two (HV1/HV2) ,but poor
discrimination power remains a problem. The only solution would appear to be to
find more polymorphic sites within mtDNA. The suggestion has been made that
besides the mtDNA control region, the polymorphisms within mtDNA coding area
should be used for forensic biologists in order to greatly increase the
discrimination power of mtDNA .
In this study, we have sequenced the mtDNA
coding area nt8162-8483 and nt13070-13299 of 100 unrelated healthy Han Chinese
individuals. We have presents the single nucleotide polymorphisms (SNP) sites
and 9-bp length-polymorphism of the mtDNA intergenic COⅡ/tRNALys region,
which may be of crucial importance to forensic testing. The lengths of the
amplicons were 322bp and 230bp respectively. There were 24 mitochondrial
haplotypes defined by 21 variable positions in both regions. The gene diversity
was estimated at 75.11%, and the probability of two randomly selected
individuals having identical mtDNA types was 25.64%.
Conclusions
The polymorphic sites within mtDNA coding area
can be useful in combination with mtDNA control region in order to increase the
discrimination power.
Contact: yachengliu@163.com
P-160
Study of microvariation of
allelic frequency distribution of 17 STR’s in each of the Azores
islands population
Lopes V1, Carvalho M1, Andrade L1,
Anjos MJ1, Serra A1, Balsa F1, Brito P1,
Oliveira C1, Batista L1, Gamero JJ2,
Corte-Real F3, Vieira DN3, Vide MC1
1Forensic Genetic Service.
National Institute of Legal Medicine. Largo da Sé Nova, 3000 Coimbra. Portugal
2Departament of Legal Medicine,
Faculty of Medicine, University of Cádiz,
Spain
3National Institute of Legal
Medicine. Largo da Sé Nova,
3000 Coimbra. Portugal
We performed a
study of the allelic frequency distribution of 17 STR’s along each of the
islands to look out for statistical differences among islands. From the global
population we selected only those individuals whose both parents were born in
the same island.
DNA was extracted
by Chelex from air-dried
blood stains of healthy and unrelated
individuals from Azores archipelago and
amplified with two commercial multiplex kits: AmpFlSTR®
Identifiler™ (Applied Biosystems) and PowerPlex® 16 System (Promega). The detection was
carried out on ABI
Prism™ 310 Genetic Analyzer with internal standards (LIZ-500 and
I.L.S. 600 respectively) and allelic ladders supplied with each kit.
The allele frequency
distribution of the seventeen STR’s present in the multiplex systems in
each of the islands is in equilibrium of Hardy-Weinberg.
The
microvariation study of the allelic frequency distribution among the islands
was performed with software Arlequin 2.000 to obtain the genetic distances
between the islands and the correspondent P values by the sum of squared size
difference (RST) method. The phylogenetic tree derives from Phylip 3.5c
software using the Neighbor-joining method.
There were no
significant differences among the islands with the exception of Flores (in the most occidental group) with some P values
not reaching 0.05.
P-161
Y-STR polymorphisms from Basque-speaking region of Cinco Villas
(Navarra) in the context of the Pyrenean genetic landscape.
López-Parra
AM1, Tavares L2, Gusmão L2, Mesa MS4,
Prata MJ2, Amorim A2,3, Arroyo-Pardo E1
1
Depto. Toxicología y Legislación Sanitaria, Facultad de Medicina, Universidad
Complutense, Madrid, Spain.
2
Instituto de Patología e Imunologia Molecular da Universidade do Porto
(IPATIMUP) Portugal.
3
Faculdade de Ciências, Universidade do Porto, Portugal.
4 Depto
Zoología y Antropología Física, Facultad de Biología, Universidad Complutense,
Madrid, Spain.
The Iberian Peninsula
presents a complex geographical landscape with mountain ranges as in Northern Spain. That enhances the genetic isolation of
small populations and consequently significant differentiation. We have studied
a set of 42 samples from the Basque-speaking region of Cinco Villas (Navarra),
located at the western part of the Pyrenees,
for a total of 15 Y-STRs (Minimum Haplotype plus DYS460, DYS461, DYS437,
DYS438, DYS439, GATA H4, GATA C4). Thirty five different haplotypes were
detected (haplotype diversity: 0.9919± 0.0069) and only seven were found in two
individuals. Data from this population were compared with those from other
Pyrenean and Iberian populations. Statistical analyses revealed that Cinco
Villas region clusters with other samples of the Basque Country and also with
other non Basque-speaking population from Pyrenees.
