This document discusses functional genomics, which studies how genes and intergenic regions contribute to biological processes on a genome-wide scale. It describes techniques used at the DNA, RNA, and protein levels, including genetic interaction mapping, ChIP-seq, DNase-seq, FAIRE-seq, ATAC-seq, microarrays, SAGE, RNA-seq, yeast two-hybrid systems, and affinity purification mass spectrometry. Major projects like ENCODE and GTEx aim to identify all functional elements in genomic DNA and understand the role of genetic variation in gene expression across tissues.

1. Functional Genomics
Seminar
By
Farah Kayed Al-Hyari
Supervisor
Dr. Mohammad Shatnawi

2. What is Functional Genomics?
 Study of how genes and intergenic regions of the
genome contribute to different biological processes.
 Studies genes (locus) on a “genome- wide” scale.
 To determine how the individual components of a
biological system work together to produce a particular
phenotype from gene by transcription and translation
 Dynamics expression of gene products in a specific
context (protein function)

4. Gene: is a section of DNA have sequence of
nucleotide (A,T,C,G)

5. Design by ahmed aljammal

10. Goals of functional Genomics
1. To understand the function of genes and protein
2. To study biochemical ,cellular , physiological
properties of each gene product.
3. To study special(site) and temporal genetic
variation on chromosome
4. Determination the DNA sequence of each gene

12. Techniques used in Functional Genomics
 At the DNA level :
1- Genetic interaction mapping
 is a powerful technique to systematically reveal functional
relationships between genes which often also reveal the
presence of a physical interaction
 GI mapping involves the pairwise perturbation of genes
(e.g. knockout, knockdown or overexpression) in order to
elucidate how one gene modulates the phenotype of the other
 GI mapping is often used to uncover new functions of genes,
enabling a hierarchical organization of gene products into
functional complexes and pathways

14. 2-DNA/protein interaction (chip-sequencing)
 is a powerful method for identifying genome-wide DNA
binding sites for transcription factors and other proteins
(purified, and sequenced).
 The application of next-generation sequencing (NGS) to
ChIP has revealed insights into gene regulation events that
play a role in various diseases and biological pathways, such
as development and cancer progression.
 ChIP-Seq enables thorough examination of the interactions
between proteins and nucleic acids on a genome-wide
scale.

16. 3-DNA accessibility assay:
A) DNase-seq:
- DNase-seq identifies open
regions of chromatin
- DNase-seq relies upon preferential
digestion of regions of chromatin
that are unprotected by bound
proteins
- leaving behind accessible regions
that are known
as DNase I hypersensitive sites
(DHSs)

17. B) FAIRE-seq
- FAIRE-seq is dependent on
crosslinking of chromatin-
interacting proteins to DNA using
formaldehyde
- Chromatin is then sheared and
regions that are unbound by
proteins (e.g., histones) remain in
the aqueous layer of a phenol-
chloroform extraction
- While crosslinked DNA remains
in the organic layer

18. C) ATAC-seq
- ATAC-seq relies on the
hyperactive Tn5 transposase to
insert sequencing
adapters at accessible regions of
the genome.
- Following transposition
genomic DNA can be isolated
and amplified by PCR
- Then subjected to deep
sequencing.

20.  At the RNA level :
1- Microarrays:
 Is one of the most recent advances being used for cancer
research it provides assistance in pharmacological approach
to treat various diseases including oral lesions.
 Microarray helps in analyzing large amount of samples
which have either been recorded previously or new samples
it even helps to test the incidence of a particular marker in
tumors.
 Microarray analysis is a well-known method for studying
the expression profiles of protein-coding genes it has also
been adopted to detect the expression of small RNA genes

24. 2-SAGE (Serial Analysis of Gene Expression)
 is a powerful tool which provides quantitative
and comprehensive expression profile of genes
in a given cell population.
 It works by isolating short fragments of genetic
information from the expressed genes that are
present in the cell being studied
 Produces a snapshot of the mRNA population
in the sample of interest

26. 3-RNA sequencing
 Is a standard technique for transcript discovery and differential
gene expression analysis in life science laboratories
 This powerful technique is now within the reach of most
scientists thanks to innovations in next generation sequencing
(NGS) technologies which have dramatically lowered the cost of
sequencing.
 One of the most common uses of RNA-seq is to determine how
gene expression changes in response to disease pathologies,
therapeutic intervention, or other stimuli.
 Differential gene expression (DGE) analysis compares different
experimental samples and uses toolkits to evaluate if the
observed difference between normalized read counts of a gene is
statistically significant

29.  At the protein level :
1- Yeast two-hybrid system
 The yeast 2-hybrid (Y2H) assay is a well-established
technique to detect protein-protein interactions.
 This is an extremely powerful tool for researchers and is
often used alongside one or two other methods to examine
the multitude of interactions that take place in cells.

30. PRINCIPLES
1.Y2H assay relies on the expression of a reporter gene (such as
lacZ or GFP), which is activated by the binding of a particular
transcription factor.
2.The transcription factor is comprised of a DNA-binding
domain (BD) and an activation domain (AD).
3.The query protein of interest fused with the BD is known as
the Bait, and the protein library fused with the AD is referred to
as the Prey.
4.In order to activate the reporter gene expression a
transcriptional unit must be present at the gene locus which is
only possible if Bait and Prey interact.

32. THEORY
 Expression of a reporter gene requires the binding
of a transcription factor which normally consists
of two functionally and structurally independent
domains: DNA-binding (DB) and activation (AD)
domains
 The DB domain binds to the particular DNA
sequence upstream of the reporter gene, while the
AD domain activates reporter gene expression

33. 2-Affinity purification and mass spectrometry
(Ap/MS)
• Is a highly effective method for isolating and identifying
binding partners to a target protein.
• One of the most common methods used for AP-MS
experiments is expressing the target protein with a unique
peptide sequence tag.

34. Projects on Functional Genomics
1- The ENCODE project – The ENCODE (Encyclopedia
of DNA elements)
Project is an in-depth analysis of the human genome whose
goal is to identify all the functional elements of genomics
DNA , in both coding and noncoding regions.
2-The Genotype –Tissue Expression (GTEx) project
Human genetics project aimed at understanding the role of
genetic variation in shaping variation in the transcriptome
across tissue .

36. Conclusion
1. By reaching the structure of the gene, we were able to solve many
problems, the most important of which are diseases and their
treatment
2. Also through these techniques some taxonomy researchers were
able to distinguish between cell types and their physiological
behavior
3. Through the development of these technologies scientists have been
able to find simpler ways than before to understand biotechnologies
and transfer genes to target cells
4. Also one of the most important of these technologies is microarray
which is a very important technique used in pharmacology and
medicine to know the response of cancer cells to treatment