Lecture 12

Bioteknologi Molekuler
Modern
TIM DOSEN BIOTEKNOLOGI FARMASI
2019/2020
Overview

DNA, RNA, Protein

Omics
Overview
 Perkembangan teknologi di bidang biologi memacu terjadinya
percepatan perkembangan teknologi, khususnya dalam bidang
Kesehatan
 Terpecahkannya informasi DNA dan kode genetik dan adanya
kemajuan informasi pada masa genomic menyebabkan arah
Bioteknologi menuju
 Overview
If the genomic era can be said to have a precise birth date, it was in the midst of the appearance of
the series, on April 14, 2003. That was when the international effort known as the Human Genome Project
put a close to the pregenomic era with its announcement (available at 
http://www.genome.gov/11006929. opens in new tab) that it had achieved the last of the project's original
The extent and pace of progress in
goals, the complete sequencing of the human genome.
genomics are suggested by the fact that this achievement occurred 11 days shy
of the 50th anniversary of the publication of Watson and Crick's seminal
description of the DNA double helix. If science, technology, and medicine have consistently
demonstrated anything, it is that they proceed at an ever-quickening pace. That we have gone in the past 50
years from the first description of the structure of our DNA to its complete sequencing
gives some indication of how much the impact of genomic medicine on the health care
of today's neonates will increase by the time they turn 50 years of age.
 Pre- Post-
Genomic
genomic genomic
Era
Era Era
>>Pewarisan sifat oleh >> Diketahui adanya >>Pemanfaatan informasi

Genomic Era

Post-Genomic
Pre-Genomic
Gregor Mendell struktur Kromosom oleh genetik untuk kepentingan
Rosalind Franklind umat manusia
>>Diketahui adaya >>Pemanfaatan informasi
informasi genetik berupa genetik untuk mengetahui
susunan nukleotida oleh berbagai macam
Watson & Crick mekanismeyang terjadi
>>Terpecahkannya kode dalam tubuh organisme
genetik oleh Holley, Khorana
& Nirenberg
>> Sequencing (Sanger &
Gilbert)
>> PCR technology (Kary
Mullis)
>>HuGO Project
Genome

Omics
 DNA
Genomic Era

RNA

Protein
DNA
RNA
Protein
Genome

Omics
Omics
 Omics merupakan suatu istilah bagi perkembangan teknologi biologi yang melampaui sel,
jaringan, dan organisme dengan mengintegrasikan berbagai macam platform untuk mengetahui
berbagai fenomena yang terjadi dalam suatu sistem biologi.
 The advantage of the omics study is that they reveal specific results that promote
understanding
 This technology can help to understand the etiology of disease condition through the process
of screening, diagnosis, and prognosis and also for the biomarker discovery to be made easy as
they involve simultaneous investigation of multiple molecules
 Further Omics is of great use in drug discovery and toxicity assessment
 1950s : Rosalind
1865 : Gregor 1950s : Organism
Franklin (DNA
Mendell Classifications (2
structure by X-ray
(Inheritance) domains of Life)
Diffraction)

1968 : Holley, 1977-1980 :

1962 : Watson & Khorana, Maxam & 1983 : Karry
Crick (ATGC) Nirenberg Gilbert Mullis (PCR)
(Genetic Codes) (Sequencing)
OMICs
Genomicss
 Definisi
 an interdisciplinary field of biology focusing on the structure, function, evolution, mapping, and
editing of genomes
 a study of the function and structure of genome, which comprise the complete set of all genes,
regulatory sequences, and non-coding regions within an organism’s DNA
 The study of genomics collectively characterizes, quantifies expression and its associated
regulatory network.

This discipline in genetics relies on sequencing and bioinformatics approach to

sequence, assemble, and analyse all the gene coding and non-coding sequences and how these
genetic components interact to produce an organism and all its functions
Genomics-application

Functional Comparative
Genomics Genomics

Epigenome & Clinical

Epigenetics Genomics
 >> focus on the dynamic regulation of >> enables scientists to identify genes
gene expression and protein-protein and obtain a broader insight into the
interactions, to elucidate DNA function structure–function relationships of
at the levels of genes, transcripts, and genes.
proteins in a genome-wide context >> Comparative in silico analysis
identifies gene expression profiles,
functional genomics

Comparative Genomics
protein–protein interactions, and genetic
and regulatory interactions. These in
turn enable us to obtain a broad insight
into the origin and evolution of cellular
interactions.
>> Genome comparison has enabled
us to generate an evolutionary pattern
for living organisms.
 >> The multitude of chemical >> Genomic approaches have been
entities regulating the expression of instrumental in management of
the genome without affecting the chronic illnesses, such as diabetes
sequence of DNA are known as the and inflammatory bowel disease
constituents of the epigenome,
and the phenomenon associated
Epigenetics & Epigeomics