This fact suggests a common genetic background throughout the whole Pyrenean
mountain range or an important gene flow between these mountain populations,
irrespectively of the language presently spoken.
Contact: earroyop@med.ucm.es
P-162
Microgeographic mitochondrial DNA
patterns in the South Iberia
López-Soto
M1, Salas A 2, Sanz P 1, Carracedo A2
1 National
Institute of Toxicology and Forensic Sciences, Depart. of Seville, Ministry of Justice, Spain
2 Unidad de Genética,
Facultad de Medicina, Universidad de Santiago de Compostela, Galicia, Spain
Along history, Andalusia (South of the Iberian
Peninsula) has been a territory occupied by many civilizations
coming from Europe and North
Africa. Here we aim to identify its mitochondrial composition by
analyzing the two hypervariable regions (HVS-I and HVS-II) and selected coding
region SNPs of the mitochondrial DNA (mtDNA). A total of 419 individuals from
28 villages (belonging to different provinces and with more than 200 years of
history) have been sampled. This sampling has been design in order to uniformly
cover the geographic area of South Iberia.
Historical record indicates that these villages have experience little recent
migration. Preliminary results revealed that 94% of the haplotypes belong to
typical European haplogroups, 2.1% are Sub-Saharan lineages, and only 1.6%
North African. AMOVA analysis indicates that the main percent (97.6%) of the
variability in these populations is found between individuals, 2.2% between villages
of the same province, and 0.25% between provinces. In addition, haplotype
diversity is high (0.99) in Andalusia in
comparison with other Iberian and European populations. The results point to a
lack of significant demographic impact (at least in the maternal mtDNA side) of
North Africa despite the close geographic
proximity and eight centuries or Arabian colonization.
P-163
Multiplex STR and
mitochondrial DNA testing for paraffin embedded specimen of healthy and
malignant tissue: Interpretation issues
Lu C1,2, Budimlija ZM1, Popiolek
DA3, Illei P3,West BA3, Prinz M1
1 Office of Chief Medical
Examiner, Department of Forensic Biology, New
York, NY
2 John Jay College of Criminal
Justice, New York, NY
3 New York University School of Medicine,
Department of Pathology, New York,
NY
DNA-based short tandem repeat (STR) typing is a
powerful tool for the confirmation of suspected sample mix-ups or the presence
of contamination in histology material (1). Histology specimens also have
potential as reference samples in body identification efforts. But
microsatellite analysis of tumor DNA has shown substantial allelic instability
(2,3) which might impair correct sample associations. The aim of this study was
to compare STR typing results for healthy and malignant tissues to mtDNA data
for the same sample sets, evaluate the results and formulate interpretation
rules, for example, for the determination of Loss of Heterozygosity (LOH). Ten
different types of carcinoma were represented in the anonymous study material
(endometrial adenocarcinoma – types I and II, granulosa cell tumor,
adenosarcoma, malignant mixed Mullerian tumor, adenocarcinomas of prostate,
lung, colon and cecum and cutaneous melanomas). Healthy and carcinogenic
tissues were collected from each individual and embedded in paraffin. All
slides were set up in a double blind manner where the researchers did not know
which tissue was healthy or cancerous. After standard organic extraction,
samples were typed using the PowerPlex®16 multiplex STR system (Promega Corp., Madison, WI)
and the Linear Array Mitochondrial DNA HVI/HVII Region-Sequence Typing Kit
(Roche Applied Science, Indianapolis,
IN). Many of the tested samples
yielded partial profiles that showed characteristics of degraded DNA. Tissue
fixation and embedding have been shown to negatively affect DNA quality. DNA
degradation results in reduced peak intensity for high molecular weight alleles
and increased stochastic effects causing heterozygous peak imbalance and
allelic drop out. Several samples did display additional STR alleles and LOH.
The mtDNA assay is less affected by DNA degradation but more prone to detect
DNA contamination. Another critical issue for mtDNA testing that must be
addressed during interpretation is heteroplasmy (4). In order to distinguish
LOH from degradation based allelic drop out, the interpretation guidelines need
to incorporate signal intensity and molecular weight of affected alleles. For
RFU values below 300, LOH cannot be determined for loci > 350bp. Alleles
smaller than 350bp should still display full types and a heterozygote balance ≥
0.7. For mtDNA testing on clinical specimen mutation and heteroplasmy issues
will be difficult to establish unless the histology samples can be processed
under ultra clean conditions. Contact: PRINZ@ocme.nyc.gov
References: 1. Popiolek DA, Prinz MA, West
BA, Nazzaruolo BL, Estacio SM, Budimlija ZM (2003) American Journal of Clinical
Pathology 120:746-751
2.Vauhkonen H, Hedman M, Vauhkonen M, Kataja M, Sipponen
P, Sajantila A (2004). Forensic Science International 139:159-167.
