BIOCHEMISTRY o Australia, Belgium, Canada, Denmark,
Human Genome Project Germany, Israel, Italy, Netherlands,
Dr. Guerrero
AFCG, REM, SMB
GENOME
 Organism’s complete set of DNA
 DNA – paired strands
o Each strand is made of four nucleotide
bases
o Bases: Adenine (A), Thymine (T),
guanine (G) and cytosine (C)
 A-T, C-G
I. Informative Data
HUMAN GENOME
 Comprised of 23 pairs of chromosomes
o 22 autosomes and 1 sex chromosome
 Smallest human chromosome: Y, 50M bp
 Largest Human Chromosome: 1, 250M bp
 Karyotype: analysis of chromosomes via
microscope based on shape (size and banding
pattern)
II. What is the Human Genome Project?
GOALS
 Identify the approximate 100,000 genes in
human DNA
 Determine the sequences of the more than 3 B
bases that make up human DNA
 Store information in database
 Develop tools for data analysis
 Address ethical, legal and social issues that
arise from genome research
HUMAN GENOME PROJECT
 International effort to determine the sequence
of the human genome and identify the genes
that it contains
 Ran from 1990 - 2003
 US government project coordinated by DOE
(Department of Energy) and NIH (National
Institutes of Health)
 Budget: 3 billion dollars
 15-year effort
 International consortium
o US, France, UK, Japan
Human Genome Project | Bacolor, Carag, Miguel
Russia, Sweden, China
MAJOR INTERNATIONAL ORGANIZATIONS INVOLVED
 HUGO (Human Genome Organization)
 EC (European Council)
 UNESCO
III. Deciphering the Human Genome
EXPERIMENTAL PROCEDURES
a) Restriction Fragment Polymorphism (RFLP)
o Identification of sequence variations in
DNA sites that can be cleaved by
restriction enzymes
b) Pulsed-field Gel Electrophoresis
o Laboratory technique used by scientists
to produce DNA fingerprint for bacterial
isolates
o PFGE Steps:
1. Bacterial cells from an agar
plate
2. Mixes bacterial cells with
melted agarose and pours into
a plug mold
3. Bacterial cells are lysed so DNA
is free
4. Loads DNA gelatin plug into a
gel and into an electric field
that separates DNA fragments
5. Stained so DNA can be seen
under UV light
c) PCR (Polymerase Chain Reaction)
o In vitro amplification of specific nucleic
acids
o PCR Markers
 Based on short, repetitive DNA
sequences widely distributed in
the human genome
a) Yeast Artificial Chromosome (YAC)
o Genetically engineered chromosomes
derived from the DNA of yeast
o To isolate and propagate very large
segments of DNA in a yeast host
b) Sequence Tagged Site
o Short unique DNA sequence that can be
amplified by PCR (easily detected)
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 c) Positional Cloning
o Markers are used for gene hunts
o Once the gene is located, physical maps
are used to obtained flanking DNA
segments for further detailed study
(mostly pertains to regulation of gene
function)
o Identifying gene inflicting a disease
(inheritable disease)
IV. Mapping and Sequencing the Human Genome
MAPPING
 Divide each chromosome into small segments
 Arranging them sequentially on the
chromosome
GENETIC MAP
 Depicts the order by which genes are arranged
along a chromosome
 Sequence (genetic map) is facilitated by known
markers: genes or other DNA stretches
 Distances between markers are measured in
centimorgans (cM)
o Unit of measure of recombination
frequency
o 1 genetic cM is about 1 M base pairs
(bp) (on physical distance)
 Assisted in chromosomal location of several
inherited diseases (Sickle Cell disease, cystic
fibrosis, Tay-Sachs disease, Fragile X syndrome,
Myotonic dystrophy, ataxia telangiectasia)
GENE LINKAGE MAP
 Shows the relative location of a specific DNA
marker along the chromosome
 Constructed by observing how frequently two
markers are ‘inherited’ together
 The closer the markers are to one another on
the same chromosome, the more tightly linked
they are, the more likely they will be passed to
the next generation
PHYSICAL MAP
 Shows actual sites of genes on the genome
 Comprised of landmarks, (restriction enzymes
and STS) providing reference points relative to
which DNA sequence such as genes can be
localized
 Restriction Fragment Length Polymorphism
(RFLP)
Human Genome Project | Bacolor, Carag, Miguel
o Sequence variations in DNA sites that
can be cleaved by restriction enzymes
 Short Tandem Repeat Polymorphism (STRP)
o Advantages:
 Repeated up to thousands of
times throughout the genome
 Even distribution throughout
the genome
 Amplifiable by PCR
 Number of repeats vary among
individuals
V. Mapping Methods
MACRO-RESTRICTION MAPS
a) Top-down mapping:
o Fragmenting chromosomes with a rare
restriction enzyme into large pieces ->
ordered -> subdivided -> mapped
o Smaller pieced mapped together
o Result: more continuity and less gap
than the contig method, but it has a
lower map resolution
b) Contig Map (Bottom-up mapping)
o Cutting a chromosome into small
pieces, each cloned and ordered,
forming contiguous DNA blocks
c) STS-Content Mapping
o Provides the means to establish these
overlaps between each clone and its
nearest neighbors
o If 2 clones share even a single STS, they
can reliably be assumed to overlap.