Clinical Genomics
with it is defined as epigenetics.
>> These chemical components
have the ability to turn expression
of genes on and off for formation of
functionally specialized cells and
are invariably heritable
Transcriptomics - overview
 The presence of mRNA in the sample reflects the abundance level of the corresponding gene.
 Gene expression involves the detection and classification of mRNA mixture in a specific
sample. The goal of gene expression profiling is to differentiate the mRNA mixtures from
different samples. Contrary to genotyping, gene expression categorizes the level of
gene expression. The variation of the transcriptome can be seen over time between cell
types and change according to environmental conditions (Hubank 2004).
 Transcriptome constitutes all transcripts present in a cell including mRNA, miRNA, noncoding
RNAs, and small RNAs.
 Transcriptomics identifies the quantity of RNA and transcriptional structure and quantifies the
differential expression levels of transcripts spatially and temporally during various
developmental stages and under varying physiological conditions.
Transcriptomics - definition

 Analysis of a complete set of transcripts in a cell or transcriptome

structure at a given time
 The study of the RNA, RNA variants, transcriptome complexity,
and gene expression analysis
 Transcriptomics - usage
 Transcriptomics guides us to analyze all RNA transcripts including noncoding
RNAs (ncRNAs) and small RNAs, splicing variants and pattern, transcriptional
units, transcriptional start sites, and posttranscriptional modifications.
 We can analyze the differential expression of gene population under different
conditions and developmental stages.

Transcriptomics is now the first and foremost assay to understand an

organism’s biology where it reveals the information on expression of a gene, its
regulation, and downstream signaling
 Transcriptomics - principles
 Transkriptomik menganalisa :
 Structure and dynamics of the transcriptome
 mRNA, where mRNA is converted to cDNA followed by fragmentation, labeling, hybridization, and
probing

Sequencing is followed by bioinformatic approach for data analysis.

Transcriptomics – technology/devices

Serial/Cap Analysis of Gene

Expression Sequence Tag (EST)
Expression (SAGE)

Microarray RNA-seq
Transcriptomics – technology/devices (lanjutan 2)
 Expression Sequence Tag (EST)
 ESTs are unique sequences pointing to expressed genes in the mapped cDNA clone.
 EST was useful for unknown gene identification but could not quantify expressed genes.

 Serial/Cap Analysis of Gene Expression (SAGE)

 SAGE was invented in 1995 by Dr. Victor Velculescu from John Hopkins.
 SAGE is a powerful technique designed for direct quantitation (digital analysis) of gene
expression and also identifies novel gene expression in a cell population.
 Transcriptomics – technology/devices (lanjutan)
Microaray
 Microarray was developed to monitor multiple gene expressions in a given time simultaneously.
 Scientific ingenuity has led to transition from two-dimensional to three-dimensional microarray followed by suspension bead arrays, which are now
useful in clinical implications.
 Ada 2 jenis Microarray untuk Transkriptomik:
1. Low-density spotted arrays or printing microarrays – picoliter volume of cDNA is required, and control and treated samples are labeled with
different fluorophores and hence can be incorporated in same array. The oligonucleotide probes (50–70 nucleotide in length) or PCR products
are spotted onto the glass slides with spot size ranging from 80 to 150 μm. The information obtained from these arrays is the relative gene
expression between two conditions as absolute quantification cannot be done.
2. In situ (on-chip) synthesized or high-density arrays or gene chips consist of synthetically designed oligonucleotides on glass slides or wafers
with the help of modified photolithographic technology (by Affymetrix) or inkjet technology (Agilent). The probes are usually 20–25 bp long,
and multiple probe sets are used that enhance the sensitivity and specificity of the array compared to low-density array. Hence, a single gene is
assayed by several short oligo probes. The major advantage of these gene chips is absolute measurement of the RNA expression in each sample,
and the main drawback is inability to simultaneously compare two biological samples in a same array (Lowe et al. 2017).
RNA-seq
 RNA sequencing is a novel method applied for mapping and quantifying the transcriptome. RNA sequencing is an advanced technique which
utilizes deep sequencing approach to analyze the transcriptome. The major advantage of RNA-Seq over microarray is the discovery of novel RNA
species.
 Transcriptomics - applications
Disease Profilling
• Microarray and RNA-Seq approach have allowed to compare the differential gene expression in normal and disease
tissue samples from patients.
• The strategy has been widely used in identifying cancer and in immune-related diseases.

