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  • Human population phylogenetic studies using mithochondrial DNA

    Dr Rym KEFIMIGOD- Institut Pasteur -Tunis

  • Plan: I- Introduction

    II- Example: Phylogenetic and Neandertal enigma.

    II- Example I: Mitochondrial DNA diversity of the prehistoric population from Taforalt (12,000 years- Morocco).

  • The main aim of Human population genetics is to find answers concerning: Introduction

    Differentiation in single population (Bertranpetit et al 1995; Ann Hum Genet)

  • the migration patterns in certain geographic areas.

  • human evolution, and the spread of modern humans (Richards et al 1996, Am J Hum Genet )

  • Mitochondrial DNAan important tool for Human population genetics

    Mitochondrial dna is an important tool for Human population genetics because of such features as its maternal inheritance and absence of recombination. Consequently a substantial number of mt DNA mutations have accumulated sequentially along radiating maternal lineages that have diverged as human populations colonized different geographical regions of the world. Another virtue of mtDna is that a high mutation rate compared to nuclear dna, allows the genealogy to be captured in a fair amount of details.The most variable region of the mtDNA is the 1122 bp non coding control region called D- Loop. This region is located between 16024 and 576 bp. The variation is concentrated in two fragments: HVS1 and HVS2

  • The studies of mtDNA polymorphism in human populations were based:

    initially on restriction enzyme (RFLP) analysis :

    Low resolution restriction and high resolution restriction mapping (Horai et al 1984, Johnson et al 1983, Horai et al 1984, Cann et al 1987, Torroni et al 1993 ) ,

    Brown and Wallace in 1970: Pioneers in mtDNA investigation Mitochondrial DNA Studies history:

    Research on Human mtDNA as a molecular marker was pioneerd by Wesley Brouwn and Douglas Wallace in the late 1970 The studies of mtDNA polymorphism in human populations were based: initially on restriction enzyme (RFLP) analysis : Low resolution restriction uses 5 to 6 enzymes whereas high resolution restriction mapping uses at least 12 enzymes

  • on combined method using sequence analysis and restriction mapping (Bertranpetit et al 1995, Vigilant et al 1991, Richards et al 1996, Richards et al 1998, Macaulay 1999, Torroni et al 2001, Maca-Meyer et al 2001, Salas et al 2002)

    on sequence analysis of the mtDNA control region

  • Mitochondrial Eve Cann et al; Nature 1987 147 individuals from five geographic populations: Europe, Africa, Asia, Australia, New Guinea

    have been analysed by high-resolution restriction mapping

    Accordingly, Cann and collaborators analysed>>>>>, They noted that Subsaharan african individuals presents the most variable mtdna sequences. Moroover, a parsimonious tree constructed with 133 sequences displays 2 clusters. The basical cluster is composed exclusively of subsaharn african haplotype wheras the second cluster groups sequences from the five geographicals areas. Authors concluded that, mtDNA have been diverged from an ancestral lineage originated in Africa. Cann et al called this hypothesis the : Mitochondrial Eve

  • Different mtDNA lineages have been diverged from an ancestral women originated in Africa. Mitochondrial Eve

    Cann et al postulated that

  • RFLPs studies of mtDNA from a wide range of Human populations have revealed a number of stable polymorphic sites in the mtDNA coding region . Mutations observed in both mtDNA coding region and control region in modern human populations have occurred on these pre-existing haplogroups

    Define the individual mtDNA type or haplotypes Define related groups of mtDNA called haplogroups

    RFLPs studies of mtDNA from a wide range of Human populations have revealed a number of stable polymorphic sites in the mtDNA coding region . These define related groups of mtdna s called haplogroups. Most of the mutations observed in both mtDNA and control region in modern human populations have occurred on these pre-existing haplogroupes and define the individual mtdna type or haplotypes

  • Alignment of sequences with mtDNA reference (CRS) using Blast 2 sequences

    Haplotype and Haplogroup+ RFLP analysis

    alignement of maktar sequence with CRS using blast 2 sequence were performed in the aim to define haplotype and haplogroups

  • Examples :

    Individual 1:

  • Mitochondrial haplogroups

    The majority of haplogroups have been shown to be continent specific. L1 , L2 and L3 group sub- Saharan African lineages. H, I, J, K T U V W and X. encompass almost all mtDNA from European north African and Western Asian Caucasian . Finally Haplogroups A, B, C, E, F , G and M embrace the majority of the lineages described for Asia, Oceania and native Americans.

  • The phylogenetic relationships between haplotypes were inferred in the first studies by

    Maximum parsimony tree

    Then by Neighbor- joining trees

    Later by Median joining network.

