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We are fascinated with the question of human origin.
Palaeontology and archaeology are the branches that traditionally deal with the reconstruction of human origin and history.
Molecular biology is contributing to this area through DNA analysis.
HUMAN GENOME DIVERSITY PROJECT
HUMAN GENOME DIVERSITY PROJECT SEEKS TO UNDERSTANDTHE GENETIC RELATIONSHIP BETWEEN DIFFERENT POPULATIONSBY USING STANDARD DNA MARKERS LIKE
mtDNA
“Y” CHROMOSOME
HGDP IN INDIA- CCMB IS ONE OF THE PIONEER INSTITUTE WHICH IS ANALYSING PRIMITIVE TRIBES AND CASTE AT MOLECULAR LEVEL. AROUND 6000 BLOOD SAMPLES OF DIFFERENTPOPULATIONS HAVE BEEN ANALYSED FOR THEIR GENETIC DIVERSITY
TWO HYPOTHESES REGARDING EVOLUTION OF MAN
OUT OF AFRICA THEORY
MULTI REGIONAL THEORY
MITOCHONDRIAL DNA IS COMMONLY USED IN THE CONSTRUCTION OF EVOLUTIONARY TREES.
EVOLUTION OF MAN
MITOCHONDRIAL DNA WAS FIRST RECORDED BY
ELECTRON MICROSCOPY IN 1963 BY NASS & NASS
IT WAS ISOLATED FROM YEAST MITOCHONDRIA
BY SCHATZ
IN 1981 THE MITOCHONDRIAL GENOME WAS COMPLETELY
SEQUENCED BY ANDERSON et al.
mtDNA IS MATERNALLY INHERITED [GILES ET AL 1980]
mtDNA IS A SEMI-AUTONOMOUS MOLECULE, WHICH DOES NOT
UNDERGO RECOMBINATION.
CIRCULAR MOLECULE OF 16569 BP LENGTH
37 GENES
13 CODING PROTEINS
2 rRNAS, 22 tRNAS
THE ONLY NON-CODING REGION IN THE MTDNA IS
SITUATED BETWEEN THE GENES CODING FOR
tRNA-PRO AND t-RNA-PHE.
IT IS ABOUT 1120bp IN HUMANS
MITOCHONDRIAL DNA MAP
THE CONTROL REGION IS HIGHLY STRUCTURED,
WITH A CONSERVED CENTRAL REGION (CCR)
FLANKED BY TWO HIGHLY DIVERGENT
PERIPHERAL DOMAINS.
IN HUMANS THE TWO PERIPHERAL REGIONS ARE
CALLED HYPER VARIABLE REGIONS ONE AND TWO (HVR-I
AND HVR-II).
THE CONTROL REGION IS RELATIVELY TOLERANT OF
HIGH MUTATION RATE,
THE MT GENOME IS INHERITED INTACT OVER
THOUSANDS OF GENERATIONS, WITHOUT THE
CONFOUNDING EFFECT OF CROSSOVER WITH A
PATERNAL CHROMOSOME.
THESE FEATURES OF mtDNA HAVE BEEN EXPLOITED FOR
EVOLUTIONARY AND POPULATION GENETIC STUDIES
MAKING IT A SUITABLE CANDIDATE FOR THE STUDY.
THIS PROJECT AIMS TO FIND GENOMIC DIVERSITY
WITHIN AND BETWEEN ENDOGAMOUS POPULATIONS BY
EVOLUTION USING A STANDARD DNA MARKER –mtDNA
ISOLATION OF DNA
QUANTIFICATION OF DNA
DILUTING THE DNA SAMPLES
AGAROSE GEL ELECTROPHORESIS
POLYMERASE CHAIN REACTION
PCR FOR 9bp
PCR Reaction Mixture Requirement
PCR BufferMgCl2
dNTPsPrimer APrimer BTaq PolymeraseDDWDNA (10ng/μl)
Volume(μl)
10.60.81.01.00.5
1.14
PCR Conditions
Steps
12345678
Conditions
950C for 5min950C for 1min550C for 1min720C for 1minGo to 2, 35 times720C for 5min40C for everEnd
THE PRODUCTS WERE THEN RESOLVED IN 6% POLYACRYLAMIDEGEL .
NO DELETIONS AND INSERTIONS WERE OBSERVED.
POLYMERASE CHAIN REACTION
HVR I PRIMER
PCR Reaction Mixture
Requirement Volume(μl)
PCR BufferMgCl2
dNTPsPrimer FPrimer RTaq PolymeraseDDWDNA (10ng/μl)
10.60.350.20.20.53.154
PCR Conditions
Steps Conditions
12345678
940C for 1min940C for 30sec600C for 45sec720C for 2 min30secGo to 2, 34 times720C for 7min40C for everEnd
SEQUENCING PCR
THESE PCR PRODUCTS ARE SEQUENCED IN ABI PRISM 3700 AUTOMATED DNA ANALYSER
Requirement
BigDye™Primer FDDWDNA (10ng/μl)
Volume (μl)
1.80.12.11
PCR Reaction Mixture PCR Conditions
Steps
123
4
Conditions
950C for 10sec500C for 5sec600C for4min, 30cycles40C for ever & end
THE SEQUENCES WERE ASSEMBLED IN AUTOMATED DNA SEQUENCER
ANALYSIS OF FLOURESCENT LABELLED DNA MOLECULES. THIS INSTRUMENT COMPRISES OF A ELECTROPHORETIC APPARATUS WITH A 5000 V POWER SUPPLY.
ARGON ION LASER.
CCD CAMERA.
MACINTOSH COMPUTER WITH SOFTWARE SUCH AS AUTO ASSEMBLER. THE SEQUENCES WERE ASSEMBLED AND COMPARED WITH ANDERSON mt DNA SEQUENCE AND MUTATION SITES WERE NOTED.
NOTE ALL THE MUTATION SITES AFTER ASSEMBLING THE DNA SEQUENCES WITH THE ANDERSON SEQUENCE .
MJ NETWORK ,THE NETWORK WAS DRAWN USING HVR 1 SEQUENCES THROUGH NETWORKING 3.0 VERSION SOFTWARE OF FLUXUS ENGINEERING.
A TOTAL OF 15 HAPLOTYPES WERE OBTAINED . OUT OF 15 ,10 ARE UNIQUE AND REST FORM COMMON SETS OF HAPLOTYPES.
MJ NET WORK
MEDIAN JOINING NETWORK OF CHENCHU TRIBE
NJ NET WORK
NEIGHBOUR JOINING TREE WAS CONSTRUCTED WITH THE DATA FROM 44 CHENCHU INDIVIDUALS WITH THE DATA FROM THE INDIVIDUALS OF OTHER INDIAN POPULATIONSUSING CLUSTAL X, ARLEQUIN ,PHYLIP SOFTWARES .
THE CHENCHU POPULATION HAS CLOSE RELATOION WITH DUNGRI BHILS ,DUNGRI GARASIAS, IRULA NILGIRIS
NJ NETWORK OF CHENCHU TRIBE