Ecology
• transcriptome analyses of gene expression are usedto study the molecular adaptations to environmental challenges
• Transcriptomics has been applied to analyze gene expression and identification of pathways in response to abiotic
and biotic environmental stresses

Evolution
• Transcriptomic technology is now being used to study the cause of phenotypic variation correlating with the
divergence in the population

Gene Function Annotation

• Transcriptomics has been useful in identifying the gene structure, function, and their specific phenotype
 Transcriptomics – applications on medical
sciences

>> Cancer is caused due to genetic changes >> Analysis of molecular signatures at
leading to altered expression of oncogenes or inflammatory sites reveals the type of
tumor suppressor genes. inflammation and help in diagnosis of
>> Microarray has widely been used for infection.
identifying cancer progression, classification >> SAGE technique has identified differential
of tumors, and drug sensitivity or resistance. gene expression in response to IgE or

Immunity and
inflammation
Cancer

>> Microarray-based expression profiling stimulating factors and has determined

identifies cellular changes occurring during several novel genes to be stimulation
transformation of noninvasive to invasive cell responsive.
undergoing metastasis. It also aids in >> RNA-Seq has been potentially useful in
identifying biomarkers for different cancers immune disorders or diseases where it can
and have revealed certain genes responsible dissect two cell populations and identify T-
for chemoresistance cell and B-cell receptor repertoire in patients
>> Transcriptome profiling is also useful in (Byron et al. 2016).
drug discovery whereby drug effects in a cell
could also be monitored
Proteomics
Study of the complete proteins from a source and the techniques implied to study these
proteins and their interactions
There exist two modes of proteomics, one that involves the study of only proteins as analysis of
gene products and the other that is more inclusive and comprises a combination of protein
analysis and genomics/transcriptomics
 >> The quantification of >> large-scale analysis of >> This is a broader
the expression of the protein structures category and involves
proteome followed by >> Structural analysis of several proteomics
comparison of the protein proteins is essential for approaches ranging from
expression among samples determining the protein discovery and identification
that differ by a factor being interactions and for drug- of novel proteins/protein

Structure-based
Protein Expression-

Functions-based
Based Proteomics

proteomics

Proteomics
studied. binding studies. complexes, study and
>> The factor could be a characterization of proteins
>>Structure-based for elucidating protein
disease, a drug treatment proteomics is done mainly
or an environmental effect. signalling, interaction and
through X-ray disease mechanisms
>> The differentially crystallography and NMR
expressed proteins spectroscopy.
identified are highly
valuable as disease-/
condition-specific proteins
or as potential drug targets
or as diagnostic markers.
 >> identifying the structure of protein >> the use of proteomics for
complexes or the proteins present improving gene annotations and
inside the cell or a particular organelle genomics. A parallel and combined
creating a “three-dimensional cell analysis of the proteome and the
map” (Blackstock and Weir 1999). genome accelerates
>> These studies determine where >> the discovery of regulatory
Cell Map Proteomics

Proteogenomics
the proteins are located and their mechanisms such as post-
interactions with other proteins transcriptional and post-translational
modifications (Gupta et al. 2007).
>> This approach helps in analyzing
and comparing multiple genomes
(Gupta et al. 2008).
Metabolomic
the high-throughput study of metabolites which serve as an integral part of
the metabolism.
to provide a comprehensive snapshot of the physiological state of an
organism at a given time state

an interdisciplinary field, and it requires the input and data information from
all other fields of ‘omics’ such as through genomics, through transcriptomics
and partly through proteomics

Its focus is on the study of the metabolic components such as enzymes,

substrates and products
 Metabolomic-Application
Functional
Toxicology Genomics Nutrigenomics
* The metabolic profiling of * The knowledge about the * The combination of
the urine samples or the metabolome would be knowledge of various field
blood samples can be used useful to predict whether a of biology such as
for the determination of the gene was affected by transcriptomics, genomics,
toxicity levels insertion/deletion in the metabolomics and
* This approach can also be genome of an organism proteomics using analytical
used for analysing the * The prediction of the techniques with nutritional
disease conditions function of the unknown principles for humans
associated with the liver gene
and the kidney
 Biomarker Personalized
discovery Medicine
* The biomarkers can be considered as * Individual samples from the metabolites
the metabolites, which can be used for the and their concentrations could be used for
classification of two groups of samples (The disease diagnosis and treatment. The
disease group and the healthy group) response of the individual to certain
* Biological samples from the bile, urine medications can be detected by checking
or seminal fluids are important source of the metabolite concentrations
metabolic information, and this
information can be processed though
metabolomics by metabolic profiling or
fingerprinting for the identification of
biomarkers
The End