a presentation on application of proteomics

1. APPLICATIONS OFTRANSCRIPTOMICS IN MODERN
BIOTECHNOLOGY

2. Presented to Dr.M.naveed
Presented by
Javeria sami
Pakeeza rubab
Rashiqa nosheen
Amara hameed
Laiba shoaib
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3. CONTENTS
1. INTRODUCTION
2. HISTORY
3. SCOPE AND AIMS OF TRANSCRIPTOMICS
4. TYPES OF RNA
5. FUNCTIONS OF RNA
6. APPLICATIONS OF TRANSCRIPTOMICS
7. TECHNIQUIES OF TRANSCRIPTOMICS
8. ETHICAL ISSUES

4. INTRODUCTION
• TRANSCRIPTION
It is the process where from DNA,the single stranded RNA is formed.
• TRANSCRIPTS
The readouts of genes are called transcripts
• TRANSCRIPTOME
A complete set of transcript in a cell and their quantitiy in a specific developmental stages or physical condition.
TRANSCRIPTOMICS
The set of all RNA molecules including mRNA, tRNA and rRNA.& non coding and coding RNA produced
in one population of cell.

5. History
• 1970 libraries of of silkmoth Mrna transcripts were collected and converted to
complementary DNA (Cdna) for storage using reverse transcriptase)
• 1980 ( sanger method was used to sequence random transcripts,producing expressed
sequence tags (ESTs)
• 1990 ( RNA microarray red expressed sequence tag blue and serial/cap analysis of gene
expression 1990s ( ESTs came to prominence as an efficient method to determine the
gene content of an organism without sequencing the entire genome)
• 1991 ( The first attempt at capturing a partial human transcriptome was published
• 2008 ( two human transcriptomes , composed of millions of transcript-derived
sequences covering 16000 genes

6. Aims of transcriptomics
• To determine the transcriptional structure of gene
• To catalogue all species of transcript,including mRNAs, non-coding RNAs and
small RNAs
• In terms of their start sites, 5’ and 3’ ends , splicing patterns and other post
transcriptional modifications
• To quantify the changing expression levels of each transcript during different
conditions

7. RNA
It stands for RIBONUCLEIC ACID it is single
stranded nucleic Acid and plays in a role in
transferring information from DNA to protein
forming system of the cell structure.

8. Structure of RNA

9. TYPES OF RNA
TYPES Abbr functions
1 Messenger RNA mRNA Codes for protein
2 Ribosomal RNA rRNA translation
3 Transfer RNA tRNA translation
4 Small nuclear RNA snRNA Splicing and other functions
5 Small nucleolar RNA Sno RNA Nucleotide modification of RNAs
6 Micro RNA miRNA Gene regulation
7
8
Small interfering RNA
Spliced Leader RNA
RNase P
Gene regulation
TRNA maturation

10. APPLICATIONS IN TRANSCRIPTOMICES
• 1. Agriculture and plant breeding
• 2. Stem cells
• 3. Use in risk assessment of chemicals
• 4. Health n disease
• 5. Expression profiling for disease
• 6. Responses to environment

11. TRANSCRIPTOMICES IN RELATIONTO
PLANT BREEDING & AGRICULTURE
• Microsatellite markers
• Detection
• RNAi

12. • In agricultural crops
• RNA interference technology
• RNAi is an ancident evolutionary
mechanism adopted by plants as a defence
strategy
• It is applicable to plants to acquire new traits
• Small RNA characterization
• it is non-protein coding small RNA
molecules ranging from 20-30 nt that have a
role in development,genome maintaince and
plant responses to environmental stresses

13. • Two major groups
• Micro RNAs : these are about 21 nt and usually have a post transcriptional
regulatory role by directing cleavage of a specific transcript
• Short interfering RNAs : these are usually 24nt long and influence de novo
methylation or other modifications to silence genes
• eQTLs: metabolite,protein and transcript profile can also be directly mapped
onto a segregating population to provide information on loci that control
gene expression level, protein modification
• Association with traits eQTL, pQTL, or mQTL

14. Stem cell relation to transcriptomics

15. EXOSOMES FOR CARDIAC REPAIR
• Transfering stem cells
• Approaches to enhance the efficacy of stem cell therapies
• Cell free components
• Promote cardiac function in the pathological heart
• Variety of RNA closely associated with gene expression
• Variety of cellular processes
• Non –coding RNA

16. Transcriptomics biomarker in safetly and risk
assessment of chemicals
• Markers for exposure to chemicals and therapeutic agents
• Transcriptomic marker
• Mechanism of cell injury,cell death or carcinogenic transformation
• Predictor of heath
• Drug safety assessments are more limited

17. APPLICATIONS OF TRANSCRIPTOMICS IN
HEALTH & DISEASE
• Application of transcriptome in autoimmune disease
• Transcriptome analysis of adrenocortical cells in health and disease
• Single cell transcriptomics to explore the immune system in health and disease

18. Expression profiling for disease
Study of transcriptomics also known as expression profiling.
Gene expression profiling
The identification & characterization of mixture of RNA that is present in specific
sample .