    Trees were based on distance data calculated from

    Nucleotide sequence or RFLPs data or Nucleotide sequence combined with RFLPs data.

    The phylogenetic relationships between haplotypes were inferred in the first studies by Maximum parsimony tree and than by neighbor- joining trees and later by median joining network.

  • Macaulay et al, 1999; , Am J Hum Genet,

    This neighbour- joining tree shows the relationship between different haplogroups. Numbers indicate mutation characteristic of haplogroup, number between parenthesis indicate the RFLP analysis. This tree is rooted with Neandertal sequence We note that African Haplogroups ( L1, L2 and L3) occupy the base of the tree and they are the most ancient haplogroups. This results reinforce the postulate of Mit eve

  • Dr Rym KEFI and Dr Eliane BERAUD-COLOMB

    U600 INSERM-FRE2059 CNRS Laboratoire d'Immunologie, Hpital de Sainte-Marguerite- Marseille- FranceExample I: Mitochondrial DNA diversity of the prehistoric population from Taforalt (12,000 years- Morocco).

  • Knowledge of the settlement of Northern Africa regionStudy of molecular diversity of modern Human populationsStudy of archaeological specimens and their environment

    La connaissance du peuplement du Nord de l'Afrique se construit suivant deux approches: l'une travers l'tude des spcimens archologiques et de leur environnement, l'autre travers l'tude de la diversit molculaire des diffrents marqueurs gntiques ports par les individus composant les populations actuelles

  • Anthropologic dataTransition from Homo erectus towards Homo sapiens archaic

    The oldest human presence in North Africa is dated of 700.000 years BP and corresponds to the osseous remainders of Atlanthropus maurtanicus. discovered in Ternifine in Algeria. The specimens of Sidi Abdderrahman sites could shouw the transition from Homo erectus towards Homo sapiens sapiens. Around 40.000 years, the appearance of Homo sapiens sapiens was associated with the developement of an original industry : Aterian, specific of North Africa.

  • Ibero-Maurusian manIbero-Maurusian industry

  • The main Ibero- Maurusian site are Taforalt in Morocco, Columnata and Afalou in Algeria

  • Ibero-maurusian man1-Europian origin? (Vallois 1969, Ferembach 1985) 2- Near East origin ? (Vandermeersch 1978)4- North African origin ? (Camps 1989, Dutour 1995)3- Subsaharian origin? (Ferembach 1976)Origins ???

    the origin of the ibero-maurusians is a multi-field debate: four assumptions were proposed

  • Ancestral indigenous component: U6- (Paleolithic: 45.000 years)

    Eurasiatic component: T, H, U, J(Neolithic?: 9000 years)

    Sub-Saharan component : L (Historic?)

    Former studies using Mitochondrial DNA (Crte-Real et al. 1996; Rando et al. 1998; Comas et al. 2000; Brakez et al. 2001; Esteban et al. 2004) showed that the genetic structure of North Africa is composed of 3 components: Genetic data

    Concerning the Tunisian population, only fewstudies relating to the analysis of mitochondrial diversity of 155 Berber and 47 non Berber, were recently published (Plaza et al. 2003, Fadhlaoui-Zid et al. 2004). Represented by U6 haplogroup old of 45.000 years . Eurasiatic component include haplogroup like T, H, U . Old probably of 9000 corresponding to Neolithic period . And finally subsaharan component represented by L haplogroup appeared in Norh Africa probably since historic period

  • to contribute to the knowledge of North Africa settlement

    We proposed to analyse the mitochondrial DNA diversity of the prehistoric population from Taforalt (13,000 years BP- Morocco). Aim:

    In the aim to contribute to the knowledge of North Africa settlement

  • The cave of Taforalt in Morocco The cave of Taforalt is Located at 55 km in the North-West of Oujda

  • Ancient DNA was extracted from 31 bone remains from Taforalt

    Phenol/ChloroformeExtractionDissolution of bone powders ADNHypervariable segment 1 (HVS1) of control region (D-Loop) was amplified by PCR and sequenced (R. Kefi et al; C.R.Palvol 2003)

    La manipulation de lAND a necessite des precautions drastiques de laboratoires

  • Mitochondrial DNA diversity of Taforalt population Genetic structure of Taforalt:Eurasiatic Component :

    H, U, JT, V: 90,5%North African component:

    U6: 9,5%In modern Human population, JT is presents only in: 1,6% Berbers from the North of Morocco 1,8% of Sicilians, 1,6% of Italians.

    Feuil1

    spcimensdbut et finPolymorphismesHaplogroupes

    de la squence

    Taf I16054-1645