3. Poetsch M, Petersmann A, Woenckhaus C, Proetzl C,
Dittberner T, Lignitz E, Kleist B (2004) Forensic Science International 145:1-6
4. Calloway CD, Reynolds RL, Herrin Jr GL, Anderson WW (2000)
American Journal of Human Genetics 66:1384-1397
P-164
Disparity
between self-identified ethnicity and mtDNA ancestral lineages:
a
case study in Kenyan populations
Luiselli D1,
Boattini A1, Flamigni ME1,
Castrì L1, Pettener D1
1Department of Biology, Unit of Anthropology, University of Bologna, Bologna, Italy
Genetic studies of African populations can be frequently biased by
different sampling criteria or erroneous assessment of ethnicity. In this study
we analyze mtDNA variation of Turkana, Samburu and Rendille populations, three
pastoralist nomadic ethnic groups of Kenya, with the aim of proving that
genetic data should always be associated to individual biodemographic
information. The simple use of self-identified ethnicity is often misleading.
The social structure of African Pastoralists is patrilineal and requires a
detailed reconstruction of the real marital migration patterns. In fact, in
exogamous marriages, the brides loses her ethnicity and acquires the groom’s
one, creating a disparity between the real ancestral maternal lineage and the
declared one.
The data were collected in the
Loyangalani village, district of Marsabit, and in Morijo village. Buccal swab
samples were obtained from 107 individuals, and the geographic and ethnic
origin of each subject as well as of his four grandparents was carefully
ascertained by oral interviews. All mtDNAs were subjected to sequencing of the
control-region hypervariable segment I (HVS-I), and surveyed for 13 RFLPs polymorphic markers in the coding
region.
Biodemographic results show consistent
admixture between the three ethnic groups, with different patrilineal and
matrilineal migration patterns. As concerns maternal lineages, admixture
between Turkana, Samburu and Rendille are 27%, 9% and 42% respectively. The
genetic data are analyzed both taking into account the self-identified
ethnicity and the genealogic reconstruction of real ancestral lineages up to the third generation. In the former case AMOVA (Fst=0,036;
p=0.029, 1000 iterations) shows absence of genetic homogeneity among the three
groups, while in the latter (Fst=-0,016; p=0,819, 1000 iterations) homogeneity
is high.
The striking difference in results using the two clustering criteria
suggests caution in the analysis and interpretation of mitochondrial genetic
data of African populations. When samples are collected without detailed
biodemographic information, the use of self-identified ethnicity can lead to
cultural groupings that are largely
independent from their genetic origin.
P-165
Enzyrim: a
new additive to increase the DNA yield from different materials such as teeth,
blood or saliva
Mályusz V1, Schwark
T1, Simeoni E1, Ritz-Timme S2, von Wurmb-Schwark N1
1Institute of Legal
Medicine, University
of Kiel, Kiel, Germany
2Institute of Legal
Medicine, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany
Enzyrim is an enzyme mixture normally used
for bone maceration. It is cheap, easy to handle, non-toxic and disposal is
simple. When extracting DNA from Enzyrim treated teeth we discovered that the
amount of extracted DNA was unexpectedly high.
We then systematically investigated different
biological materials using three extraction kits, the Invisorb Forensic kit,
the SPS Spin Swab kit (both Invitek, Germany) and the NucleoSpin Blood
Quick pure kit (Macherey Nagel,
Germany).
DNA was extracted from buccal swabs, dried
blood spots on filter paper, whole blood and toothpowder. All DNA extractions
were performed according to the manufacturer´s recommendations as well as after
addition of Enzyrim to the lysis step of each kit.
DNA quality and quantity was tested on ethidium
bromide stained agarose gels. Absolute quantification was done using real time
PCR. The DNA samples were also employed to genetic fingerprinting using the
Powerplex ES and the AmpFlSTRIdentifiler kit.
The application of Enzyrim greatly improves the
DNA yield from forensically important materials and does not hamper DNA amplification.
Thus Enzyrim apparently is a very useful additive for the optimisation of DNA
extraction in the forensic routine.
Tidak ada komentar:
Posting Komentar