19. Response to environment
• Transcriptome allow identification of genes
• Biotic and abiotic stress
• Novel transcriptional networks

20. APPLICATIONS OFTRANSCRIPTOMICES
IN AUTOIMMUNE DISEASES
• To identify molecular pathways involved in the inflammatory processes
• To investigate the transcriptional changes
• Differential expression of miRNA , a class of small noncoding RNAs that regulate
gene expression
• Deregulated in autoimmune diseases
• Role of miRNA
• Blood transcriptome
• Examples of successful blood transcriptome

21. Techniques in transcriptomics
• These are the following techniques in trasncriptomics:
• Real-time PCR
• Microarray
• Next generation sequencing

22. Real-time PCR
• RealTime PCR is based on the detection of the fluorescence produced by a
reporter molecule which increases, as the reaction proceeds. a technology
used for quantification of DNA sequences amplified in PCR

23. History of real-time PCR
• 1993: First DNA detection method by using of EtBr.
• 1996: useTaqMan detection method instead of EtBR.
• 1996-1997: ABI first introduced the real-time PCR.
• (Since many more instrument manufacture and many more detection
method have been developed

24. Role of real-time PCR
• Detection
• Quantification
• analysis

25. How does real-time PCR work
• To amplify a segment of DNA using PCR, the sample is first heated so the
DNA denatures, or separates into two pieces of single-stranded DNA. Next,
an enzyme called "Taq polymerase" synthesizes - builds - two new strands of
DNA, using the original strands as templates.

26. • On x-axis number of PCR cycle, y-axis show the
fluorescent of amplification.
• During exponential phase the amount of PCR product
approximately double in each cycle.
• As a reaction proceed reaction component are
consumed and ultimately one or more of the
components become limiting.in this point we show that
the reaction become slow and enter the plateau phase.
• Fluorescence at background levels and increase of
fluorescence are not detectable then amplified product
accumulate to yield a detectable fluorescence signal the
cycle number at which occurred is called quantification
cycle (Cq).
• Large amount of template the reaction will have low or
early.
• Small amount of template the reaction will high or late
Cq.

27. Microarray
• A microarray is a laboratory tool used to detect the expression of thousand
of gene at the same time

28. DNA microarray procedure:
• There are following step of microarray:
• Collection sample
• Isolate mRNA
• Hybridization
• Detection the relative intensities of fluorescent under microarray scanner
• Analyzed data

30. NEXT GENERATION SEQUENCING
It is also known as high throughput approaches to DNA
sequencing. It is also called massive parallel
sequencingNext-generation sequencing (NGS), also known
as high-throughput sequencing, is the catch-all term used
to describe a number of different modern sequencing
technologies including: Illumina (Solexa) sequencing.

31. Types of next generation sequencing
• ILLUMINA OR SOLEXA SEQUENCING
• DNA NANOBALL SEQUENCING
• SOLID SEQUENCING

32. ILLUMINA OR SOLEXA SEQUENCING
• The Illumina Solexa sequencing technology
uses sequencing-by-synthesis on an eight-
channel flowcell to produce more than 10
million reads per channel with read lengths up
to 100bp. Individual fragments of a genomic
DNA library are amplified on a flowcell via
bridge-PCR to generate clusters of identical
fragments.

33. DNA NANOBALL SEQUENCING
• It is used to determine the entire
genomic sequence of an
organism.This technology has been
used for multiple genome sequencing
project and is scheduled to be used
for more.

34. Solid sequencing
• Solid (Sequencing by
Oligonucleotide Ligation and
Detection) is a next generation DNA
sequencing technology developed
by Life Technologies and has been
commercially available since 2006.
This next generation technology
generates hundreds of millions to
billions of small sequence reads at
one time

35. ETHICAL ISSUES
• Issues in ownership of an individual’s DNA
• Security and sharing of genomic data
• Increasing use of genetic variation screening both in newborn and adult
• Screening for genetic variations can be harmful increasing in anxiety in
individuals.

36. THE END