d) Radiation Hybrid Mapping
o Involves fragmentation of
chromosomes in cultured cells with high
doses of X-rays -> Incorporation of
fragments into stable cell lines
VI. Terminologies
CONTIG (CONTIGUOUS)
 Set of overlapping DNA segments that together
represent a consensus region of DNA
 Organized set of DNA clones that collectively
provide redundant cloned coverage of a region
too long to clone in one piece
COSMID
 Often used as a cloning vector in genetic
engineering
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  Can be used to build genomic libraries
 Designed for cloning fragments of DNA (20k-40K
base pairs)
FISH (FLUORESCENCE IN SITU HYBRIDIZATION)
 Physical mapping technique employing
fluorescein-labelled DNA probes that can detect
segments of the human genome by DNA
GEN BANK
 Public database of DNA sequence operated by
NIH
 Accessible freely and without restrictions to all
scientists in industry and academe
I. Gene Therapy
 The prime benefit to be derived from the Human
Genome Project, in which
 Defective genes inflicting congenital diseases are
replaced by functional genes
 Genes are likely to be the key targets for new types
of therapy (eg. if a gene is mutated so it is
permanently active, gene therapy could inactivate
the protein, or if a gene has been turned back on or
a second, normal copy is introduced)
II. Benefits Of Human Genome Project
MEDICAL BENEFITS
 Improved diagnosis of disease
 Earlier detection of predispositions to disease
 Rational drugs design
 Gene therapy and control systems for drugs
 Pharmacogenomics “personal drugs”
 Organ replacement
MICROBIAL GENOME RESEARCH
 New energy sources (biofuels)
 Environmental monitoring to detect pollutants
 Protection from biological and chemical warfare
 Safe, efficient toxic waste clean up
DNA FORENSICS
 Identify potential suspects at crime scenes
 exonerate wrongly accused persons
 identify crime and catastrophe victims
 establish paternity and other family relations
 detect bacteria and other organisms that may
pollute air, water, soil, and food
 match organ donors with recipients in transplant
programs
Human Genome Project | Bacolor, Carag, Miguel
 determine pedigree for seed or livestock breeds
AGRICULTURE AND LIVESTOCK
 disease, insect, and drought-resistant crops
 healthier, more productive, disease-resistant
farm animals
 more nutritious produce
 bio-pesticides
 edible vaccines incorporated into food products
 new environmental cleanup uses for plants like
tobacco
EVOLUTION AND HUMAN MIGRATION
 study migration of different population groups
based on female genetic inheritance
 study mutations on the evolutionarily stable Y
chromosome to trace lineage and migration
 compare breakpoints in the evolution of
mutations with ages of populations and historical
events
RISK ASSESSMENT
 assess health damage and risks caused by
radiation exposure, including low-dose exposures
 assess health damage and risks caused by
exposure to mutagenic chemicals and cancer-
causing toxins
 reduce the likelihood of heritable mutations
III. More Information (FYIs)
 The Human Genome Project assembles 12,000
bases every minute
 15 billion raw base pairs were sequenced to
reach the two billion milestone
 Each area of a chromosome is sequenced at least
four to five times to ensure that the data
deposited into the GenBank is accurate
 2 billion of the 3 billion “letters” that constitute
the genetic instruction book of humans have
been deciphered and deposited into GenBank
 As per 2003’s technology, the Human Genome
Project is as complete as it can be.
 Small gaps that are unrecoverable in any current
sequencing method remain.
 New technologies will have to be invented to
obtain the sequence of these regions.
 Even though the Human Genome Project has
been completed, scientists continue to develop
and apply new technologies to the few
remaining refractory problems.
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  Continue to support a wide range of research to
develop new sequencing technologies, to
interpret the human sequence and to use the
newfound understanding of the human genome
to improve human health
 About 100 cancer genes have been already been
found, most of which are from rare leukemia and
lymphomas (<10% of all human cancer)
 Common adult cancers (breast, colon, prostate,
lung and ovary) which account for 80% of the
cancer burden -> only about 30 genes are known.

 REFERENCES
 PowerPoint presentation of Dra. Charisse
Guerrero
 Human Genome Project FAQs

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